Open Access

KEGG orthology-based annotation of the predicted proteome of Acropora digitifera: ZoophyteBase - an open access and searchable database of a coral genome

  • Walter C Dunlap1, 2,
  • Antonio Starcevic4,
  • Damir Baranasic4,
  • Janko Diminic4,
  • Jurica Zucko4,
  • Ranko Gacesa4,
  • Madeleine J H van Oppen1,
  • Daslav Hranueli4,
  • John Cullum5 and
  • Paul F Long2, 3Email author
BMC Genomics201314:509

DOI: 10.1186/1471-2164-14-509

Received: 6 March 2013

Accepted: 15 July 2013

Published: 26 July 2013

Abstract

Background

Contemporary coral reef research has firmly established that a genomic approach is urgently needed to better understand the effects of anthropogenic environmental stress and global climate change on coral holobiont interactions. Here we present KEGG orthology-based annotation of the complete genome sequence of the scleractinian coral Acropora digitifera and provide the first comprehensive view of the genome of a reef-building coral by applying advanced bioinformatics.

Description

Sequences from the KEGG database of protein function were used to construct hidden Markov models. These models were used to search the predicted proteome of A. digitifera to establish complete genomic annotation. The annotated dataset is published in ZoophyteBase, an open access format with different options for searching the data. A particularly useful feature is the ability to use a Google-like search engine that links query words to protein attributes. We present features of the annotation that underpin the molecular structure of key processes of coral physiology that include (1) regulatory proteins of symbiosis, (2) planula and early developmental proteins, (3) neural messengers, receptors and sensory proteins, (4) calcification and Ca2+-signalling proteins, (5) plant-derived proteins, (6) proteins of nitrogen metabolism, (7) DNA repair proteins, (8) stress response proteins, (9) antioxidant and redox-protective proteins, (10) proteins of cellular apoptosis, (11) microbial symbioses and pathogenicity proteins, (12) proteins of viral pathogenicity, (13) toxins and venom, (14) proteins of the chemical defensome and (15) coral epigenetics.

Conclusions

We advocate that providing annotation in an open-access searchable database available to the public domain will give an unprecedented foundation to interrogate the fundamental molecular structure and interactions of coral symbiosis and allow critical questions to be addressed at the genomic level based on combined aspects of evolutionary, developmental, metabolic, and environmental perspectives.

Keywords

Acropora digitifera KEGG orthology Database Annotation Proteome Genome Coral Symbiosis Cnidaria

Background

All of the reef-building corals (Scleractinia; phylum Cnidaria) that create the vast calcium carbonate deposits of coral reefs have evolved an endosymbiotic partnership with photosynthetic dinoflagellates of the genus Symbiodinium (Dinophyceae), commonly known as zooxanthellae, which reside within the gastrodermal cells of their scleractinian host [13]. Coral-algal symbiosis is a cooperative metabolic adaptation necessary for survival in the shallow oligotrophic (nutrient-poor) waters of tropical and subtropical marine environments [4, 5] that drives the productivity of coral reefs [6]. Coral reefs provide habitat and trophic support for many thousands of marine species, the richness of which rival the biological biodiversity of tropical rainforests [7]. Underlying the basic requirements of corals for growth, reproduction and survival are special needs to accommodate symbiont-specific host recognition, to control innate and responsive immune systems, and what is likely to emerge from future research is the extent to which the host is involved in direct regulation of its endosymbiont populations. Much is understood about the cellular biology of cnidarian-dinoflagellate symbiosis (reviewed in [8]), but less is known at the molecular level of coral symbiology. There is little opposition to the contention that environmental and anthropogenic disturbances are causing alarming losses to coral reefs ([9] and reference therein). Threats to productivity are being imposed by the disruption of coral symbiosis (apparent as “coral bleaching”) caused in response to increasing thermal stress attributed to global warming [10, 11], from an increase in stress-related coral disease [1214], from the discharge of domestic and industrial wastes, pollutants from agricultural development and the transport of sediments in terrestrial runoff [15, 16], and potentially from imminent declines in coral calcification owing to rising ocean acidification [1719]. Accordingly, we require a better understanding of the molecular stress responses and adaptive potential of corals. Such information is necessary to predict bleaching events and so better inform effective management policies for the conservation of coral reef ecosystems [2024].

To understand how coral holobionts respond to environmental change at the molecular level, the identification of genes that may respond by transcription to stress is of primary importance [25]. Thus, the use of transcriptomic methodologies to identify stress-responsive genes has been highly successful [2632]. Transcriptome high-throughput profiling has allowed changes in gene expression across thousands of genes to be measured simultaneously. Fuelled by data-generating power, the number of coral based studies utilising transcriptomics to investigate molecular responses to environmental stressors has expanded greatly by the acquisition of expressed sequence tag (EST) gene libraries, the fabrication of microarray biochips used to estimate levels of mRNA expression, and by direct analysis using next-generation, high-throughput sequencing. However, much of this work has been conducted using the aposymbiotic state of pre-settlement coral larvae, so transcribed genes relevant to metamorphosis and the cytobiology of the adult polyp are limited to a few recent studies [3336]. The transcriptome additionally does not provide the structural framework and essential regulatory elements of the functional genome for comprehensive evaluation. Recently, deep metatranscriptomic sequencing of two adult coral holobiomes has been made available on searchable databases: PocilloporaBase for Pocillopora damicornis[36] and PcarnBase for Platygyra carnosus[37]. In contrast, high-throughput metaproteomic analyses to quantify the product yield of stress-response genes of the coral holobiome are yet to be widely adopted by the coral reef scientific community, despite the proteome being the ultimate measure of the coral phenotype [38, 39].

The early accumulation of transcriptomic data revealed that a small proportion of coral ESTs matched genes known previously only from other kingdoms of life, implying that the ancestral animal genome contained many genes traditionally regarded as ‘non-animal’ that have been lost from most animal genomes [40]. Furthermore, an unexpected revelation from EST data is the greater extent to which coral sequences resemble human genes than those of the Drosophila and Caenorhabditis model invertebrate genomes [41, 42]. Comparative genomic analysis has revealed higher genetic divergence and massive gene loss within the ecdysozoan lineages. Hence, many genes assumed to have much later evolutionary origins are likely to have been present in an ancestral or early-diverged metazoan [43]. While much of the animal kingdom remains yet to be explored, examples of the metazoan phylum Cnidaria provide a unique insight into the deep evolutionary origins of at least some vertebrate gene families [42]. Thus, the complete genomic sequence of a coral is likely to reveal many genes previously assumed to be strictly vertebrate innovations. To date, cnidarian genomes have been published for the sea anemone N. vectensis[42] and the hydroid Hydra magnipapillata[44]. Only the coral genome of Acropora digitifera is available without restriction on use of its published sequence [45], but the compiled sequence has not been fully annotated. At the time of this writing, the genome assembly of Acropora millepora has been released to the public domain [46], also without full annotation, but an embargo is imposed on use of this data that is highly restrictive to the progress of further studies. Understanding how genomic variation affects molecular and organismal biology is the ultimate justification of genome sequencing, and annotation is an essential step in this process. We envisage that unrestricted access to annotation of the A. digitifera genome will provide an unprecedented foundation to freely interrogate the generic molecular structure, possible endobiotic interactions and the response of coral to environmental stress. Accordingly, we offer annotation of the predicted proteome of A. digitifera on the open access and searchable database, ZoophyteBase [47]. Use of the ZoophyteBase search engines will allow genes of encoded proteins to be identified that can be examined in context of the cellular physiology, processes of ecological significance, the evolutionary and developmental biology of corals and the functional metabolism of the holobiont that collectively underpin the health of coral reefs.

Construction and content

ZoophyteBase is an open access and searchable database of complete annotation of the predicted proteome of the coral A. digitifera[48]. It was constructed using the MEGGASENSE system, which is a general system for constructing annotation databases with different sorts of input data (DNA reads, assembled genomes, predicted proteomes) and the possibility of using different combinations of analysis tools to create the annotation (Gacesa et al, in preparation). In the case of ZoophyteBase, hidden Markov model (HMM) profiles [49] were chosen as the annotation tool rather than the more common BLAST searches [50]. HMM profiles are constructed from multiple alignments of protein families and contain information about conserved differences in amino acid residues as well as deletions and insertions [49]. This is particularly important for a coral database, as corals are evolutionarily distant to most other organisms. This means that known homologous sequences present in the databases will usually have relatively low similarity, making BLAST searches inaccurate. The statistical information in an HMM profile gives more sensitive and accurate detection of sequence homology. An additional advantage of HMM profiles is that the statistical significance of hits (the expected value) is much more accurate than that calculated by BLAST programs.

The quality of sequence annotation is limited by the accuracy of information provided in any database used. It is well known that there are many problems with annotation in the large uncurated databases such as the NCBI GenBank nr sequences. Widely accepted, the most accurate database for functional annotation is the KEGG database [51]. The KEGG database organises sequences as groups of KEGG orthologues. These are sets of homologous sequences from as wide a range of organisms as possible having an assigned molecular function. These functions are arranged in a hierarchical fashion and grouped in biological pathways. The sequences belonging to KEGG orthologues were used to construct HMM profiles for annotating the coral sequences. Accordingly, the 23,524 predicted proteins encoded in the coral genome were analysed using HMM profiles. If a protein showed a highly significant correlation (“hit”) to a single HMM profile, this was used to create a “trusted” annotation of the sequence. Choosing a cut-off for this criterion is not trivial, because longer sequences tend to have more significant e-values. For construction of ZoophyteBase the criterion 1e-5 was used. This resulted in 19,044 predicted proteins giving “trusted” sequence annotation. For many of these proteins there were two or more highly significant hits to established HMM profiles. In these cases, the most significant correlation was used to construct our “best-fit” annotation file, but other hits can be viewed by the database user so that expert knowledge can be employed to override the automatic annotation function. In 8,004 out of 19,044 predicted proteins which were annotated, more than one annotation was assigned based on non-overlapping regions within the protein which were used to construct the “best-fit” annotation file. We interpreted these as “fusion” events generated by the in silico protein prediction method used, and these proteins were treated as multiple instead of single encoded proteins. Hence, this analysis resulted in the annotation of 33,195 proteins in total, generated from the original 23,524 predicted coral proteins. This is a very conservative annotation scheme, so it can be assumed that most of the annotations are biologically meaningful. Almost 81% (19,044 out of 23,524) of the predicted proteome was assigned using this method.

Utility

The MEGGASENSE system was used to generate a web interface for ZoophyteBase. The home page (Figure 1A) allows the use of several functions. A text version of the entire annotation can be downloaded for manual inspection. There is a proteome overview that gives statistics about the database and a breakdown of the annotated functions into different categories of genes. A particularly useful feature of ZoophyteBase is the ability to use text queries employing a search engine that provides a relevant inquiry in the absence of an exact match between key words of a search and those described for a functional protein. The search engine uses text from the KEGG-database, PubMed and other sources to establish links between query words to access protein data using an intelligent Google-like search engine implemented by the search platform Lucene/Solr [52]. This helps to overcome the common problem that different terminology is used by different groups of researchers. The use of this search function is illustrated by using the query “phagocytosis” (Figure 1B). This inquiry finds 42 hits to KEGG orthologue profiles. One of the hits corresponds to amphiphysin (a synaptic vesicle protein) with annotation of two protein homologues encoded in the coral genome. On the data page there is a brief description of the function of amphiphysin together with a PUBMED literature reference. The sequences of the predicted coral proteins (Figure 1C) can be retrieved, and it is also possible to analyse such data with computer aided drug design methods [53] to extract conserved domains. There are also two tools for the user to examine matches to protein sequences. The user can carry out a BLAST search against the coral protein sequence or analyse the predicted sequence against HMM profiles used to annotate the coral proteome. These tools require only the user to paste their queury into the sequence window.
https://static-content.springer.com/image/art%3A10.1186%2F1471-2164-14-509/MediaObjects/12864_2013_Article_5245_Fig1_HTML.jpg
Figure 1

Graphical overview of the user-web interface for ZoophyteBase during a typical search. The home page allows several search functions (A). Text queries using an intelligent Google-like search engine is illustrated by using the query “phagocytosis” (B). This finds 42 hits to KEGG orthologue profiles. One of the hits corresponds to amphiphysin with annotation of two protein homologues encoded in the coral genome. On the data page there is a brief description of the function of amphiphysin together with a PUBMED literature reference. The sequences of the predicted coral proteins can be retrieved (C).

In this manuscript we demonstrate the utility of ZoophyteBase by presenting predicted gene-encoded proteins revealed by annotation of the A. digitifera genome that have physiological, biological and environmental significance. We discuss features of importance in coral physiology: (1) regulatory proteins of symbiosis, (2) planula and early developmental proteins, (3) neural messengers, receptors and sensory proteins, (4) calcification and Ca2+-signalling proteins, (5) plant-derived proteins, (6) proteins of nitrogen metabolism, (7) DNA repair proteins, (8) stress response proteins, (9) antioxidant and redox-protective proteins, (10) proteins of cellular apoptosis, (11) microbial symbioses and pathogenicity proteins, (12) proteins of viral pathogenicity, (13) toxins and venom, (14) proteins of the chemical defencesome and (15) coral epigenetics.

Discussion

Regulatory proteins of symbiosis

Metabolic cooperation is a key feature of coral-algal symbiosis that allows reef-building corals to inhabit the often nutrient-poor waters of tropical oceans [54]. In this phototropic symbiosis, fixed carbon produced by resident algae is released to the host for nutrition, and the algal symbionts benefit by acquiring the inorganic nutrient wastes of host metabolism [2, 55]. The symbiotic dinoflagellates reside and proliferate within a specialised phagosome (the symbiosome) maintained within host gastrodermal cells. This arrangement requires complex biochemical coordination by the coral at various metabolic stages that includes endocytosis (phagocytosis) by post-settlement polyps to acquire algal symbionts, accord symbiosome recognition to arrest phagosomal maturation for sustained organelle homeostasis, activate symbiophagy or exocytosis to eliminate damaged symbionts [56, 57], and regulate apoptotic or exocytotic pathways to remove excess or impaired populations, all of which have long been recognised as essential to preserve the stability of coral symbiosis [58]. Although these processes are poorly understood in corals, it has been realised from studies of the sea anemone Aiptasia pulchella, a related anthozoan also containing Symbiodinium sp. endosymbionts, that the persistence of algal-containing symbiosomes in Cnidaria relies on the exclusion or retention of small Rab GTPase family proteins that are key regulatory components of vesicular trafficking and membrane fusion in eukaryotic cells [59]. Significantly, ApRab3 and ApRab4 accumulate in the biogenesis of maturing symbiosomes of A. pulchella[60, 61], and mature symbiosomes enveloping healthy dinoflagellates have tethered ApRab5 [62], a checkpoint antagonist of downstream ApRab7 and ApRab11 proteins that would otherwise direct autophagy of the symbiont cargo [63, 64].

Our annotation of the A. digitifera genome reveals sequences encoding putative Rab homologues of the Ras superfamily of proteins (Table 1). In a comparison of cnidarian Rab proteins, eight proteins of A. digitifera matched homologues of Aiptasia pulchella, twenty-nine matched proteins encoded by the aposymbiotic freshwater H. magnipapillata and the aposymbiotic anemone N. vectensis genomes, while seven Rab and Rab-interacting proteins of A. digitifera did not match other cnidarian proteins (Table 2). Significantly, the eight homologues of A. digitifera that matched exclusively Rab proteins of A. pulchella included homologues of the aforementioned ApRab3, ApRab4 and ApRab5 proteins attributed to the maintenance of healthy symbiosomes in Aiptasia, while homologues of the autophagic ApRab7 and ApRab11 proteins are found also in N. vectensis. While Rab GTPase and their effector proteins coordinate consecutive stages of endocytic vesicular transport [65, 66], soluble N-ethylmaleimide-sensitive factor attachment receptor (SNARE) proteins are essential for Rab assembly to complete endosomal fusion of vesicle membranes [67], a process by which Rab proteins impart specificity by binding distinct Rab and SNARE partner proteins prior to membrane fusion [68]. Genes encoding syntaxin-like SNARE proteins have been unambiguously identified [69] from coral EST database libraries constructed from expressed mRNA isolated from various early life stages of Acropora aspera, A. millepora, A. palmata and Orbicella faveolata (= Monastraea faveolata), as well as from the genome of the sea anemone N. vectensis[70]. In metazoans, vacuolar r-SNARE receptor proteins comprise the syntaxin, synaptobrevin and VAMP family proteins, of which there are eight syntaxin and syntaxin-binding proteins (plus two plant-like syntaxins). Additionally, there are one t-SNARE target protein to direct vacuolar morphogenesis, two synaptosomal proteins, one synaptosomal complex ZIP1 protein (yeast homologue), one synaptobrevin membrane protein of secretory vesicles, ten vesicle-associated membrane proteins (VAMPs), a vacuolar protein-8 regulator of autophagy, four vacuolar-sorting proteins and two SEC22 vesicle trafficking protein encoded in the genome of A. digitifera (Table 1), many of which may interact to provide metabolic transport between the endoplasmic reticulum and Golgi apparatus [71]. Included in this vast but yet unexplored repertoire of vacuolar-acting proteins are the syntaxin-binding amisyn and tomosyn regulators of SNARE complex assembly and disassembly [72, 73], which may control membrane fusion in the phagocytic establishment and dis-sociation of coral symbiosis.
Table 1

Regulatory proteins of symbiosis in the predicted proteome of A. digitifera

Gene sequence

KEGG Orthology

Encoded protein description

v1.06849

K06110

Exocyst complex component 3

v1.00063; v1.01826

K06111

Exocyst complex component 4

v1.06336; v1.06337; v1.15354

K07195

Exocyst complex component 7

v1.04340 [+ 4 other sequence copies]

K14966

Host cell factor

v1.01629; v1.19166

K12481

Rabenosyn-5

v1.18447 [+ 26 other sequence copies]

K07976

Rab family, other (similar to Rab-6B)

v1.02380

K12480

Rab GTPase-binding effector protein-1

v1.01032

K13883

Rab-interacting lysosomal protein

v1.14682; v1.03256; v1.07709

K12484

Rab11 family-interacting protein-1/2/5

v1.13055; v1.13176; v1.16348

K12485

Rab11 family-interacting protein-3/4

v1.01275

K07932

Rab-like protein-2B

v1.17629 [+ 13 other sequence copies]

K07933

Rab-like protein-3

v1.03299; v1.09653

K07934

Rab-like protein-4

v1.08498

K07935

Rab-like protein-5

v1.16155 [+5 other sequence copies

K07874

Ras-related protein Rab-1A

v1.09098

K07875

Ras-related protein Rab-1B

v1.13558; v1.08983

K07877

Ras-related protein Rab-2A

v1.14260

K07878

Ras-related protein Rab-2B

v1.07500; v1.20532; v1.07498

K07884

Ras-related protein Rab-3D

v1.21242; v1.07502

K07880

Ras-related protein Rab-4B

v1.01341; v1.05619

K07888

Ras-related protein Rab-5B

v1.07125

K07889

Ras-related protein Rab-5C

v1.09239

K07893

Ras-related protein Rab-6A

v1.10443; v1.13335

K07897

Ras-related protein Rab-7A

v1.03086; v1.17122; v1.07231

K07916

Ras-related protein Rab-7 L1

v1.02275 [+ 4 other sequence copies]

K07901

Ras-related protein Rab-8A

v1.24612

K07899

Ras-related protein Rab-9A

v1.00411

K07900

Ras-related protein Rab-9B

v1.10697; v1.01515

K07903

Ras-related protein Rab-10

v1.22278; v1.04408; v1.12528

K07905

Ras-related protein Rab-11B

v1.07033; v1.23028

K07881

Ras-related protein Rab-14

v1.02275

K07908

Ras-related protein Rab-15

v1.16455; v1.14911; v1.14959

K07910

Ras-related protein Rab-18

v1.04714

K07911

Ras-related protein Rab-20

v1.01878; v1.12184

K07890

Ras-related protein Rab-21

v1.09930

K06234

Ras-related protein Rab-23

v1.13579; v1.12841

K07912

Ras-related protein Rab-24

v1.10183

K07913

Ras-related protein Rab-26

v1.08199

K07885

Ras-related protein Rab-27A

v1.13978; v1.18893

K07917

Ras-related protein Rab-30

v1.03085; v1.06007; v1.07729

K07918

Ras-related protein Rab-32

v1.24721

K07919

Ras-related protein Rab-33A

v1.18892

K07920

Ras-related protein Rab-33B

v1.16060

K07876

Ras-related protein Rab-35

v1.15894

K07922

Ras-related protein Rab-36

v1.03080

K07923

Ras-related protein Rab-38

v1.21391

K07924

Ras-related protein Rab-39A

v1.14786

K07928

Ras-related protein Rab-40

v1.05611 [+ 13 other sequence copies]

K08502

Regulator of vacuolar morphogenesis (t-SNARE domain)

v1.18253

K08520

SEC22 vesicle trafficking protein A/C

v1.15499

K13814

t-SNARE domain-containing protein 1

v1.05749

K08516

Synaptobrevin homologue YKT6

v1.13229

K12768

Synaptonemal complex protein ZIP1

v1.16533; v1.17141

K08508

Synaptosomal-associated protein, 23 kDa

v1.05301

K08509

Synaptosomal-associated protein, 29 kDa

v1.19071

K04560

Syntaxin 1A

v1.04614; v1.22747

K08486

Syntaxin 1B/2/3

v1.16462

K08490

Syntaxin 5

v1.20758; v1.21534

K08498

Syntaxin 6

v1.22836; v1.15499

K08488

Syntaxin 7

v1.01959; v1.24227

K08501

Syntaxin 8

v1.02007; v1.06683; v1.12727

K08491

Syntaxin 17

v1.21308; v1.11830; v1.01582

K08492

Syntaxin 18

v1.22100; v1.09457

K08518

Syntaxin binding protein 5 (tomosyn)

v1.18555

K08519

Syntaxin binding protein 6 (amisyn)

v1.12938

K08500

Syntaxin of plants SYP6

v1.06575

K08506

Syntaxin of plants SYP7

v1.14699

K08507

Unconventional SNARE in the endoplasmic reticulum protein 1

v1.23782 [+ 38 other sequence copies]

K08332

Vacuolar protein 8

v1.15282; v1.24603; v1.01672

K12196

Vacuolar protein-sorting-associated protein 4

v1.17791 [+ 4 other sequence copies]

K12479

Vacuolar protein sorting-associated protein 45

v1.20907

K11664

Vacuolar protein sorting-associated protein 72

v1.15996 [+ 5 other sequence copies]

K12199

Vacuolar protein sorting-associated protein VTA1

v1.15614

K08510

Vesicle-associated membrane protein 1 (synaptobrevin)

v1.13353

K13504

Vesicle-associated membrane protein 2 (synaptobrevin)

v1.12458; v1.07528

K13505

Vesicle-associated membrane protein 3 (cellubrevin)

v1.19735; v1.21831; v1.07186

K08513

Vesicle-associated membrane protein 4 (Golgi transport)

v1.05299

K08514

Vesicle-associated membrane protein 5 (exocytosis)

v1.13557; v1.24610

K08515

Vesicle-associated membrane protein 7 (exocytosis)

v1.12279

K08512

Vesicle-associated membrane protein 8 (endobrevin)

v1.00261; v1.08699; v1.04334

K06096

Vesicle-associated membrane protein A

v1.20177

K10707

Vesicle-associated membrane protein B

v1.15472; v1.03568

K06027

Vesicle-fusing ATPase

v1.11431; v1.10487

K08517

Vesicle transport protein SEC22

v1.06393; v1.13003; v1.08735; v1.04261

K08493

Vesicle transport interaction with t-SNAREs 1

Table 2

Distribution of Rab homologues of Aiptasia puchella , Hydra magnipapillata and Nematostella vectensis in the predicted proteome of A. digitifera

A. digitifera Rab protein

Cnidarian encoding Rab homologue

Rab-like protein- 2B, Rab-2B Rab-3D, Rab-4B, Rab-5B, Rab-26, Rab-32, Rab-38

A. puchella

Rab-like protein-3, Rab-36

N. vectensis

Rab-2A, Rab-23

A. puchella, H. magnipapillata

Rab-like protein-6B, Rab-6A, Rab-7 L1, Rab-10, Rab11B, Rab-30, Rab-33B

A. puchella, N. vectensis

Rab effector protein-1, Rab11-interacting protein-3/4

H. magnipapillata, N. vectensis

Rab-like protein-4, Rab-like protein-5, Rab-1A, Rab5C, Rab-7A, Rab-8A, Rab-9A, Rab-14, Rab-18, Rab-20, Rab-21, Rab-24, Rab-27A, Rab-35

A. puchella, H. magnipapillata, N. vectensis

Rab-interacting lysomal protein, Rab11-interacting protein-1/2/5, Rab-1B, Rab-9B, Rab-3A, Rab-39A, Rab-40

No match

In the final step of exocytosis there is a cytosolic influx of calcium which binds to synaptotagmin to actuate completion of membrane SNARE protein assembly with exocytic docking to form the conducting channel for trans-membrane vesicular transport on activation by vesicle-fusing ATPase [74]. As synaptotagmin proteins are not included in the KEGG database, Zoophytebase was used for BLAST searches with all known synaptotagamin sequences [27]. Synaptotagamin proteins from A. digitifera were found having similarity to homologues from diverse invertebrate and vertebrate organisms, including one from the human genome (Table 3). Other Ca2+-sensing proteins of A. digitifera, such as calmodulin and the calcium binding protein CML, are given with calcification and Ca2+-signalling proteins.
Table 3

Synaptotagmin proteins in the predicted proteome of A. digitifera

Gene sequence

GenBank Accession

Genome encoded homologue

v1.08623

GI:268530614

Caenorhabditis briggsae: XP_002630433 (worm)

v1.20682; v1.10560; v1.02080; v1.10015

GI:150416761

Platynereis dumerilii: ABR68850 (worm)

v1.10269; v1.04412

GI:288869516

Nasonia vitripennis: NP_001165865 (wasp)

v1.01508

GI:29378331

Lymnaea stagnalis: AA093847 (snail)

v1.18613

GI:391339919

Metaseiulus occidentalis: XP_003744294 (mite)

v1.07402

GI:260834895

Branchiostoma floridae: XP_002612445 (lancelet)

v1.01542

GI:149067023

Rattus norvegicus: EDM16756 (rat)

v1.20683

GI:383860584

Megachile rotundata: XP_003705769 (bee)

v1.17688

GI:48529130

Oreochromis niloticus; XP_003452067 (fish)

v1.15777; v1.14902

GI:269785031

Saccoglossus kowalevskii: NP_001161667 (worm)

v1.17175; v1.11521

GI:11559313

Halocynthia roretzi: BAB18864 (ascidian)

v.1.03344; v1.03345

GI:12658419

Manduca sexta; AF331039 (moth)

v1.16152

GI:395729192

Pongo abelii: XP_003780414 (orangutan)

v1.10268

GI:327283049

Anolis carolinensis: XP_003226254 (lizard)

v10.2778

GI:125984480

Drosophila pseudoobscura XP_001356004.1 (fly)

v1.02083; v1.02777

GI:226490194

Schistosoma japonicum: CAX69339.1 (fluke)

v1.04326

GI:167744962

Homo sapiens: 2R83_A (human)

v1.14682; v1.04180

GI:241704658

Ixodes scapularis: XP_002411967 (tick)

Intriguingly, annotation of the A. digitifera genome reveals a host cell factor (K14966), but this is not related to the elusive “host factor” of symbiosis demonstrated to be present in tissue homogenates of corals and other marine invertebrates that harbor Symbiodinium spp. endosymbionts [7577]. Instead, this mammalian transcriptional coactivator host cell factor (HFC-1) is known to mediate the enhancer-promoter assemblies of herpes simplex (HSV) and varicella zoster (VZV) viruses for activation of the latent state for replication [78], such that the coral HCF homologue may have similar relevance as a viral checkpoint transcriptional coactivator of virulence in A. digitifera. HCF-1 expression is coupled also to chromatin modification [79, 80] suggesting that the coral protein homologue may have an additional role in epigenetic reprogramming of the chromatin histone-DNA complex at different stages of development.

Planula and early developmental proteins

In this section we discuss predicted proteins encoded in the A. digitifera genome having functional homology to known proteins specific to early embryonic development, planula larvae function and morphogenesis, which are given in Table 4. Annotation of the coral genome reveals a large set of homeobox proteins involved in the regulation of anatomical development during morphogenesis. The homeobox is a highly conserved DNA sequence (homeodomain) within genes that binds to DNA in a sequence-specific manner [81] often at the promoter region of their target gene to affect transcription in the developing embryo. Amonst these transcriptional regulators, Hox genes are essential to metazoan development as their expressed proteins differentiate embryonic regions along the anterior-posterior axis (the Hox code) and are recognised for their contribution to the evolution of morphological diversity [82]. Hox genes are well characterised in cnidarians and, given their importance in embryonic development, it is not surprising that molecular evidence from the Cnidaria reveal that the genetic origins of Hox genes predate the cnidarian-bilaterian divergence [8385] yet had evolved after divergence of the sponge and eumetazoan lineages [86]. Hox genes of cnidarians are typically located in a conserved genomic collinear cluster, which is apparent also for A. digitifera, whereby the order of the genes on the chromosome is the same as that of gene expression in the developing embryo. Included in our annotation are genes encoding two LIM homeobox proteins and a LIM homeobox transcription factor (Lhx) having conserved roles in neuronal development [87], which in N. vectensis are responsible for the development of neural networks in developing larvae and juvenile polyps [88]. Unlike N. vectensis[89], the coral genome expresses a homeobox BarH-like protein that in vertebrates directs neurogenesis [90]. Distinct from homeodomain proteins, but serving similar functions, are various protein activators, regulators and receptors of cellular morphogenesis. Annotation of the coral genome has revealed multiple sequence alignments to a protein homologue of the dishevelled-associated activator of morphogenesis 1 (Daam1) that initiates cytoskeleton formation via the control of actin assembly. Daam1 was found crucial for gastrulation in Xenopus[91], wherein Daam1 mutants of Drosophilia exhibit trachea defects [92], and in mammals Daam1 is highly expressed in multiple developing organs and is deemed essential for cardiac morphogenesis [93]. Similar morphogenetic genes express regulatory proteins that are necessary for vacuole biogenesis in yeasts [94]. Others express bone morphogenetic proteins (and their BMP receptors), which are potent multi-functional growth activators that belong to the transforming growth factor beta (TGFbeta) cytokine superfamily of proteins that in humans have various functions during embryogenesis, skeletal formation, neurogenesis and haematopoiesis [95]. However, since many of the homeobox and morgenetic proteins (Table 4) are homologues of proteins with functions ascribed to higher organisms, their precise function in A. digitifera cannot be ascertained by KEGG orthology alone.
Table 4

Planula and early developmental proteins in the predicted proteome of A. digitifera

Gene sequence

KEGG Orthology

Encoded protein description

v1.09797; v1.11180; v1.08414

K03776

Aerotaxis receptor (oxygen sensing)

v1.07838 [+5 other sequence copies]

K07822

Archaeal flagellar protein FlaC

v1.14039; v1.11310; v1.11309

K05502

Bone morphogenetic protein 1

v1.01025; v1.17008; v1.15796; v1.23658

K04662

Bone morphogenetic protein 2/4

v1.02299; v1.07696; v1.10675

K04663

Bone morphogenetic protein 5/6/7/8

v1.06335; v1.01763

K04673

Bone morphogenetic protein receptor type-1A

v1.13481

K13578

Bone morphogenetic protein receptor type-1B

v1.10550 [+4 other sequence copies]

K04671

Bone morphogenetic protein receptor type-2

v1.00912 [+4 other sequence copies]

K13579

Bone morphogenetic protein receptor type-1, invertebrate

v1.19370

K14624

C-C motif chemokine 2

v1.23163

K12499

C-C motif chemokine 5

v1.08576

K05511

C-C motif chemokine 15/23

v1.09229

K05512

C-C motif chemokine 19/21

v1.09305

K08373

C-C chemokine receptor-like 2

v1.04942

K04179

C-C chemokine receptor type 4

v1.02658

K04245

Chemokine-like receptor 1

v1.21300

K12671

C-X-C motif chemokine 10

v1.16396; v1.21991

K10035

C-X-C motif chemokine 16

v1.23712

K11522

Chemotaxis family two-component system response regulator PixG

v1.09435

K13490

Chemotaxis family, histidine kinase sensor response regulator (WspE-like)

v1.14142; v1.05300

K05874

Chemotaxis protein I, serine sensor receptor (MCP family)

v1.07361

K05877

Chemotaxis protein IV, peptide sensor receptor (MCP family)

v1.17411

K03414

Chemotaxis protein CheZ

v1.16104

K00575

Chemotaxis protein methyltransferase CheR

v1.15537 [+ 7 other sequence copies]

K08482

Circadian clock protein KaiC

v1.14925 [+ 4 other sequence copies]

K02223

Circadian locomoter output cycles kaput protein

v1.06432 [+ 9 other sequence copies]

K04512

Dishevelled associated activator of morphogenesis

v1.17637 [+ 70 other sequence copies]

K10408

Dynein heavy chain, axonemal

v1.00202 [+5 other sequence copies]

K10409

Dynein intermediate chain 1, axonemal

v1.04986; v1.09649; v1.23645

K11143

Dynein intermediate chain 2, axonemal

v1.08695; v1.09481; v1.23153

K10411

Dynein light chain 1, axonemal

v1.11684

K10412

Dynein light chain 4, axonemal

v1.23322; v1.01131; v1.04207

K10410

Dynein light intermediate chain, axonemal

v1.14083

K02401

Flagellar biosynthetic protein FlhB

v1.16997

K02420

Flagellar biosynthetic protein FliQ

v1.02867

K02396

Flagellar hook-associated protein 1 FlgK

v1.18101; v1.13427

K02408

Flagellar hook-basal body complex protein FliE

v1.04339; v1.07633

K06603

Flagellar protein FlaG

v1.17895[+5 other sequence copies]

K02383

Flagellar protein FlbB

v1.21111

K02413

Flagellar protein FliJ

v1.17651 [+ 13 other sequence copies]

K02415

Flagellar protein FliL

v1.01971 [+ 6 other sequence copies]

K02418

Flagellar protein FliO/FliZ

v1.14031

K02423

Flagellar protein FliT

v1.08025

K02394

Flagellar P-ring protein precursor FlgI

v1.02396; v1.15777

K02409

Flagellar M-ring protein FliF

v1.20693

K09451

Homeobox protein aristaless-like 4

v1.24732 [+5 other sequence copies]

K09452

Homeobox protein aristaless-related

v1.15788; v1.19334; v1.04164

K09313

Homeobox protein cut-like

v1.01801

K09319

Homeobox protein engrailed

v1.16835; v1.06323

K09320

Homeobox even-skipped homologue protein

v1.0412; v1.054771

K09354

Homeobox protein expressed in ES cells 1

v1.13604

K09324

Homeobox protein goosecoid

v1.06346; v1.08163

K09325

Homeobox protein goosecoid-like

v1.17295; v1.17294

K09361

Homeobox protein, BarH-like (vertebrate neurogenesis)

v1.07457

K09316

Homeobox protein DLX, invertebrate

v1.11157; v1.08573; v1.15250

K09317

Homeobox protein EMX

v1.01800

K09321

Homeobox protein GBX

v1.10929; v1.06346; v1.05443; v1.07458

K09310

Homeobox protein GSH

v1.13684; v1.24444

K08025

Homeobox protein HB9

v1.16254; v1.16064

K08024

Homeobox protein HEX

v1.07458; v1.06706; v1.06705

K09339

Homeobox protein HLX1

v1.06347; v1.06348; v1.17294

K09302

Homeobox protein HoxA/B2

v1.06125

K09306

Homeobox protein HoxA/B/C6

v1.19818

K09304

Homeobox protein HoxA/B/C/D4

v1.06706

K09301

Homeobox protein HoxA/B/D1

v1.02056

K09353

Homeobox protein LBX

v1.06347; v1.06348

K09328

Homeobox protein Unc-4

v1.24342; v1.04552

K09318

Homeobox protein ventral anterior

v1.03823; v1.10070; v1.04435

K09309

Homeobox protein Nkx-1

v1.12852 [+ 4 other sequence copies]

K08029

Homeobox protein Nkx-2.2

v1.21630

K09345

Homeobox protein Nkx-2.5

v1.10625

K09347

Homeobox protein Nkx-2.8

v1.10625; v1.13865; v1.05476

K09348

Homeobox protein Nkx-3.1

v1.21628; v1.05475; v1.05477

K09995

Homeobox protein Nkx-3.2

v1.06135; v1.10071

K09349

Homeobox protein Nkx-5

v1.14702

K08030

Homeobox protein Nkx-6.1

v1.14917; v1.11907

K09350

Homeobox protein Nkx-6.2

v1.00777; v1.21453

K09322

Homeobox protein MOX

v1.00602 [+ 6 other sequence copies]

K09326

Homeobox protein OTX

v1.16722; v1.12785

K09374

LIM homeobox protein 3/4

v1.11281; v1.05135

K09375

LIM homeobox protein 6/8

v1.07988; v1.22037

K09371

LIM homeobox transcription factor 1

v1.09328 [+ 5 other sequence copies]

K10394

Kinesin family member 3/17

v1.09196; v1.12479

K11525

Methyl-accepting chemotaxis protein PixJ (MCP family)

v1.17028; v1.13473

K08473

Nematode chemoreceptor

v1.13159; v1.00655

K09330

Paired mesoderm homeobox protein 2

v1.15178; v1.10962; v1.16587; v1.01557

K02633

Period circadian protein

v1.23288; v1.13857

K04627

Pheromone a factor receptor

v1.22464; v1.17135

K11213

Pheromone alpha factor receptor

v1.05611 [+ 13 other sequence copies]

K08502

Regulator of vacuolar morphogenesis

v1.04431

K09333

Retina and anterior neural fold homeobox-like protein

v1.17636

K09331

Short stature homeobox protein

v1.14704

K09340

T-cell leukemia homeobox protein

v1.11765

NA 1

Tektin

v1.04154

K02669

Twitching motility protein PilT

1 NA KEGG orthology designation not assigned.

Another protein encoded in the A. digitifera genome is a retina and anterior neural fold homeobox-like (RAX) protein that may activate the development of primitive coral photoreceptors [96, 97], including a blue light-sensing, cryptochrome photoreceptor that in A. millepora is implicated in the detection of light from the lunar cycle of night time illumination to signal synchronous coral spawning [98, 99]. Photosensitive behaviours and the circadian rhythms of corals are well described, and diurnal cycles of gene transcription that regulate circadian biological processes in the coral A. millepora have been reported [100]. Such traits in A. millepora appear regulated by an endogenous biological clock entrained to daily cycles of solar illumination [101]. Annotation of the A. digitifera genome reveals a circadian timekeeper protein KaiC [102] that in cyanobacteria is activated during the diurnal phosphorylation rhythm [103, 104]. In Synechococcus elongatus, KaiC regulates the rhythmic expression of all other proteins encoded in the genome [105], yet no homologue of any of the prokaryotic clustered circadian kiaABC genes has been identified in eukaryotes [106]. In Drosophila, KaiC together with a homologue of the eukaryotic period (Per) circadian protein drives circadian rhythms in eclosion (hatching) and locomotor activity [107]. Nevertheless, a circadian locomotor output cycles kaput (CLOCK) homologue (Table 4) was found in our annotation. Since CLOCK proteins serve as an essential activator of downstream elements in pathways critical to the regulation of circadian rhythms in eukaryotes [108], it would be worthy to examine how transcription of the RAX-like homeobox protein in this coral contributes to the development of circadian functions by activation of kaiC, per and Clock genes. Such a study might reveal that components of the animal circadian clock are more ancient than data previously suggested [109].

Broadcast-spawning corals, such as A. digitifera, release gametes, and the fertilised eggs develop into planula larvae within the water column until they have reached settlement competency, find a suitable hard substrate, attach and develop into the polyp on metamorphosis. Coral sperm and planula larvae achieve motility using flagella (sperm) or cilia (larvae) as their locomotor organelles. The eukaryotic axonemal proteins of cilia and flagella are composed of a dynein ATPase protein to provide mechanochemical energy transduction together with the principle structural proteins of the ciliary/flagellar microtubules [110]. The flagellar/ciliary microtubules consist of filaments composed of α- and β-tubulins, microtubule-stabilising tektins and kinesin motor proteins [111113]. The coral genome encodes members of the dynein axonemal (flagella and cilia) proteins (Table 4) and many of the dynein cytoplasmic proteins (not tabulated), the latter being involved in intracellular organelle transport and centrosome assembly. The coral genome encodes α- and β-tubulins and members of the eukaryotic kinesin superfamily proteins (not tabulated). Amongst the many kinesin proteins encoded in the coral genome is the kinesin family member 3/17 protein, which is a direct homologue of the kinesin-II intraflagellar transport protein FLA10 essential for flagella assembly in the alga Chlamydomonas[114]. The microtubule-stabilising tektin protein, which is required for cilia and flagella assembly [113], is also encoded in the coral genome [note: there is no KEGG orthology profile assigned to this protein]. It was a surprise, however, to find a large complement of prokaryotic flagellar proteins encoded in the coral genome consisting of archaeal flagellar (FlaC and FlaG), bacterial filament (FibB, FlliE, FliF, FliJ, FliK, FliO/FliZ, FliQ and FliT) homologue components (Table 4). Included also are the prokaryote homologues FlgN and FlbB that regulate transcriptional activation of flagellar assembly [115, 116] and FlhB which controls the substrate specificity of the entire prokaryotic flagellar apparatus [117]. Encoded in the coral genome is a flagella-independent Type IV twitching mobility protein PilT that affords social gliding translocation in many prokaryotic organisms controlled by complex signal transduction systems that include two-component sensor regulators [118]. It is unlikely that these genes are derived from contamination from bacterial DNA. Such contamination would manifest itself by the random occurrence of bacterial genes from the whole genome including many housekeeping genes. In this case, the genes occur as members of groups with specialised functions, suggesting that multiple horizontal gene transfers between bacteria and the coral genome have occurred [119]. Their precise function in A. digitifera remains unknown; homologues of these prokaryotic genes have not been described previously in any other eukaryote genome.

Linked closely with flagellar/ciliary proteins are the sensory receptors that signal chemoattraction or avoidance to direct cellular motility. The coral genome reveals a variety of genes that encode chemoreceptor and chemotaxis proteins (Table 4). The chemoreceptor proteins of A. digitifera include an oxygen-sensing aerotaxis receptor that in bacteria invokes an avoidance response to anoxic micro-environments [120]. Encoded also are a nematode sensory chemoreceptor homologue [121], two homologous pheromone factor receptor proteins that in fungi activate a species-specific mating response [122], three chemotaxis protein sensor receptors belonging to the methyl-accepting chemotaxis family of proteins (MCPs) in bacteria and archaea [123], and two proteins (CheZ and CheR) and two regulators (PixG and WspE) of the two-component signal transduction (TCST) system for activation of gene expression. In bacteria and archea, as well as some plants, fungi and protozoa [124], TCST systems mediate many cellular processes that respond to a broad range of environmental stimuli via activation of a specific histidine (or serine) kinase sensor and its cognate response regulator [125]. There are 77 sequence matches to various elements of the TCST family of proteins in the A digitifera genome (data not tabulated). Included also are genes encoding members of the chemotactic cytokine (chemokine) family of sensory proteins that on secretion directs chemotaxis in nearby responsive cells by stimulating target chemokine receptors; both chemokine and chemokine receptor proteins are encoded in the coral genome. Significantly, sensory chemokines/chemokine receptors are found in all vertebrates, some viruses and some groups of bacteria, but none have been described previously for invertebrates [126].

Neural messengers, receptors and sensory proteins

Corals and other cnidarians are the earliest extant group of organisms to have a primitive nervous system network [127] thought to be evolved from a eumetazoan ancestor prior to the divergence of Cnidaria and the Bilateria [128, 129]. Unlike marine sponges (Porifera) that predate synaptic innovation [130], cnidarians possess a homogenous nerve net that, although lacking any form of cephalization, accommodates fundamental neurosensory transmission across the nerve net to end in a motoneural junction to coordinate tentacle movement required for feeding and predator avoidance [131]. The nervous systems of cnidarians consist of both ectodermal sensory cells and their effector cells and endodermal multipolar ganglions capable of neurotransmission [132]. At the functional level, synaptic transmission in cnidarians relies on fast neurotransmitters (glutamate, GABA, glycine) and slow neurotransmitters (catecholamine, serotonin, neuropeptides) for sensory-signal conduction [133]. At the ultrastructural level, many cnidarian neurons have multifunctional traits of sensory, neurosecretory and stimulatory attributes [134]. Significantly, the genome of A. digitifera encodes the expression of a ciliary neurotrophic factor, which is a polypeptide hormone and nerve growth factor that promotes neurotransmitter synthesis, neurite outgrowth and regeneration [135]. Additionally, the coral genome encodes nerve growth factor and neurotrophic kinase receptors, a survival motor neuron protein, a survival neuron splicing factor, the neural outgrowth protein neurotrimin, and a neurotrophin growth factor attributed to signalling neuron survival, differentiation and growth (Table 5). Encoded for neuron regulation and development are several neuron cation-gated channels, a neuronal guanine nucleotide exchange factor, a neurotransmitter Na+ symporter, several neurogenic differentiation proteins, a neuronal PAS domain transcription factor for activation of neurogenesis, the axon guidance protein neurophilin-2, a neural crest protein of embryonic neural development, neural ELAV-like transcription proteins of neurogenesis, a Notch protein (79 sequence domain matches) and a neutralized protein subset of the Notch signalling pathway that promotes neuron proliferation in early neurogenic development. Structural elements of the coral nerve net include neurofilament polypeptides and neuronal adhesion proteins.
Table 5

Neuronal and sensory proteins in the predicted proteome of A. digitifera

Gene sequence

KEGG Orthology

Encoded protein description

v1.01918 [+ 5 other sequence copies]

K01049

Acetylcholinesterase

v1.18087; v1.14516

K04136

Adrenergic receptor alpha-1B

v1.06394

K04137

Adrenergic receptor alpha-1D

v1.09628; v1.15688; v1.00966

K04140

Adrenergic receptor alpha-2C

v1.19831; v1.20450

K04142

Adrenergic receptor beta-2

v.17293

K00910

beta-Adrenergic-receptor kinase

v1.13740 [+ 5 other sequence copies]

K04828

Amiloride-sensitive cation channel 1, neuronal (degenerin)

v1.23541 [+ 6 other sequence copies]

K04829

Amiloride-sensitive cation channel 2, neuronal

v1.09323 [+ 4 other sequence copies]

K04439

beta-Arrestin

v1.07723; v1.22465

K04641

Bacteriorhodopsin

v1.08062

K05420

Ciliary neurotrophic factor

v1.03288 [+ 5 other sequence copies]

K02295

Cryptochrome

v1.20011; v1.20036; v1.20084; v1.18607

K04948

Cyclic nucleotide gated channel alpha 1

v1.21470

K04951

Cyclic nucleotide gated channel alpha 4

v1.21783; v1.01466; v1.01466; v1.01466

K05326

Cyclic nucleotide gated channel, invertebrate

v1.03645

K05391

Cyclic nucleotide gated channel, other eukaryote

v1.21256

K08762

Diazepam-binding inhibitor (GABA receptor, acyl-CoA-binding protein)

v1.22156 [+ 6 other sequence copies]

K00503

Dopamine beta-monooxygenase

v1.21775: v1.15989

K04148

Dopamine D1-like receptor

v1.14160; v1.01697

K04144

Dopamine receptor D1

v1.05089; v1.20018

K04145

Dopamine receptor D2

v1.14030; v1.23273

K04146

Dopamine receptor D3

v1.20536

K13088

ELAV-like protein 1

v1.18658 [+ 5 other sequence copies]

K13208

ELAV-like protein 2/3/4

v1.05774 [+ 18 other sequence copies]

K04313

G protein-coupled receptor 6

v1.00572; v1.18152

K08404

G protein-coupled receptor 17

v1.23842

K04316

G protein-coupled receptor 19

v1.03948

K08411

G protein-coupled receptor 26

v1.09271

K08383

G protein-coupled receptor 34

v1.05595

K04243

G protein-coupled receptor 37 (endothelin receptor type B-like)

v1.04019

K08409

G protein-coupled receptor 45

v1.19913; v1.09821; v1.04291

K08450

G protein-coupled receptor 56

v1.05404

K04321

G protein-coupled receptor 63

v1.02179; v1.10397

K08451

G protein-coupled receptor 64

v1.23269 [+ 5 other sequence copies]

K08408

G protein-coupled receptor 68

v1.21091

K08421

G protein-coupled receptor 84

v1.11008

K04302

G protein-coupled receptor 85

v1.21884; v1.01951

K08452

G protein-coupled receptor 97

v1.03243 [+ 13 other sequence copies]

K08378

G protein-coupled receptor 103

v1.13790; v1.18939

K08453

G protein-coupled receptor 110

v1.09442; v1.14019

K08455

G protein-coupled receptor 112

v1.24009

K08456

G protein-coupled receptor 113

v1.04290

K08459

G protein-coupled receptor 114

v1.06608; v1.24223

K08457

G protein-coupled receptor 115

v1.10800 [+ 6 other sequence copies]

K08458

G protein-coupled receptor 116

v1.07662 [+ 6 other sequence copies]

K08462

G protein-coupled receptor 125

v1.09663; v1.08981

K08463

G protein-coupled receptor 126

v1.24252

K08464

G protein-coupled receptor 128

v1.02750 [+ 26 other sequence copies]

K08465

G protein-coupled receptor 133

v1.05774 [+ 11 other sequence copies]

K08466

G protein-coupled receptor 144

v1.05497; v1.13272; v1.01323

K08436

G protein-coupled receptor 152

v1.08653 [+ 5 other sequence copies]

K08467

G protein-coupled receptor 157

v1.11807; v1.10392; v1.10394

K08469

G protein-coupled receptor 158

v1.07294; v1.00247

K08439

G protein-coupled receptor 161

v1.05167

K08442

G protein-coupled receptor 176

v1.08677; v1.23465; v1.19865; v1.06986

K12762

G protein-coupled receptor GPR1

v1.13395

K08291

G protein-coupled receptor kinase

v1.18529; v1.07599; v1.05558

K12487

G protein-coupled receptor kinase interactor 2

v1.02481

K04619

G protein-coupled receptor family C group 5 member B

v1.22242

K04622

G protein-coupled receptor family C group 6 member A

v1.08625; v1.13650; v1.13048; v1.18694

K04599

G protein-coupled receptor Mth (Methuselah protein)

v1.07465; v1.10540

K08341

GABA(A) receptor-associated protein (autophagy-related protein 8)

v1.09831 [+ 30 other sequence copies]

K05270

Gamma-aminobutyric acid (GABA) receptor, invertebrate

v1.18702; v1.11701

K05183

Gamma-aminobutyric acid (GABA) A receptor beta-3

v1.04252 [+ 6 other sequence copies]

K05185

Gamma-aminobutyric acid (GABA) A receptor epsilon

v1.06325

K05186

Gamma-aminobutyric acid (GABA) A receptor gamma-1

v1.00048

K05188

Gamma-aminobutyric acid (GABA) A receptor gamma-3

v1.07506 [+ 6 other sequence copies]

K04615

Gamma-aminobutyric acid (GABA) B receptor 1

v1.07506 [+ 24 other sequence copies]

K04616

Gamma-aminobutyric acid (GABA) B receptor 2

v1.06426; v1.10563; v1.01138

K05192

Gamma-aminobutyric acid (GABA) receptor theta

v1.15485

K05198

Glutamate receptor, ionotropic, AMPA 2

v1.09807

K05200

Glutamate receptor, ionotropic, AMPA 4

v1.04764

K05207

Glutamate receptor, ionotropic, delta 2

v1.15247 [+ 12 other sequence copies]

K05313

Glutamate receptor, ionotropic, invertebrate

v1.15247 [+ 7 other sequence copies]

K05202

Glutamate receptor, ionotropic, kainate 2

v1.00617

K05203

Glutamate receptor, ionotropic, kainate 3

v1.09688 [+ 6 other sequence copies]

K05208

Glutamate receptor, ionotropic, N-methyl D-aspartate 1

v1.21204 [+ 4 other sequence copies]

K05212

Glutamate receptor, ionotropic, N-methyl-D-aspartate 2D

v1.01622

K05214

Glutamate receptor, ionotropic, N-methyl-D-aspartate 3B

v1.01418 [+ 5 other sequence copies]

K05387

Glutamate receptor, ionotropic, other eukaryote

v1.04275

K05194

Glycine receptor alpha-2

v1.10737; v1.06885

K05195

Glycine receptor alpha-3

v1.05488

K05271

Glycine receptor alpha-4

v1.08900; v1.06885

K05196

Glycine receptor beta

v1.18634

K05397

Glycine receptor, invertebrate

v1.14569; v1.14570

K09071

Heart-and neural crest derivatives-expressed protein

v1.16783 [+ 4 other sequence copies]

K02168

High-affinity choline transport protein

v1.13837

K07608

Internexin neuronal intermediate filament protein, alpha

v1.01671

K04309

Leucine-rich repeat-containing G protein-coupled receptor 4

v1.09480; v1.05605

K04308

Leucine-rich repeat-containing G protein-coupled receptor 5

v1.15300 [+ 8 other sequence copies]

K08399

Leucine-rich repeat-containing G protein-coupled receptor 6

v1.17524 [+ 14 other sequence copies]

K04306

Leucine-rich repeat-containing G protein-coupled receptor 7

v1.21700; v1.03578; v1.17196

K04307

Leucine-rich repeat-containing G protein-coupled receptor 8

v1.16104

K08396

Mas-related G protein-coupled receptor member X

v1.08718; v1.02042; v1.02042

K04604

Metabotropic glutamate receptor 1/5

v1.22794 [+ 7 other sequence copies]

K04605

Metabotropic glutamate receptor 2/3

v1.15331

K04607

Metabotropic glutamate receptor 4

v1.01418

K04608

Metabotropic glutamate receptor 6/7/8

v1.21698; v1.04544; v1.21739

K14636

MFS transporter, solute carrier family 18 (acetylcholine transporter) 3

v1.05751; v1.19720; v1.22165; v1.02336

K04134

Muscarinic acetylcholine receptor

v1.11550

K04129

Muscarinic acetylcholine receptor M1

v1.01913 [+ 4 other sequence copies]

K04131

Muscarinic acetylcholine receptor M3

v1.18723

K04132

Muscarinic acetylcholine receptor M4

v1.08171

K04133

Muscarinic acetylcholine receptor M5

v1.07408 [+ 34 other sequence copies]

K02583

Nerve growth factor receptor (TNFR superfamily member 16)

v1.15265 [+ 91 other sequence copies]

K06491

Neural cell adhesion molecule

v1.13789; v1.24010; v1.03980

K09038

Neural retina-specific leucine zipper protein

v1.24586; v1.16386; v1.16387

K08052

Neurofibromin 1

v1.05520; v1.15407; v1.07950

K04572

Neurofilament light polypeptide

v1.19724

K04573

Neurofilament medium polypeptide (neurofilament 3)

v1.15787 [+ 4 other sequence copies]

K09081

Neurogenin 1 (neurogenic differentiation protein)

v1.00345; v1.05338; v1.10997

K08033

Neurogenic differentiation factor 1

v1.07355; v1.14517

K09078

Neurogenic differentiation factor 2

v1.08832

K09079

Neurogenic differentiation factor 4

v1.06678; v1.06677

K01393

Neurolysin

v1.16238 [+ 19 other sequence copies]

K06756

Neuronal cell adhesion molecule

v1.20460; v1.16967

K06757

Neurofascin NFASC (cell adhesion molecule CAMs)

v1.22060; v1.03561

K07525

Neuronal guanine nucleotide exchange factor

v1.03908

K09098

Neuronal PAS domain-containing protein 1/3

v1.00089

K05247

Neuropeptide FF-amide peptide

v1.21565

K08375

Neuropeptide FF receptor 2

v1.06392 [+ 11 other sequence copies]

K04209

Neuropeptide Y receptor, invertebrate

v1.08609 [+ 31 other sequence copies]

K06819

Neuropilin 2

v1.11492 [+ 5 other sequence copies]

K03308

Neurotransmitter:Na+ symporter, NSS family

v1.16744 [+ 8 other sequence copies]

K06774

Neurotrimin

v1.05353

K03176

Neurotrophic tyrosine kinase receptor type 1

v1.20055

K04360

Neurotrophic tyrosine kinase receptor type 2

v1.03803

K04356

Neurotrophin 3

v1.09523

K04803

Nicotinic acetylcholine receptor alpha-1 (muscle)

v1.11940

K04806

Nicotinic acetylcholine receptor alpha-4

v1.01548

K04808

Nicotinic acetylcholine receptor alpha-6

v1.05056; v1.12097

K04809

Nicotinic acetylcholine receptor alpha-7

v1.07222; v1.11069

K04810

Nicotinic acetylcholine receptor alpha-9

v1.18231 [+ 32 other sequence copies]

K05312

Nicotinic acetylcholine receptor, invertebrate

v1.24404

K04813

Nicotinic acetylcholine receptor beta-2 (neuronal)

v1.06514; v1.23640

K04815

Nicotinic acetylcholine receptor beta-4

v1.18634

K04816

Nicotinic acetylcholine receptor delta

v1.18231 [+ 32 other sequence copies]

K05312

Nicotinic acetylcholine receptor, invertebrate

v1.05293 [+ 78 other sequence copies]

K02599

Notch protein

v1.15348 [+ 4 other sequence copies]

K04256

c-Opsin protein

v1.01972

K08385

G0-Opsin protein

v1.13345 [+ 5 other sequence copies]

K04255

r-Opsin protein

v1.00749; v1.03435

K00504

Peptidylglycine monooxygenase

v1.12323 [+ 11 other sequence copies]

K00678

Phosphatidylcholine-retinol O-acyltransferase

v1.18340 [+ 6 other sequence copies]

K09624

Protease, serine, 12 (neurotrypsin, motopsin)

v1.08030 [+ 9 other sequence copies]

K01931

Protein neuralized

v1.04431

K09333

Retina and anterior neural fold homeobox-like protein

v1.01789; v1.06542

K00061

Retinol dehydrogenase

v1.05804 [+ 6 other sequence copies]

K11150

Retinol dehydrogenase 8

v1.22340; v1.14029

K11151

Retinol dehydrogenase 10

v1.24399; v1.07017

K11154

Retinol dehydrogenase 16

v1.19667; v1.16885; v1.24371

K00909

Rhodopsin kinase

v1.12432; v1.15302; v1.07505

K09516

all-trans-Retinol 13,14-reductase

v1.09104 [+ 6 other sequence copies]

K05613

Solute carrier family 1 (glial high affinity glutamate transporter), member 2

v1.19779; v1.08769; v1.22032

K05617

Solute carrier family 1 (high affinity Asp/glutamate transporter), member 6

v1.19293; v1.19292

K14387

Solute carrier family 5 (high affinity choline transporter), member 7

v1.10901; v1.19493

K05336

Solute carrier family 6 (neurotransmitter transporter), invertebrate

v1.24615 [+ 10 other sequence copies]

K05034

Solute carrier family 6 (neurotransmitter transporter, GABA) member 1

v1.07932

K05046

Solute carrier family 6 (neurotransmitter transporter, GABA) member 13

v1.01817

K05036

Solute carrier family 6 (neurotransmitter transporter, dopamine) member 3

v1.20691; v1.16333; v1.15484; v1.02123

K05038

Solute carrier family 6 (neurotransmitter transporter, glycine) member 5

v1.15484; v1.15484

K05042

Solute carrier family 6 (neurotransmitter transporter, glycine) member 9

v1.18461; v1.09068; v1.02237; v1.20880

K05333

Solute carrier family 6 (neurotransmitter transporter) member 18

v1.02239; v1.13836; v1.09067

K05334

Solute carrier family 6 (neurotransmitter transporter) member19

v1.21997 [+ 5 other sequence copies]

K12839

Survival of motor neuron-related-splicing factor member 30

v1.21997 [+ 6 other sequence copies]

K13129

Survival motor neuron protein

Cnidarians differentiate highly specialised sensory and mechanoreceptor cells involved in the capture of prey and for defence against predators. Their stinging cells, termed nematocysts or cnidocytes, are stimulated by adjacent chemosensory cells. Nematocysts trigger the release of a stinging barb (cnidae tubule) via ultra-fast exocytosis on physical contact with ciliary mechanoreceptors of the cnidocyte to deliver the discharge of its venom [136]. Despite considerable advances in the sensory biology of cnidarians, knowledge of the specific receptor genes that regulate cnidocyte function remains incomplete. In Hydra, and perhaps other cnidarians, cnidocyte discharge is controlled by an ancient light-activated, opsin-mediated phototransduction pathway [137] that precedes the evolution of cubozoan (box jellyfish) eyes [138]; cubozoans are the most basal of animals to have eyes containing a lens and ciliary-type visual cells similar to that of vertebrate eyes [139]. These G-coupled opsin photoreceptors of the retinylidene-forming protein family encoded in the genome of A. digitifera include rhodopsin, bacteriorhodopsin, c-opsin, r-opsin and G0-opsin (Table 5), but not the Gs-subfamily of opsin receptors reported to be present in sea anemones, hydra and jellyfish [140], that together with cyclic nucleotide-gated (CNG) ion channel proteins, arrestin (β-adrenergic receptor inhibitor) and other retino-protein receptors, are usual components of the bilaterian phototransduction cascade. Present also are genes to express rhodopsin kinase and β-adrenergic receptor kinase which are related members of the serine/threonine kinase family of proteins that specifically initiate deactivation of G-protein coupled receptors. Additional proteins of retinol metabolism of the phototransduction pathway encoded in the A. digitifera genome are retinol dehydrogenase, all-trans-retinol 13,14 reductase and phosphatidylcholine (lichthin)-retinol O-acyltransferase, a neural retina-specific leucine zipper protein that is an intrinsic regulator of photoreceptor development and function, and a retina and anterior neural fold homeobox-like protein that modulates the expression of photoreceptor genes within the rhodopsin promoter. The genome of A. digitifera encodes also a blue light-sensing, cryptochrome photoreceptor thought to signal synchronous coral spawning by detecting illumination from the lunar cycle [98, 99].

The A. digitifera genome reveals genes to express a broad array of neurotransmitter receptor proteins (Table 5), including glycine and glutamate neuroreceptors, adrenergic receptors that target non-dopamine catecholamines (i.e., epinephrine and norepinephrine), dopamine, muscarinic and nicotinic acetylcholine receptors, sensory G protein-coupled receptors and γ-aminobutyric acid (GABA) ligand-gated ion channel and G protein-coupled receptors (and inhibitors), several of which are encoded in high copy numbers. Cellular trafficking of neurotransmitters to presynaptic terminals is essential for neurotransmission, and significantly the genome of A. digitifera encodes a wide range of solute carrier neurotransmitter transporters, including a high affinity choline transporter and an acetylcholine-specific protein belonging to the major facilitator superfamily (MFS) of secondary transporters. Encoded also is dopamine β-monooxygenase that catalyses the conversion of dopamine to norepinephrine in the catecholamine biosynthetic pathway, which is necessary for cross-activation of adrenergic neuroreceptors [141]. Notably, the A. digitifera genome encodes acetylcholinesterase that is expressed at neuromuscular junctions and cholinergic synapses where its protease activity serves to terminate synaptic transmission.

The primitive nervous networks of cnidarians are strongly peptidergic with at least 35 neuropeptides identified from different cnidarian classes [142]. Our annotation of the sequenced A. digitifera genome, however, revealed only the neuropeptide FF-amide neurotransmitter, a RF amide related peptide, and its neuropeptide FF and Y receptors (Table 5). Neuropeptides are usually expressed as large precursor proteins which comprise multiple copies of “immature” neuropeptides. Our annotation did not readily reveal these precursor neuropeptide proteins, but we did find enzymes required for their processing, for example, a variety of carboxypeptidase enzymes (not tabulated) that remove propeptide carboxyl residues at basic peptidase sites, and the mature peptide neurotransmitters that are finished by consecutive modification by peptidylglycine (α-hydroxylating) monooxidase (PHM) and peptidyl α-hydroxyglycine α-amidating lyase (PAL) enzymes, both of which are commonly expressed in mammals as a single bifunctional peptidylglycine monooxygenase (K00504/EC 1.14.17.3) [143]. Our extensive catalogue of animal-like neural and sensory proteins revealed by genome annotation is testament that essential neurobiological features were developed in the primitive neural networks of early eumetazoan evolution.

Calcification and Ca2+-signalling proteins

The massive structures of coral reefs evident today are a construction of aggregated calcium carbonate deposited over long geological time by scleractinian corals and other calcifying organisms, yet our understanding of the molecular processes that regulate the biological processes of coral calcification is limited [144]. Ca2+ transfer from seawater to the calicoblastic site of coral calcification occurs by passive diffusion through the gastrovascular cavity [145] and by active calcium transport [146]. Active entry of Ca2+ through the oral epithelial layer is regulated by voltage-dependent calcium channels, such as demonstrated by the L-type alpha protein cloned from the reef-building coral Stylophora pistillata[147]. Ca2+ transport across the calioblastic ectoderm to the extracellular calcifying site is facilitated by the plasma-membrane ATP-dependent calcium pump that in S. pistillata resemble the Ca2+-ATPase family of mammalian proteins [148]. By 2H+/Ca2+-exchange at the calioblastic membrane, Ca2+-ATPase removes H+ (from the net reaction Ca2+ + CO2 + H2O CaCO3 + 2H+) thereby increasing the saturation state of CaCO3 to sustain calcium precipitation [146]. Importantly, located also at the calicoblastic membrane is carbonic anhydrase [149] which is required to catalyse the intermediate step of calcification by the reversible hydration of carbon dioxide (CO2 + H2O HCO3- + H+). In coral phototrophic symbiosis, despite numerous studies describing the well-known phenomenon of light-enhanced calcification, the relationship linking symbiont photosynthesis to coral calcification has been elusive [150, 151]. Nonetheless, efforts to better understand the calcifying response of scleractinian corals to environmental change and ocean acidification are gaining traction [149, 152, 153].

Voltage-gated calcium channels (VGCCs) have been examined extensively in mammalian physiology for converting membrane potential into intracellular Ca2+ transients for signalling transduction pathways (reviewed in [154]). VGCC signalling affects cellular processes to include muscle contraction, neuronal excitation, gene transcription, fertilisation, cell differentiation and development, proliferation, hormone release, activation of calcium-dependent protein kinases, cell death via necrosis and apoptosis pathways, phagocytosis and endo/exocytosis. Remarkably, annotation of the genome of A. digitifera reveals sequences encoding homologues of all the VGCC (α, αδ, β, and γ) subunits of the molecular (L, N, P/Q and R) phenotypes expressed in mammalian physiology (Table 6). There are multiple sequences encoding three variants of Ca2+-transporting ATPase, of which at least one is necessary for coral calcification. There is only one sequence match for expressing carbonic anhydrase in the genome of A. digitifera, which may reflect the high catalytic efficiency of this calcifying enzyme [155], although a BLAST search of ZoophyteBase does reveal scaffolds with low e-values which on future experimental inspection might uncover multiple copies of this enzyme essential for calcification. There are multiple sequences that express solute carrier Na+/Ca2+- and Na+/K+/Ca2+-exchange families of transport proteins that with expression of the coral Ca2+/H+-antiporter may regulate cellular pH and Ca2+ homeostasis.
Table 6

Calcification and Ca 2+ -signalling proteins in the predicted proteome of A. digitifera

Gene sequence

KEGG Orthology

Encoded protein description

v1.06452; v1.06451; v1.24424; v1.16923

K07300

Ca2+:H+ antiporter

v1.01669 [+ 9 other sequence copies]

K01537

Ca2+−transporting ATPase

v1.22367; v1.22366; v1.22365

K05850

Ca2+ transporting ATPase, plasma membrane

v1.19074

K05853

Ca2+ transporting ATPase, sarcoplasmic/endoplasmic reticulum

v1.22416; v1.22417; v1.15682; v1.00750

K14757

Calbindin D28

v1.24568 [+ 9 other sequence copies]

K01672

Carbonic anhydrase

v1.09241

K08272

Calcium binding protein 39

v1.02323 [+ 39 other sequence copies]

K13448

Calcium-binding protein CML

v1.05162 [+ 21 other sequence copies]

K13412

Calcium-dependent protein kinase

v1.09352

K07359

Calcium/calmodulin-dependent protein kinase kinase

v1.06475; v1.07555;v1.00945; v1.00159; v1.21122

K08794

Calcium/calmodulin-dependent protein kinase I

v1.06475; v1.01061; v1.21150; v1.22443

K04515

Calcium/calmodulin-dependent protein kinase II

v1.00159

K05869

Calcium/calmodulin-dependent protein kinase IV

v1.21927; v1.01218; v1.22226; v1.06623; v1.13703

K06103

Calcium/calmodulin-dependent serine protein kinase

v1.13460

K08284

Calcium channel MID1

v1.20738; v1.01401

K12841

Calcium homeostasis endoplasmic reticulum protein

v1.22794 [+ 11 other sequence copies]

K04612

Calcium-sensing receptor

v1.10079 [+ 17 other sequence copies]

K02183

Calmodulin

v1.10994

K14734

S100 calcium binding protein G

v1.02488 [+ 14 other sequence copies]

K05849

Solute carrier family 8 (sodium/calcium exchanger)

v1.23153 [+ 9 other sequence copies]

K13749

Solute carrier family 24 (sodium/potassium/calcium exchanger)

v1.14863

K12304

Soluble calcium-activated nucleotidase 1

v1.18656 [+ 13 other sequence copies]

K04858

Voltage-dependent calcium channel alpha-2/delta-1

v1.13222

K04860

Voltage-dependent calcium channel alpha-2/delta-3

v1.08078 [+ 9 other sequence copies]

K05315

Voltage-dependent calcium channel alpha 1, invertebrate

v1.03896 [+ 6 other sequence copies]

K05316

Voltage-dependent calcium channel alpha-2/delta, invertebrate

v1.04798

K05317

Voltage-dependent calcium channel beta, invertebrate

v1.22788

K04863

Voltage-dependent calcium channel beta-2

v1.09999

K04872

Voltage-dependent calcium channel gamma-7

v1.02505

K04873

Voltage-dependent calcium channel gamma-8

v1.03648[+ 6 other sequence copies]

K04850

Voltage-dependent calcium channel L type alpha-1C

v1.03648; v1.17267

K04851

Voltage-dependent calcium channel L type alpha-1D

v1.03648; v1.13219; v1.21895

K04857

Voltage-dependent calcium channel L type alpha-1S

v1.06313; v1.01656; v1.23096

K04344

Voltage-dependent calcium channel P/Q type alpha-1A

v1.08078 [+ 10 other sequence copies]

K04849

Voltage-dependent calcium channel N type alpha-1B

v1.07968

K04852

Voltage-dependent calcium channel R type alpha-1E

v1.01364; v1.13467; v1.08705

K04854

Voltage-dependent calcium channel T type alpha-1G

v1.15414; v1.14241; v1.09595

K04855

Voltage-dependent calcium channel T type alpha-1H

Implicit to coral calcification is Ca2+ regulation that affects signalling of other vital cellular functions. Cellular Ca2+ is mediated by the calcium-sensing receptor calmodulin (18 sequence matches) and other messenger calcium-binding effectors (Table 6), including the calcium-binding protein CML (40 protein domain sequence matches). Calcium/calmodulin-protein kinase proteins are arguably key to Ca2+-signalling in coral symbiosis but, with the exception of activation of sperm flagellar motility [156], their precise role has not been elaborated.

Plant-derived proteins

Endosymbiosis has contributed greatly to eukaryotic evolution, most notably to the genesis of plastids and mitochondria derived from prokaryotic antecedents. Genetic integration by endosymbiont-to-host transfer (EGT) or replacement (EGR) has been a significant force in early metazoan innovation, whereby nuclear transferred genes may even adopt novel functions in the host cell or replace existing versions of the protein that they encode [157]. Prokaryote-to-eukaryotic gene transfer has been widespread in evolution, but examples of genetic exchange between unrelated eukaryotes, such as between algal symbionts and their multicellular eukaryote host, are considered rare (reviewed by [158, 159]). One such example is aroB (3-dehydroquinate synthase) transferred to the genome of the sea anemone N. vectensis, which sequence best fits that of the dinoflagellate Oxyrrhis marina[119]. Close inspection of the amino acid sequence of the aroB gene product, as reported by Shinzato et al. [45], clearly shows this protein to be 2-epi-5-epi-valiolone synthase (EVS), a sugar phosphate cyclase orthologue that catalyses the conversion of sedoheptulose 7-phosphate to 2-epi-5-epi-valiolone found to be a precursor of the mycosporine-like amino acid (MAA) sunscreen shinorine in the cyanobacterium Anabaena variabilis[160]. Additionally, the EVS gene of N. vectensis has a distinctive O-methytransferase fusion that is identical in O. marina[161]. The shikimate pathway is essential to apicomplexan parasites of the genera Plasmodium, Toxoplasma and Cryptosporidium and of Tetrahymena ciliates to express a pentafunctional aroM gene similar to that of Ascomycetes, which is thought to have been conveyed by fungal gene transfer to a common ancestral progenitor [162]. In a separate example, H. viridis expresses a plant-like ascorbate peroxidase gene (HvAPX1) during oogenesis in both symbiotic and aposymbiotic individuals [163], whereby peroxidase activity is coincident with oogenesis and embryogenesis that in Hydra acts as a ROS scavenger to protect the oocyte from apoptotic degradation [164]. The sacoglossan (sea slug) molluscs Elysia chlorotica and E. viridis (Plakobranchidae) acquire plastids on ingestion of the siphonaceous alga Voucherea litorea (termed “kleptoplasty”) and, by maintaining sequestered plastids in an active photosynthetic state, has emerged as a model organism for the transfer of nuclear-encoded plant genes from algal symbiont to its animal host [165]. In this symbiosis, the family of light-harvesting genes psbO, prk (phosphoribokinase) and chlorophyll synthase (chlG) are entrained in the genome of Elysia chlorotica (reviewed in [166, 167]), although there is debate whether these genes are transcriptionally expressed (compare [168] and [169]). Also, phylogenomic analysis of the predicted proteins of the aposymbiotic unicellular choano-flagellate Monosiga brevicollis, considered to be a stem progenitor of the animal kingdom [170, 171], reveals 103 genes having strong algal affiliations arising from multiple phototrophic donors [172]. Such notable examples illustrate the transfer of algal genes to animal recipients.

KEGG orthology-based annotation of the predicted proteome of A. digitifera reveals a plethora of sequences presumed to be of algal origin (Table 7). Like E. chlorotica, the coral genome has encoded the photosystem II (PSII) protein PsbO of the oxygen-evolving complex of photosynthesis, as well as the PSII light-harvesting complex protein PsbL that is important in protecting PSII from photo-inactivation [173]. Encoded also are the photosystem I subunit proteins PsaI and PsaO. Additionally encoded are the photosystem P840 reaction center cytochrome c551 (PscC) protein and the photosynthetic reaction center M subunit protein, the light-harvesting proteins complex 1 alpha (PufA), the complex II chlorophyll a/b binding protein 6 (LHCB6), the cyanobacterial phycobilisome proteins AcpF and AcpG, the phycocyanin-associated antenna protein CpcD, the phycocyanobilin lyase protein CpcF and the phycoerythrin-associated linker protein CpeS. Like E. chlorotica, the coral genome encodes chlorophyll synthase (ChlG), a chlorophyll transporter protein PucC, a light-independent nitrogenase-like protochlorophyllide reductase enzyme that is sensitive to oxygen [174] and a red chlorophyll reductase essential to the detoxification of photodynamic chlorophyll catabolites arising from plant/algal senescence [175]. Three chlorosome proteins of the photosynthetic antenna complex of green sulphur bacteria, a bacteriochlorophyll methyltransferase involved in BChl c biosynthesis [176] and the retinylidene bacteriorhodopsin of phototrophic Archaea are also encoded in the coral genome. Present are genes encoding subunit 6 of the cytochrome B6f complex that links PSII and PSI via the plastoquinone pool, together with chloroplast ferredoxin-like NapH and NapG proteins and their 2Fe-2S cluster protein. The coral genome, however, encodes sequences for NAD+-ferredoxin reductase (HcaD; not tablulated), rather than the required NADP+-ferredoxin reductase of photosynthesis. Annotation of the A. digitifera genome revealed genes unexpectedly encoding ferredoxin hydrogenase [EC:1.12.7.2] and that of its small subunit protein (Table 7) involved in light-dependent production of molecular hydrogen having its [Fe-Fe]-cluster coupled to the photosynthetic transport chain via a charge-transfer complex with ferredoxin (see [177]).
Table 7

Plant-derived proteins in the predicted proteome of A. digitifera

Gene sequence

KEGG Orthology

Encoded protein description

v1.14452

K09843

(+)-Abscisic acid 8′-hydroxylase

v1.18868

K14496

Abscisic acid receptor PYR/PYL family (PYL)

v1.21983; v1.05890

K03342

p-Aminobenzoate synthetase / 4-amino-4-deoxychorismate lyase (PabBC)

v1.15436

K02822

Ascorbate-specific IIB component, PTS system (PTS-Ula-EiiB)

v1.11187; v1.13966

K00423

L-Ascorbate oxidase

v1.20081; v1.22465

K13604

Bacteriochlorophyll C20 methyltransferase (BchU)

v1.07723

K04641

Bacteriorhodopsin (BoP)

v1.21858

K04040

Chlorophyll synthase (ChlG)

v1.01742

K08945

Chlorosome envelope protein A (CsmA)

v1.04797; v1.14208

K08946

Chlorosome envelope protein B (CsmB)

v1.18698

K08948

Chlorosome envelope protein D (CamD)

v1.18637

K02642

Cytochrome b6f complex subunit 6 (PetL)

v1.21101; v1.14192; v1.14548

K01735

3-Dehydroquinate synthase (AroB)

v1.05796

K10210

4,4′-Diaponeurosporene oxidase (carotenoid biosynthesis; CrtP)

v1.11730

K04755

Ferredoxin, 2Fe-2S (FdX)

v1.19154; v1.00014

K00532

Ferredoxin hydrogenase

v1.00014

K00534

Ferredoxin hydrogenase small subunit

v1.17698; v1.06031; v1.16647

K02574

Ferredoxin-type protein (NapH)

v1.23058

K02573

Ferredoxin-type protein (NapG)

v1.08414

K08926

Light-harvesting complex 1 alpha chain (PufA)

v1.21458

K08917

Light-harvesting complex II chlorophyll a/b binding protein 6 (LHCB6)

v1.03743

K08226

MFS transporter, BCD family, chlorophyll transporter (PucC)

v1.13030; v1.08678

K13413

Mitogen-activated protein kinase kinase 4/5, plant ((MKK4_5P)

v1.02429; v1.10744; v1.03340

K08929

Photosynthetic reaction center M subunit (PufM)

v1.03631

K02696

Photosystem I subunit VIII (PsaI)

v1.11432

K14332

Photosystem I subunit (PsaO)

v1.17422

K02713

Photosystem II protein (PsbL)

v1.18303

K02716

Photosystem II oxygen-evolving enhancer protein 1 (PsbO)

v1.12300; v1.21136

K08942

Photosystem P840 reaction center cytochrome c551 ((PscC)

v1.00280

K02097

Phycobilisome core component 9 (AcpF)

v1.10967

K02290

Phycobilisome rod-core linker protein (AcpG)

v1.02166

K02287

Phycocyanin-associated, rod protein (CpcD)

v1.19642; v1.07305; v1.19572; v1.01248

K02289

Phycocyanobilin lyase beta subunit (CpcF)

v1.10441

K05382

Phycoerythrin-associated linker protein (CpeS)

v1.13406

K10027

Phytoene dehydrogenase (desaturase; CrtI)

v1.18809; v1.06199

K02291

Phytoene synthase (CrtB)

v1.20411; v1.02037; v1.14064; v1.21095

K09060

Plant G-box-binding factor (GBF)

v1.10035

K00218

Protochlorophyllide reductase [NifEN-like; Por]

v1.21846

K05358

Quinate dehydrogenase (QuiA)

v1.03127

K13545

Red chlorophyll catabolite reductase (ACD2)

v1.05899

K00891

Shikimate kinase (AroK, AroL)

v1.21101; v1.14192; v1.05899

K13829

Shikimate kinase / 3-dehydroquinate synthase (AroKB)

v1.12938

K08500

Syntaxin of plants (SYP6)

v1.06575

K08506

Syntaxin of plants (SYP7)

v1.04929

K09834

Tocopherol cyclase (VTE1, SXD1)

v1.01022

K05928

Tocopherol O-methyltransferase

v1.05457

K09838

Zeaxanthin epoxidase (ZEP, ABA1)

Like N. vectensis and the dinoflagellate Oxyrrhis marina, the genome of A. digitifera encodes an O-methyltransferase which is immediately downstream of EVS, but the two genes are not fused. Using a ZoophyteBase BlastP search, the O-methyltransferase showed little sequence homology with the corresponding protein of A. variabilis (e-value of 6.972E-2 and Bit score of 34.27), whereas the EVS protein shared 87% absolute sequence identity to the A. variabilis EVS protein. What role, if any, these two genes play in mycosporine-like amino acid (MAA) biosynthesis in A. digitifera has yet to be determined, although it has been suggested from the transcriptome of Acropora microphthalma that MAA biosynthesis proceeds from a branch point at 3-dehydroquinate of the shikimic acid pathway as a shared metabolic adaptation between the coral host and its symbiotic zooxanthellae [40]. The 3-dehydroquinate synthase enzyme of the shikimic acid pathway, thought to be a key intermediate in an alternative MAA biosynthetic pathway in A. variabilis[178], is instead encoded by the fused aroKB gene of A. digitifera (Table 7). Additional shikimate proteins of the predicted proteome, although not limited to phototrophs, are shikimate kinase (AroK), quinate dehydrogenase (QuiA) and the conjoined p-aminobenzoate synthase and 4-amino-4-deoxychlorismate lysate (PabBC) enzyme necessary for folate biosynthesis [179]. Other plant-related gene homologues include the phytohormone abscisic acid receptor protein (PabBC) and its cytochrome P450 monooxygenase abscisic acid 8′-hydroxylase, L-ascorbate oxidase and PTS system degrading enzymes, the unique SYP6 and SYP7 syntaxins of plant vesicular transport, tocopherol cyclase and a tocopherol O-methyltransferase enzyme that converts γ-tocopherol to α-tocopherol. Essential for carotene biosynthesis are phytoene synthase (CrtB) and phytoene dehydrogenase (CrtI) enzymes. Significantly, encoded within the coral genome is zeaxanthin epoxidase that is essential for abscisic acid biosynthesis and is a key enzyme in the xanthophyll cycle of plants and algae to impart oxidative stress tolerance.

Given that viruses often mediate gene transfer processes, it is intriguing that certain bacteriophages of marine Synechococcus and Prochlorococcus cyanobacteria are reported to carry genes encoding the photosynthesis D1 (psbA), and D2 (psbD) proteins, a high-light inducible protein (HLIP) [180, 181] and the photosynthetic electron transport plastocyanin (petE) and ferredoxin (petF) proteins thought to enhance the photosynthetic fitness of their host [182184]. Accordingly, it has been suggested that the transfer of psbA by viruses associated with Symbiodinium could lessen the severity of thermal impairment to PSII and the response of corals to thermal bleaching [185]. It is yet unknown if phages or dinoflagellate-infecting viruses [186], particularly those of Symbiodinium[187], may affect gene transfer leading to complementary (or “shared”) metabolic adaptations of symbiosis [119, 188].

Proteins of nitrogen metabolism

It is well accepted that intracellular Symbiodinium spp. provide reduced carbon for coral heterotrophic metabolism by photosynthetic carbon fixation. Because of this metabolic relationship, light is a critical feature in the bioenergetics of coral symbiosis [189]. The algal photosynthate translocated to corals, however, is deficient in nitrogen at levels necessary to sustain autotrophic growth. While corals can assimilate fixed nitrogen from surrounding seawater [190], “recycled” nitrogen within the symbiosis may account for as much as 90% of the photosynthetic nitrogen demand [191]. It would not be surprising then that light would have a strong influence on the uptake and retention of ammonium by symbiotic corals. Consequently, corals excrete excess ammonium in darkness [192], and in light excretion is induced by treatment with the photosynthetic electron transport inhibitor 3-(3,4-diclorophenyl)-1,1-dimethylurea (DCMU) [193]. Since ammonia is the product of nitrogen fixation, these observations suggest that the coral holobiont may fix nitrogen in the dark, or when photosynthesis is repressed, during which coral tissues are hypoxic [194], and nitrogenase activity is not inactivated by molecular oxygen [195].

Tropical coral reefs are typically surrounded by low-nutrient oceanic waters of low productivity but, paradoxically, the waters of coral reefs often have elevated levels of inorganic nitrogen [196, 197] attributed to high rates of nitrogen fixation. While nitrogen fixation from diazotrophic epiphytes of the coral reef substrata and sediments [197, 198] and diazotrophic bacterioplankton of the coral reef lagoon [199] provide substantial quantities of fixed nitrogen for assimilation by the coral reef, mass-balance estimates show this input to be less than the community’s annual nitrogen demand [200]. Endolithic nitrogen-fixingbacteria are abundant in the skeleton of living corals where they benefit from organic carbon excreted by overlaying coral tissues to provide a ready source of energy for dinitrogen reduction [201]. Additionally, intracellular nitrogen-fixing cyanobacteria are reported to coexist with dinoflagellate symbionts in the tissues of Monastraea cavernosa and to functionally express nitrogenase activity [202]. Corals also harbour a diverse assemblage of heterotrophic microorganisms in their skele-ton, tissues and lipid-rich mucus (reviewed in [203]), and these communities include large populations of diazo-trophic bacteria [204, 205], and archaea [206]. Apart from nitrogen fixation, the coral microbiota contributes to other nitrogen-cycling processes, such as nitrification, ammonification and denitrification [207, 208]. We were surprised to find several nitrogen fixation and cycling proteins encoded in the genome of A. digitifera (Table 8), notably a nitrogen fixation NifU-like protein, the Nif-specific regulatory protein (NifA), the regulatory NAD(+)-dinitrogen-reductase ADP-D-ribosylastransferase protein, a nitrifying ammonia monooxy-genase enzyme and nitrate reductase, which are usually expressed only by prokaryotic microorganisms.
Table 8

Proteins of nitrogen metabolism in the predicted proteome of A. digitifera

Gene sequence

KEGG Orthology

Encoded protein description

v1.23444; v1.09133; v1.23443

K05521

ADP-ribosylglycohydrolase (DraG)

v1.09202

K10944

Ammonia monooxygenase subunit A

v1.03645 [+ 8 other sequence copies]

K03320

Ammonium transporter, Amt family

v1.12268; v1.12269

K06580

Ammonium transporter Rh

v1.02406

K01954

Carbamoyl-phosphate synthase (CPS)

v1.01524; v1.18283; v1.18284

K01948

Carbamoyl-phosphate synthase (CPS, ammonia)

v1.01615

K04016

Formate-dependent nitrite reductase (NrfA)

v1.16277; v1.23483; v1.13667; v1.22675

K00261

Glutamate dehydrogenase (NAD(P)+)

v1.17166; v1.11089

K01745

Histidine ammonia-lyase

v1.22825; v1.08034;v1.o8520

K05123

Integration host cell factor (INF) subunit beta

v1.11343

K05951

NAD+−dinitrogen-reductase ADP-D-ribosyltransferase (DraT)

v1.00547

K02584

Nif-specific regulatory protein (NifA)

v1.18869

K00371

Nitrate reductase 1, beta subunit

v1.06763

K08346

Nitrate reductase 2, beta subunit

v1.14858; v1.00685; v1.23148

K05916

Nitric oxide dioxygenase

v1.16954 [+ 5 other sequence copies]

K02448

Nitric oxide reductase NorD protein

v1.06115

K02164

Nitric oxide reductase NorE protein

v1.17629 [+ 12 other sequence copies]

K04748

Nitric oxide reductase NorQ protein

v1.24077 [+ 4 other sequence copies]

K13125

Nitric oxide synthase-interacting protein

v1.21801; v1.05719; v1.23577; v1.19464

K13253

Nitric-oxide synthase, invertebrate

v1.05980

K00363

Nitrite reductase (NAD(P)H) small subunit

v1.00101

K02598

Nitrite transporter NirC

v1.02355; v1.18772

K04488

Nitrogen fixation protein NifU

v1.17812

K02589

Nitrogen regulatory protein PII 1

v1.09150

K02570

Periplasmic nitrate reductase NapD

v1.01560

K02571

Periplasmic nitrate reductase NapE

v1.10035

K00218

Protochlorophyllide reductase [NifEN-like]

v1.08939

K03737

Pyruvate-flavodoxin reductase (NifJ)

v1.17373

K00365

Urate oxidase

v1.13217

K01427

Urease

v1.16409 [+ 5 other sequence copies]

K03187

Urease accessory protein

v1.13217

K01429

Urease subunit beta

v1.13217

K14048

Urease subunit gamma/beta

v1.12211 [+ 4 other sequence copies]

K00106

Xanthine dehydrogenase/oxidase

v1.12212

K13481

Xanthine dehydrogenase small subunit

(Excluding amino acid and pyrimidine/purine nucleotide synthesis or metabolism).

The presence of genes encoding proteins involved in nitrogen fixation raises speculation that corals may contribute directly to, or perhaps co-regulate, certain processes that catalyse the reduction of dinitrogen (N2) to ammonia (NH3) by the enzyme nitrogenase reductase (NifH). The functional NifH enzyme is a binary protein composed of a molybdenum-iron (MoFe) protein (NifB/NifDK), or its NifEN homologue, fused with a FeMo-cofactor (FeMoco) protein [209]. While genes encoding NifB, NifDK (or NifEN) and their FeMo-cofactor do not appear in the genome of A. digitifera, a gene encoding the NifEN-like protein protochlorophyllide oxidoreductase (POR) is present (Table 8). POR has all three subunits with high similarity to the assembled MoFe nitrogenase [210], but this homologue is unlikely to be effective in nitrogen reduction [211, 212] since its activity is light dependent [213] when tissues are highly oxic [193]. The NifU protein encoded in the coral genome preassembles the metallocatalytic Fe-S clusters for maturation of nitrogenase [214], but its assemblage without NifS, a cysteine desulfurase needed for [Fe-S] cluster assembly [215], would be incomplete, and its pre-nitrogenase receptor is also missing. Yet, the coral does have the nifJ gene that encodes pyruvate:flavodoxin oxidoreductase required for electron transport in nitrogenase reduction [216]. The regulatory NifA protein encoded in the coral genome might activate, on stimulation by the integration host factor (INF), transcription of nitrogen fixation (nif) operons of RNA polymerase [217], and both of these proteins are encoded in the coral genome. Additional to this transcriptional control, post-translational nitrogenase activity is controlled by reversible ADP-ribosylation of a specific arginine residue in the nitrogenase complex [218]. NAD(+)-dinitrogen-reductase ADP-D-ribosyltransferase (DraT) inactivates the nitrogenase complex while ADP-ribosylgly-cohydrolase (DraG) removes the ADP-ribose moiety to restore nitrogenase activity, and both of these enzymes are encoded in the coral genome. Given that genes encoding essential constituent proteins of nitrogenase assembly appear incomplete, corals are unlikely to fix nitrogen per se, but co-opted elements of the coral genome to regulate processes of nitrogen fixation by its diazotrophic consortia is a prospect worthy of exploration [219].

Nitrofying/nitrifying bacteria and archaea express the enzyme ammonia monooxygenase that converts fixed ammonia to nitrite (via hydroxylamine) and the enzyme nitrite (oxido)reductase completes the oxidation of nitrite to nitrate, and both of these enzymes are entrained in the genome of A. digitifera (Table 8). The ammonia monooxygenase subunit A (amoA) of archaeal consorts has been described in nine species of coral from four reef locations [220], but the presence of amoA in the coral genome, together with encoded ammonium transport proteins, was not anticipated. Another protein of prokaryotic origin encoded in the coral genome is nitrate reductase (periplasmic, assimilatory and respiratory), the latter being required for anaerobic respiration by bacteria [221], and unlike the nitrate reductase family of sulphite oxidase enzymes in eukaryotes, the nitrate reductases of prokaryotes (K00363) belong to the DMSO reductase family of enzymes. Also encoded in the coral genome are a nitrite transporter (NirC) and a formate-dependent nitrite reductase (NrfA) required for nitrite ammonification [222]. In addition to nitrite reduction, NrfA reduces nitric oxide, hydroxylamine, nitrous oxide and sulphite, the last providing a metabolic link between nitrogen and sulphur cycling in coral metabolism. Other enzymes of nitrogen metabolism encoded in the coral genome are the carbamoyl-phosphate synthase family of enzymes [223] that catalyses the ATP-dependent synthesis of carbamoyl phosphate used for the production of urea (ornithine cycle) to provide a ready store of fixed-N in the urea-nitrogen metabolism of corals [224]. Another nitrogen source comes from glutamate dehydrogenase (GDH) that reversibly converts glutamate to α-ketoglutarate with liberation of ammonia, and as expected [225], this enzyme is encoded in the coral genome, together with the prokaryotic nitrogen regulatory protein PII of glutamine synthase, which in bacteria is activated in response to nitrogen availability. Encoded also is histidine ammonia-lyase (histidase) that liberates ammonia (and urocanic acid) from cytosolic stores of histidine. It is now accepted that uric acid deposits accumulated by symbiotic algae provide a significant store of nitrogen for the coral holobiont [226], so it is noteworthy that the coral genome encodes urate oxidase (uricase) to catalyse uric acid oxidation to allanotoin from which urea and ureidoglycolate are produced in a reaction catalysed by allantoicase (allantoate amidinohydrolase), both of which known isoforms are present in the coral genome. Encoded in the coral genome is also urease to catalyse the hydrolysis of urea, presumably excreted by its algal symbionts, with the release of carbon dioxide and ammonia to meet the nitrogen demand of the coral holobiont during periods of low nitrogen availability. Similarly, xanthine dehydrogenase (xanthine: NAD+-oxidoreductase) acts by oxidation on a variety of purines, including hypoxanthine, to yield urate for the recycling of nitrogen in coral nutrition. Many of the aforementioned proteins of nitrogen metabolism, including Nif proteins, have been detected in the proteome of an endosymbiont-enriched fraction of the coral S. pistillata[39].

Notwithstanding consideration of the rapid diffusion rate of nitric oxide (NO) or its apparent short biological half-life [227], there is debate about the provenance of endogenously produced NO in signalling the bleaching of corals in response to environmental stress. Elevated nitric oxide synthase (NOS) activity and NO production in algal symbionts has been attributed to the thermal stress response of corals [228, 229], whereas the host is ascribed to be the major source of NO during exposure to elevated temperature [230, 231]. While our annotation may not resolve this dispute, we show (Table 8) that nitric oxide synthase enzymes (Nor D, Nor E, Nor Q and an invertebrate NOS protein) are encoded in the genome of A. digitifera, together with a nitric oxide-interacting protein (NOIP) that in higher animals regulates neuronal NOS activity [232]. Nitric oxide is an intermediate of nitrite reduction catalysed by nitrite reductase (NIR), which by further reduction produces ammonia. The coral genome also encodes nitric oxide dioxygenase (NOD) that converts nitric oxide to nitrate. Accordingly, enhanced expression of NIR (NO reduction) or NOD (NO oxidation) could ameliorate the NO-signalling response of coral bleaching presumed activated by environmental stress.

DNA repair

Cellular DNA is prone to damage caused by the products of normal metabolism and by exogenous agents. Damage to DNA from metabolic processes include the oxidation of nucleobases and strand interruptions by the production of reactive oxygen species (ROS), from alkylation of nucleotide bases, from the hydrolysis of bases causing deamination, depurination and depyrimidination, and from the mismatch of base pairs from errors in DNA replication. Damage affected by external agents include exposure to UV light causing pyrimidine dimerization and free radical-induced damage, exposure to ionising radiation causing DNA strand breaks, thermal disruption causing hydrolytic depurination and single-strand breaks, and by xenobiotic contamination to cause DNA adduct formation, nucleobase oxidation and DNA crosslinking. Most of these lesions affect structural changes to DNA that alter or prevent replication and gene transcription at the site of DNA damage. Thus, recognition and repair of DNA abnormalities are vital processes essential to maintain the genetic integrity of the coral genome. Since there are multiple pathways causing DNA damage at diverse molecular sites, there are likewise diverse and overlapping processes available to repair cellular DNA damage. Of the many nuclear repair processes, photoreactivation (photolyase), base excision repair and nucleotide excision repair are the main elements for the repair of cellular DNA damage.

Exposure to sunlight is an absolute requirement for phototrophic symbiosis, but excessive exposure of corals to solar ultraviolet radiation can inflict direct damage to DNA by pyrimidine dimerization and 6-4 photoadduct formation and cause indirect damage by the production of ROS to initiate free-radical damage. While there have been abundant studies on the sensitivity of corals to solar ultraviolet radiation, only a few have examined the effects of solar UV to cause DNA damage. Photoreactivation has been shown to be an important repair pathway for reversing UV-activated DNA damage in adult coral [233] and coral planulae [234]. UV damage to DNA was first demonstrated by the detection of unrepaired cyclobutane pyrimidine dimers (CPDs) in the host tissues and algal symbionts of the coral Porities porites, in which CPDs had increased in a UV dose-dependent manner [235], whereas CPDs and 6-4 pyrimidine-pyrimidone photoadducts in the coral Montipora verrucosa holobiont were correlated inversely with levels of coral “sunscreen” protection [236]. The effects of solar UV radiation causing DNA lesions in coral have been determined by use of the comet assay [237], and UV-induced DNA damage and repair has been examined in the symbiotic anemone Aiptasia pallida[238]. The comet assay showed also that DNA lesions in coral planulae had increased on acquiring algal symbionts, presumably from greater ROS production resulting as a by-product of photosynthesis [239]. Iron-induced oxidative stress was found likewise to enhance DNA damage in the coral Pocillopora damicornis as determined by the occurrence of DNA apurinic/apyrimidinic sites caused by hydrolytic lesions [240]. Significantly, DNA damage in the host and algal symbionts of the coral Montastraea faveo-lata was found to occur simultaneously during thermal “bleaching” stress, and DNA damage is further enhanced on exposure to greater irradiances of solar radiation [241]. Nevertheless, despite the serious risk of unrepaired DNA damage to coral survival, the DNA repair processes of corals to mitigate the detrimental effects of environmental stress have not been adequately characterised at the transcriptome level of expression [29, 242].

Our annotation of the sequenced genome of A. digitifera has revealed genes encoding a large repertoire of DNA repairing enzymes and their adaptor proteins (Table 9). Given strong evidence for DNA photoreactivation in corals having been reported [233, 234], it was surprising to find only one gene in single copy that encodes a sole photolyase enzyme for reversing pyrimidine dimer and 6-4 photoadduct formation. Notably, we found genes encoding 6 members of the ERCC family of nucleotide excision repair enzymes, together with the UV excision repair protein RAD23, for the repair of UV-induced DNA damage. More abundant are the DNA mismatch repair enzymes from the MLH, MSH, Mut and PMS protein families and related glycosylase/lyase proteins for repairing erroneous insertion, deletion and mis-incorporation of bases to arise during DNA replication and recombination. There is additionally a specific gene that encodes a 3′-endonuclease protein that has a preference to correct mispaired nucleotide sequences. Abundant also are other members of the RAD-family of DNA repair proteins, including 28 sequence copies of a gene encoding the RAD50 protein for DNA double-strand break repair that, together with members of the MRE, Rec, REV, Swi5/Sae3, XRCC and XRS families of recombination and polymerase proteins, have complementary roles in DNA repair. Apparent also in the genome are the DNA helicase proteins, including RuvB–like proteins, which are primarily involved in DNA replication and transcription, but assist also in the repair of DNA damage by separating double strands at affected sites of DNA damage to facilitate repair. Of the multiple families of ATP-dependent DNA helicase proteins encoded in the coral genome, RecQ and helicase Q predominate. Encoded in the coral genome are 5 homologues of the DNA repair alkB proteins that reverse damage to DNA from alkylation caused by chemical agents by removing methyl groups from 1-methyl adenine and 3-methyl cytosine products in single-stand DNA. Annotated also are genes encoding DNA ligase 3 for repairing single-strand breaks, DNA ligase 4 to repair double-strand breaks, and a DNA cross-link repair 1C protein with single-strand specific endonuclease activity that may serve in a proofreading function for DNA polymerase. Taken together, expressing this arsenal of DNA protection may provide corals with limited ability to transcribe gene-encoded adaptation to a changing global environment.
Table 9

DNA repair proteins in the predicted proteome of A. digitifera

Gene sequence

KEGG Orthology

Encoded protein description

v1.02961; v1.13402

K03575

A/G-specific adenine glycosylase (MutY)

v1.11766

K03919

Alkylated DNA repair protein

v1.04821

K10765

Alkylated DNA repair protein alkB homologue 1

v1.02479

K10766

Alkylated DNA repair protein alkB homologue 4

v1.20302

K10767

Alkylated DNA repair protein alkB homologue 5

v1.24450

K10768

Alkylated DNA repair protein alkB homologue 6

v1.02766; v1.09413

K10770

Alkylated DNA repair protein alkB homologue 8

v1.01590 [+ 4 other sequence copies]

K10884

ATP-dependent DNA helicase 2 subunit 1

v1.18810; v1.03166; v1.08449

K10885

ATP-dependent DNA helicase 2 subunit 2

v1.08013

K03722

ATP-dependent DNA helicase DinG

v1.03542

K14635

ATP-dependent DNA helicase MPH1

v1.06737 [+ 5 other sequence copies]

K15255

ATP-dependent DNA helicase PIF1

v1.17360; v1.21235

K10899

ATP-dependent DNA helicase Q1

v1.01081 [+ 8 other sequence copies]

K10730

ATP-dependent DNA helicase Q4

v1.16859

K10902

ATP-dependent DNA helicase Q5

v1.11661 [+ 19 other sequence copies]

K03654

ATP-dependent DNA helicase RecQ

v1.20397

K03656

ATP-dependent DNA helicase Rep

v1.18049; v1.07731; v1.05830

K10905

ATR interacting protein

v1.01679

K01669

Deoxyribodipyrimidine photo-lyase

v1.03410; v1.12968; v1.00865; v1.16876

K10887

DNA cross-link repair 1C protein

v1.07474; v1.07473; v1.01809

K10610

DNA damage-binding protein 1

v1.13116; v1.03378; v1.16328

K10140

DNA damage-binding protein 2

v1.17099 [+ 5 other sequence copies]

K11885

DNA damage-inducible protein 1

v1.05469

K06663

DNA damage checkpoint protein

v1.02859; v1.14719; v1.21030; v1.10920

K04452

DNA damage-inducible transcript 3

v1.02191

K10844

DNA excision repair protein ERCC-2

v1.19108 [+ 5 other sequence copies]

K10843

DNA excision repair protein ERCC-3

v1.22267 [+ 4 other sequence copies]

K10848

DNA excision repair protein ERCC-4

v1.15137 [+ 5 other sequence copies]

K10846

DNA excision repair protein ERCC-5

v1.18550; v1.02606; v1.14935; v1.08831

K10841

DNA excision repair protein ERCC-6

v1.20045; v1.01844; v1.11724; v1.03203

K10570

DNA excision repair protein ERCC-8

v1.15430; v1.03058

K03658

DNA helicase IV

v1.00228 [+ 4 other sequence copies]

K11665

DNA helicase INO80

v1.00136; v1.0678; v1.21529

K10776

DNA ligase 3

v1.23293; v1.19418; v1.23430; v1.15721

K10777

DNA ligase 4

v1.19248

K07458

DNA mismatch endonuclease, patch repair protein

v1.19011

K08739

DNA mismatch repair protein MLH3

v1.11513; v1.11449

K08735

DNA mismatch repair protein MSH2

v1.14781

K08736

DNA mismatch repair protein MSH3

v1.05696; v1.22444; v1.19162

K08740

DNA mismatch repair protein MSH4

v1.04904

K08741

DNA mismatch repair protein MSH5

v1.15360; v1.19426; v1.08585

K08737

DNA mismatch repair protein MSH6

v1.02429 [+ 8 other sequence copies]

K03572

DNA mismatch repair protein MutL

v1.03990

K03555

DNA mismatch repair protein MutS

v1.14015

K07456

DNA mismatch repair protein MutS2

v1.08443

K10864

DNA mismatch repair protein PMS1

v1.15229

K10858

DNA mismatch repair protein PMS2

v1.08658; v1.14152; v1.01681

K15082

DNA repair protein RAD7

v1.16407 [+ 27 other sequence copies]

K10866

DNA repair protein RAD50

v1.22193

K04482

DNA repair protein RAD51

v1.02646; v1.22076

K10958

DNA repair protein RAD57

v1.15671 [+ 4 other sequence copies]

K04483

DNA repair protein RadA

v1.16193; v1.19033

K04485

DNA repair protein RadA/Sms

v1.16079; v1.07685

K04484

DNA repair protein RadB

v1.21363; v1.22360; v1.02900

K03584

DNA repair protein RecO (recombination protein O)

v1.18390

K03515

DNA repair protein REV1

v1.04705

K10991

DNA repair protein Swi5/Sae3

v1.13920; v1.03800; v1.16133

K10803

DNA repair protein XRCC1

v1.15052

K10879

DNA repair protein XRCC2

v1.09315 [+ 4 other sequence copies]

K10886

DNA repair protein XRCC4

v1.02733; v1.24592

K10868

DNA repair protein XRS2

v1.14551; v1.23176

K10873

DNA repair and recombination protein RAD52

v1.20503 [+ 4 other sequence copies]

K10875

DNA repair and recombination protein RAD54

v1.23173; v1.16050

K10877

DNA repair and recombination protein RAD54B

v1.07227; v1.08907; v1.09439; v1.02644

K10847

DNA repair protein complementing XP-A cells

v1.11534 [+ 5 other sequence copies]

K10865

Double-strand break repair protein MRE11

v1.07939

K03660

N-glycosylase/DNA lyase

v1.16163

K03652

3-Methyladenine DNA glycosylase

v1.07231

K10726

Replicative DNA helicase Mcm

v1.05482

K04499

RuvB-like protein 1 (pontin 52)

v1.19813

K11338

RuvB-like protein 2

v1.06890

K15080

Single-strand annealing weakened protein 1

v1.17193; v1.14087

K03111

Single-strand DNA-binding protein

v1.15575

K10800

Single-strand monofunctional uracil DNA glycosylase

v1.07134

K10992

Swi5-dependent recombination DNA repair protein 1

v1.13860

K03649

TDG/mug DNA glycosylase family protein

v1.14423; v1.14399; v1.05070

K03648

Uracil-DNA glycosylase

v1.23838

K10791

Three prime repair exonuclease 2

v1.19522

K10839

UV excision repair protein RAD23

Stress response proteins

Annotation of the A. digitifera genome reveals a wide assortment of thermal shock proteins, molecular chaperones and other stress response elements that are given in (Table 10), excluding antioxidant and redox-protective proteins which are described in the next section. Heat shock proteins 70 kDa, 90 kDa, 110kDA, HspQ and HspX (the last two proteins being homologues of the bacterial heat shock factor sigma32 and α-crystallin, respectively) are encoded in the coral genome, together with several HSP gene transcription factors. HSPs play a role in various cellular functioning such as protein folding, intracellular protein trafficking and resistance to protein denaturation. HSP expression is usually increased on exposure to elevated temperatures and other conditions of biotic and abiotic stress that include infection, inflammation, metabolic hyperactivity, exposure to environmental toxicants, ultraviolet light exposure, starvation, hypoxia and desiccation [243]. HSPs and chaperones are transcriptionally regulated and are induced by heat shock transcription factors [244], of which there are several encoded in the coral genome. Since HSPs are found in virtually all living organisms, it is not surprising that cnidarian hsp transcription and protein expression (HSP60, HSP70 and HSP90) have been profiled as a stress determinant [245250] and early warning indicator of coral bleaching [251254]. The coral genome reveals also a cold shock protein encoded by the cspA gene family, but profiling its expression with other stress response proteins activated by sub-optimum cold temperatures [255] has not been reported. Additionally, the coral genome encodes transcription of a homologue of the universal stress protein A (UspA), a member of an ancient and conserved group of stress-response proteins [256, 257], which have been studied mostly in bacteria [258] but have been described also in several plants [259] and animals, including members of the Cnidaria [260]. Usp transcripts have been quantified in the thermal stress response of the coral Montastraea faveolata[261] and its aposymbiotic embryos [262]. Another gene product of potential interest is a homologue of the oxidative-stress responsive protein 1 (OXSR1) that belongs to the Ser/Thr kinase family of proteins, as do other mitogen-stress activated protein kinases (MAPKs), that regulate downstream kinases in response to environmental stress [263] by interacting with the Hsp70 subfamily of proteins [264]. Another significant response protein encoded in the coral genome (Table 10) is a homologue of the stress-induced phosphoprotein 1 (30 domain sequence alignments), known also as the Hsp70-Hsp90 organising protein (HOP) belonging to the stress inducible (STI1) family of proteins, which is a principle adaptor protein that mediates the functional cooperation of molecular chaperones Hsp70 and Hsp90 [265, 266]. It is yet to be determined if Hop1 transcription may serve as a primary indicator of environmental stress in corals.
Table 10

Stress response proteins in the predicted proteome of A. digitifera

Gene sequence

KEGG Orthology

Encoded protein description

v1.04616; v1.06277

K03694

ATP-dependent Clp protease subunit ClpA

v1.04617; v1.23486; v1.23484; v1.10207

K03695

ATP-dependent Clp protease subunit ClpB

v1.13464

K03697

ATP-dependent Clp protease subunit ClpE

v1.06903; v1.11461

K06891

ATP-dependent Clp protease adaptor protein ClpS

v1.12577; v1.09531; v1.17184

K03544

ATP-dependent Clp protease subunit ClpX

v1.09407

K08054

Calnexin (protein-folding chaperone)

v1.16781

K08057

Calreticulin (Ca2+-binding chaperone)

v1.04005

K10098

Calreticulin 3 (Ca2+-binding chaperone)

v1.02702[+ 5 other sequence copies]

K03704

Cold shock protein (beta-ribbon, CspA family)

v1.01907; v1.18998

K07213

Copper chaperone

v1.23457; v1.01713; v1.19228

K04569

Copper chaperone for superoxide dismutase

v1.08719; v1.19128

K09502

DnaJ homologue subfamily A member 1

v1.08719; v1.18432

K09503

DnaJ homologue subfamily A member 2

v1.16210; v1.22054

K09504

DnaJ homologue subfamily A member 3

v1.19128

K09505

DnaJ homologue subfamily A member 4

v1.04818 [+ 6 other sequence copies]

K09506

DnaJ homologue subfamily A member 5

v1.02841; v1.02842

K09507

DnaJ homologue subfamily B member 1

v1.00368; v1.13308; v1.16977; v1.03340

K09508

DnaJ homologue subfamily B member 2

v1.11537; v1.09205; v1.08628; v1.02840

K09511

DnaJ homologue subfamily B member 5

v1.24549 [+ 9 other sequence copies]

K09512

DnaJ homologue subfamily B member 6

v1.01573

K09513

DnaJ homologue subfamily B member 7

v1.00352; v1.09196; v1.06645

K09514

DnaJ homologue subfamily B member 8

v1.18536 [+ 4 other sequence copies]

K09515

DnaJ homologue subfamily B member 9

v1.14710

K09517

DnaJ homologue subfamily B member 11

v1.14959

K09518

DnaJ homologue subfamily B member 12

v1.09205

K09519

DnaJ homologue subfamily B member 13

v1.16242

K09520

DnaJ homologue subfamily B member 14

v1.20109; v1.03468

K09521

DnaJ homologue subfamily C member 1

v1.07111 [+ 5 other sequence copies]

K09522

DnaJ homologue subfamily C member 2

v1.21077 [+ 13 other sequence copies]

K09523

DnaJ homologue subfamily C member 3

v1.07739; v1.22910

K09524

DnaJ homologue subfamily C member 4

v1.01239 [+ 13 other sequence copies]

K09525

DnaJ homologue subfamily C member 5

v1.17629 [+ 29 other sequence copies]

K09527

DnaJ homologue subfamily C member 7

v1.18619; v1.08300; v1.23789

K09528

DnaJ homologue subfamily C member 8

v1.13575; v1.04213

K09529

DnaJ homologue subfamily C member 9

v1.05956; v1.05955; v1.21265; v1.21205

K09530

DnaJ homologue subfamily C member 10

v1.13525; v1.04120

K09531

DnaJ homologue subfamily C member 11

v1.09496 [+ 4 other sequence copies]

K09533

DnaJ homologue subfamily C member 13

v1.24546

K09534

DnaJ homologue subfamily C member 14

v1.05866

K09536

DnaJ homologue subfamily C member 16

v1.16151; v1.08307; v1.14980

K09537

DnaJ homologue subfamily C member 17

v1.16309

K09539

DnaJ homologue subfamily C member 19

v1.05241; v1.22999; v1.17372

K14258

Facilitated trehalose transporter (anhydrobiosis)

v1.12967; v1.19789

K14590

FtsJ methyltransferase [heat shock protein]

v1.02247

K09414

Heat shock transcription factor 1

v1.24112

K09416

Heat shock transcription factor 3

v1.05839

K09419

Heat shock transcription factor, other eukaryote

v1.12890 [+ 10 other sequence copies]

K03283

Heat shock 70 kDa protein 1/8

v1.07996

K09489

Heat shock 70 kDa protein 4

v1.02854; v1.07452; v1.01623

K09490

Heat shock 70 kDa protein 5

v1.14149; v1.14150

K09487

Heat shock protein 90 kDa beta

v1.07995; v1.07996; v1.16399; v1.11283

K09485

Heat shock protein 110 kDa

v1.08943; v1.05577

K11940

Heat shock protein HspQ

v1.00537; v1.00043

K03799

Heat shock protein HtpX

v1.01623

K04046

Hypothetical chaperone protein

v1.16216

K08268

Hypoxia-inducible factor 1 alpha

v1.08869; v1.15120

K09097

Hypoxia-inducible factor 1 beta

v1.22724

K09095

Hypoxia-inducible factor 2 alpha

v1.23698 [+ 16 other sequence copies]

K06711

Hypoxia-inducible factor prolyl 4-hydroxylase

v1.16737; v1.22345

K09486

Hypoxia up-regulated 1 (heat shock protein 70 family)

v1.10188

K08900

Mitochondrial chaperone BCS1

v1.17197; v1.04394

K04445

Mitogen-stress activated protein kinases

v1.16301; v1.21224; v1.19344

K04043

Molecular chaperone DnaK

v1.09682; v1.16748; v1.07471; v1.13624

K03687

Molecular chaperone GrpE

v1.01621; v1.04945; v1.15919

K04044

Molecular chaperone HscA

v1.18210

K04083

Molecular chaperone Hsp33

v1.17478; v1.16977; v1.10289; v1.19907

K04079

Molecular chaperone HtpG

v1.08895; v1.18099

K11416

Mono-ADP-ribosyltransferase sirtuin 6

v1.02024

K11411

NAD-dependent deacetylase sirtuin 1

v1.04813

K11412

NAD-dependent deacetylase sirtuin 2

v1.22049; v1.22211; v1.02221

K11413

NAD-dependent deacetylase sirtuin 3

v1.11849; v1.02221

K11414

NAD-dependent deacetylase sirtuin 4

v1.05495

K11415

NAD-dependent deacetylase sirtuin 5

v1.04868

K11417

NAD-dependent deacetylase sirtuin 7

v1.15070 [+ 4 other sequence copies]

K08835

Oxidative-stress responsive protein 1 (OXSR1)

v1.04503

K11875

Proteasome assembly chaperone 1

v1.01531

K11878

Proteasome assembly chaperone 4

v1.01210

K11879

Proteasome chaperone 1

v1.18611

K11880

Proteasome chaperone 2

v1.00599 [+ 29 other sequence copies]

K09553

Stress-induced-phosphoprotein 1 (HOP1)

v1.08830

K13057

Trehalose synthase (anhydrobiosis)

v1.22042

K03533

TorA specific chaperone

v1.16986 [+ 7 other sequence copies]

K06149

Universal stress protein A

Molecular chaperones are a diverse family of proteins expressed by both prokaryotic and eukaryotic organisms that serve to maintain correct protein folding in a 3-dimensional functional state, assist in multiprotein complex assembly and protect proteins from irreversible aggregation at synthesis and during conditions of cellular stress [267]. Additionally, heat shock proteins and their co-chaperones may regulate cell death pathways by inhibition of apoptosis [268]. The coral genome encodes a large number of DnaJ subfamily (J-domain) chaperones (Hsp40) that with co-chaperone GrpE (Table 10) regulates the ATPase activity of Hsp70 (DnaK in bacteria) to enable correct protein folding [269]. The coral genome encodes homologues of the molecular chaperones HscA (specialised Hsp70), the redox-regulated chaperone Hsp33, HtpG (high temperature protein G), members of the calnexin/calreticulin chaperone system of the endoplasmic reticulum, a mitochondrial chaperone BCS1 protein necessary for the assembly of the respiratory chain complex III and a specific chaperone of trimethyl N-oxide reductase (TorA). The coral genome also encodes hypoxia-inducible factors (HIFs) that moderate the deleterious effects of hypoxia on cellular metabolism (reviewed in [270]). In the HIF signalling cascade, the alpha subunits of HIF are hydroxylated at conserved proline residues by HIF prolyl-hydroxylases allowing their recognition for proteasomal degradation, which occurs during normoxic conditions but is repressed by oxygen depletion. Hypoxia-stabilised HIF1 upregulates the expression of enzymes principally of the oxygen-independent glycolysis pathway, and in higher animals promotes vascularisation, whereas the mammalian HIF2 paralogue regulates erythropoietin control of hepatic erythrocyte production in response to hypoxic stress [271]. The roles of HIF1 and HIF2 homologues in corals have been established, with HIF1 regulation of glycolysis critical to metabolic function during the dark diurnal anoxic state of coral respiration [193, 272].

Heat shock proteins that repair unfolded or misfolded protein have a complementary function to the ubiquitin-proteasome system (ubiquitins not tabulated) that selects damaged protein for degradation [273], such that HSP chaperones and the proteasome act jointly to preserve cellular proteostasis [274, 275]. Thus, several proteasome chaperones and assembly chaperones are encoded in the A. digitifera genome (Table 10). While proteasome chaperones serve to target aberrant proteins for ubiquination, the proteasome chaperones facilitates 20S assembly for biogenesis of the multiunit 26S proteasome that is activated in response to stress [276, 277], possibly by FtsJ (aka RrmJ), a well-conserved heat shock protein having novel ribosomal methyltransferase activity that targets methylation of 26S rRNA under heat shock control [278, 279]. The HspQ protein encoded in the coral genome, although studied almost exclusively in bacteria, is known to stimulate degradation of denatured proteins caused by hyperthermal stress, particularly DnaA that initiates DNA replication in prokaryotes [280]. Specifically, HspQ (heat shock factor sigma32) regulates the expression of Clp ATPase-dependent protease family enzymes [281, 282], of which ClpA, ClpB, ClipE, the protease adaptor protein ClpS [283] and the unfoldase ClpX protein [284] are encoded in the coral genome (Table 10). HspX is a small 16 kDa α-crystallin chaperone (Acr) protein belonging to the Hsp20 family of proteins [285] that suppresses thermal denaturation and aggregation of proteins [285]. Significantly, Acr proteins are known to bind with carbonic anhydrase [286] and may have importance in moderating stress-induced loss of calcium deposition. Thus, HspX/Acr expression may account for differences in the thermal sensitivity of corals to calcification that varies among genera [287]. In a different context, HspX is attracting considerable attention for its potential to elicit long-term protective immunity against human Mycobacterium tuberculosis infection by chaperoning a host-protective antigen [288] that by extension, but yet untested, may likewise repress virulence in the initiation and progression of microbial coral disease [289, 290].

The coral genome encodes complete membership of the human sirtuin (SIRT1-7) family of NAD(+)-dependent protein deacetylases and ADP-ribosyltransferases. Mammalian SIRT1 (a homologue of yeast Sir2) is an important regulator of metabolism, cell differentiation, stress response transcription and pathways of cellular senescence (reviewed in [291]). SIRT proteins regulate chromatin function through deacetylation of histones that promote subsequent alterations in the methylation of histones and DNA to affect, via deactivation of nuclear transcription factors and co-regulators, epigenetic control of nuclear transcription. As NAD+-dependent enzymes, SIRT1 can regulate gene expression in response to cellular NAD+/NADH redox status providing a metabolic template for epigenetic transcriptome reprogramming [292, 293]. In the human genome repertoire, SIRT1 modulates cellular responses to hypoxia by deacetylation of HIF1α [294] and inhibits nitric oxide synthesis by suppression of the nuclear factor-kappaB (NF-κB) signalling pathway [295], SIRT2 promotes oxidative stress resistance by deacetylation of forkhead box O (FOXO) proteins [296], SIRT3 decreases ROS production in adipocytes [297], SIRT4 regulates fatty acid metabolism and stress-response elements of mitochondrial gene expression [298], SIRT5 is a protein lysine desuccinylase and demalonylase of unknown function [299], SIRT6 activates base-excision repair [300] and SIRT7 inhibits apoptosis induced by oxidative stress by deacylation of p53 [301, 302]. The significance of coral SIRT proteins, by analogy, to exert stress tolerance is yet to be examined.

Metallochaperones are an important class of enzymes that transport co-factor metal ions to specific proteins [303]. The copper chaperone protein ATX1 (human ATOX1) delivers cytosolic copper to Cu-ATPase proteins and serves as a metal homeostasis factor to prevent Fenton-type production of highly reactive hydroxyl radicals. ATX1, which is strongly induced by molecular oxygen, functions additionally as an antioxidant to protect cells against the toxicity of both the superoxide anion and hydrogen peroxide [304]. Encoded also is a specific copper chaperone essential to the activation of Cu/Zn superoxide dismutase [305, 306] that is enhanced by photooxidative stress in scleractinian corals [307], although reported to be less pronounced in the host than in symbiotic algae [308]. In addition to high light exposure, reef-building corals of shallow reef flats are occasionally exposed to the atmosphere for periods that can last several hours during extreme low tides. Hence, species that are adapted to withstand acute desiccation (anhydrobiosis) have a better chance of surviving such conditions. The disaccharide trehalose is an osmolyte that in some plants and animals allows them to survive prolonged periods of desiccation [309]. The hydrated sugar has high water retention that forms a gel phase when cells dehydrate, which on rehydration allows normal cellular activity to resume without damage that would otherwise follow a dehydration/rehydration cycle. Furthermore, trehalose is highly effective in protecting enzymes in their native state from inactivation from thermal denaturation [310]. Given that A. digitifera is endemic on shallow reef flats prone to exposure at low tides [311], it is not surprising that the coral genome encodes trehalose synthase and a facilitated trehalose transporter for protection against dehydration.

Antioxidant and redox-protective proteins

Oxygen is vital for life, but it can also cause damage to cells, particularly at elevated levels. In coral symbiosis, the photosynthetic endosymbionts of corals typically produce more oxygen than the holobiont is able to consume by respiration, so that coral tissues are hyperoxic with tissue p O2 levels often exceeding 250% of air saturation during daylight illumination [193]. Furthermore, because algal symbionts reside within the endodermal cells of their host, coral tissues must be transparent to facilitate the penetration of downwelling light required for photosynthesis by their algal consorts. In clear shallow waters this entails concurrent exposure to vulnerable molecular sites of both partners to damaging wavelengths of ultraviolet radiation. The synergistic effects of tissue hyperoxia and UV exposure can cause oxidative damage to the symbiosis via the photochemical production of cytotoxic oxygen species [312] that are produced also during normal mitochondrial function [313]. Consequently, protective proteins (antioxidant enzymes) are expressed to maintain the fine balance between oxygen metabolism and the production of potentially toxic reactive oxygen species (ROS). If this balance is not maintained by regulation of oxidative and reductive processes (redox regulation), oxidative stress occurs by the generation of excess ROS, causing damage to DNA, proteins, and lipids. Corals elaborate a variety of molecular defences that including the production of UV-protective sunscreens (MAAS), antioxidants, antioxidant enzymes, chaperones and heat shock proteins, which are often inducible under conditions of enhanced oxidative stress [307], including conditions that elicit coral bleaching [314, 315]. An excellent review on the formation of ROS and the role of antioxidants and antioxidant enzymes in the field of redox biology is given by Halliwell [316].

Annotation of the A. digitifera genome reveals sequences encoding two isoforms of the antioxidant enzyme superoxide dismutase (SOD) from both the Cu/Zn and Fe/Mn families of SOD (Table 11). These metalloprotein enzymes catalyse the dismutation of superoxide to yield molecular oxygen and hydrogen peroxide, the latter being less harmful than superoxide. Superoxide can oxidize proteins, denature enzymes, oxidize lipids and fragment DNA. By removing superoxide, SOD protects also against the production of reactive peroxynitrite formed by the combination of superoxide and nitric oxide, which is a precursor reactant for production of the supra-reactive hydroxyl radical. Hydrogen peroxide per se is a mild oxidant, but it readily oxidises free cellular ferrous iron to ferric iron with production of hydroxyl radicals via the Fenton reaction. Accordingly, both the removal of hydrogen peroxide and the expression of proteins, such as transferrin, (bacterio)ferritins and metallothioneins, that bind reactive (transition) metal ions is important to protect cellular components from acute oxidative damage. Oddly, only a metallothionein expression activator was found encoded in the coral genome without finding a sequence to activate transcription of the actual metallothionein protein gene.
Table 11

Antioxidant and redox-protective proteins in the predicted proteome of A. digitifera

Gene sequence

KEGG Orthology

Encoded protein description

v1.10918

K04756

Alkyl hydroperoxide reductase subunit D

v1.11551

K03387

Alkyl hydroperoxide reductase subunit F

v1.07812

K03594

Bacterioferritin

v1.21362 [+ 4 other sequence copies]

K00429

Catalase (bacterial)

v1.17525 [+ 4 other sequence copies]

K03781

Catalase (peroxisonal)

v1.23457; v1.01713; v1.19228

K04569

Copper chaperone for superoxide dismutase

v1.20153; v1.20154

K10528

Hydroperoxide lyase

v1.19687; v1.19688; v1.18796; v1.18795

K00522

Ferritin heavy chain

v1.06441

K03674

Glutaredoxin 1

v1.19449

K03675

Glutaredoxin 2

v1.14929 [+ 5 other sequence copies]

K03676

Glutaredoxin 3

v1.13285; v1.03722; v1.03688; v1.10496

K00432

Glutathione peroxidase

v1.13174; v1.13775; v1.05473

K00383

Glutathione reductase (NADPH)

v1.14344; v1.19399; v1.01421

K01920

Glutathione synthase

v1.02173

K09238

Metallothionein expression activator

v1.09719; v1.16134; v1.18608

K07390

Monothiol glutaredoxin

v1.14890; v1.17685

K07305

Peptide-methionine (R)-S-oxide reductase

v1.14909

K00435

Peroxiredoxin

v1.14106

K13279

Peroxiredoxin 1

v1.08691

K11187

Peroxiredoxin 5, atypical 2-Cys peroxiredoxin

v1.01410

K11188

Peroxiredoxin 6, 1-Cys peroxiredoxin

v1.03688

K05361

Phospholipid-hydroperoxide glutathione peroxidase

v1.05148

K05905

Protein-disulfide reductase

v1.02922; v1.22772; v1.24164

K05360

Protein-disulfide reductase (glutathione)

v1.06810

K12260

Sulfiredoxin

v1.01713 [+ 4 other sequence copies]

K04565

Superoxide dismutase, Cu/Zn family

v1.09974; v1.20324

K04564

Superoxide dismutase, Fe/Mn family

v1.02378

K11065

Thiol peroxidase, atypical 2-Cys peroxiredoxin

v1.22324 [+ 7 other sequence copies]

K03671

Thioredoxin 1

v1.05148; v1.03230; v1.20699

K03672

Thioredoxin 2

v1.17881 [+ 5 other sequence copies]

K13984

Thioredoxin domain-containing protein 5

v1.04532; v1.24501

K09585

Thioredoxin domain-containing protein 10

v1.11551; v1.19049

K00384

Thioredoxin reductase (NADPH)

v1.10930

K14736

Transferrin

As expected from the foregoing, the genome of A. digitifera encodes the antioxidant enzyme catalase (CAT) that is highly efficient in decomposing hydrogen peroxide to yield molecular oxygen and water. Two isoforms of CAT are encoded at multiple sites. One is a peroxisomal eukaryotic CAT enzyme that targets the removal of hydrogen peroxide formed as a by-product of oxidase enzymes, and the other is a related catalase domain-containing protein presumed also to decompose hydrogen peroxide. Glutathione peroxidise (GPx) reduces both hydrogen peroxide and lipid hydroperoxides, the latter of which are formed by radical-induced lipid autoxidation. Phototrophic organisms, including higher plants, utilise ascorbate peroxidase (APx) as a primary catalyst for the reduction of hydrogen peroxide and lipid hydroperoxides. However, unlike the freshwater cnidarian H. viridis[164], there is no evidence for transfer of APx-encoding genes to A. digitifera. The antioxidant enzymes SOD, CAT, GPx and APx are well characterised in the algal and animal partners of coral symbiosis (reviewed in [317]). Additionally, the coral genome has sequences encoding alkyl hydroperoxide reductase, hydroperoxide lyase, phospholipid-hydroperoxide glutathione peroxidase, thiol peroxidase and multiple isoforms of peroxiredoxin, all of which function in the detoxification of organo-hydroperoxides that are produced as a by-product of aerobic metabolism. Additionally, sulfiredoxin (Table 11) repairs peroxiredoxins when these enzymes are inhibited by over-oxidation [318].

Thioredoxins and glutaredoxins have important secondary roles in regulating multiple pathways in many biological processes, including redox signalling of apoptotic pathways, which have been attributed to processes involved in coral bleaching [56]. Other enzymes that regulate cellular thiol-disulfide homeostasis in this coral are monothiol glutaredoxin and protein-disulfide reductase. The coral genome encodes the ubiquitous thioredoxin system of antioxidant proteins (Table 11) that act as electron donors to peroxidases and ribonucleotide reductase (the latter not tabulated). By cysteine thiol-disulfide exchange, thioredoxins function as a protein thiol-disulfide oxidoreductase [319]. In the thioredoxin system, thioredoxins are maintained in their reduced state by NADPH-dependent, flavoenzyme thioredoxin reductase [320]. Peptide-methionine (R)-S-oxide reductase can additionally rescue thioredoxin from oxidative inactivation by disulfide reduction. Related glutaredoxins share many of the functions of thioredoxins but are reduced directly by glutathione, rather than by a specific reducing enzyme, while in turn glutathione is kept in its native state by NADPH: glutathione reductase.

In recent years there has been a particular focus on the role of ROS in coral bleaching, fuelled by dire prediction of future catastrophic episodes caused by environmental change affected by global warming [321]. Early predictions of coral bleaching were based principally on physical environmental parameters, rather than on the determination of the physiological state of coral populations to such conditions. While gene expression markers are being developed to monitor sub-bleaching levels of stress in situ (e.g., [261]), Kenkel et al. [322] opined that the current challenge for implementing expression-based methods lies in identifying coral genes demonstrating the most pronounced and consistent stress response, preferably with a large dynamic range to enable reliable quantification. To this end, we offer in Table 11 the annotation of novel redox-related genes for examination as potential candidate biomarkers to monitor the physiological response of A. digitifera to environmental stress.

Proteins of cellular apoptosis

Apoptosis is the signalling of programmed cell death (PCD) that occurs in multicellular organisms in response to cellular injury. A key feature of apoptosis is the activation of endogenous endonucleases causing nuclear fragmentation, chromatin condensation and chromosomal DNA fragmentation, which typically presents in affected cells by the morphological appearance of plasma membrane blebbing and cell shrinkage. Caspases and related family member proteases are described as “executioners” of apoptosis that on post-translational activation degrade the regulatory proteins that prevent DNA degradation. Fragmentation of nuclear DNA is one of the hallmarks of apoptotic cell death that occurs by PCD stimuli in a wide variety of proliferating cells. NF-κB is a protein complex that controls the transcription of DNA that can induce the expression of nitric oxide synthesis (NOS) to produce NO that is a well-known promoter of the of the pro-apoptotic transcription factor p53 cell-cycle gatekeeper of the caspase cascade. In contrast to necrosis, which is the outcome of PCD, apoptosis mediates the fragmentation of damaged cells, which by phagocytosis are removed or degraded in phagolysosomes to spare surviving cells from the uncontrolled release of cytotoxic agents. Proteins of the caspase-mediated apoptotic cascade are regarded as products of constituent housekeeping genes that are necessary to maintain healthy multicellular function [323]. In the progression of cnidarian bleaching, apoptotic pathways are activated [322325], but not all corals that suffer bleaching are destined to die [326, 327]. Coral survival has been attributed to having a high level of apoptotic protection at the onset of coral bleaching [328] and during post-bleaching recovery [329] by specific activation of anti-apoptotic Bcl-2 proteins in surviving cells [330].

Cnidarians have a complex apoptotic protein network that has exceptional ancestral complexity and is comparable to that of higher vertebrates [331, 332]. Cnidarian metamorphosis is tightly coupled with caspase-dependent apoptosis [333] and subsequent host-symbiont selection by post-phagocytic winnowing of Symbiodinium genotypes during the establishment of coral-dinoflagellate mutualism [334]. As expected, the coral genome of A. digitifera encodes multiple isoforms of genes that transcribe the caspase family of apoptotic effectors (Table 12). Included in this signalling pathway are the pro- and anti-apoptotic Bax/Bcl regulators and Bcl-2 athanogene (DNA-binding) activators of apoptosis. Notable in our annotation dataset are multiple genes that encode the protein domains of the apoptotic protease-activating factor (Apaf) that triggers assembly of the apoptosome leading to caspase activation [335]. Additional to this arsenal of cell cycle regulators are the death associated protein-6 (DAXX), a Fas-binding adaptor of c-Jun N-terminal kinase (JNK) activation [336], death-associated protein kinase (DAPK), a mediator of calcium/calmodulin-regulated Ser/Thr kinase [337], and the programmed cell death 6-interacting protein (PDCD6IP), which binds to PDCD-6 for execution of apoptosis via the caspase-3 pathway [338]. PDCD6IP activation of apoptosis is an enigma since PDCD-6 is not encoded in the coral genome, nor is caspase-3. Other cell cycle regulators are the p53 binding and p53-associated parkin-like proteins, and the activating TP53 regulating kinase protein and TP53 apoptosis effector of TP53 gene expression.
Table 12

Proteins of cellular apoptosis in the predicted proteome of A. digitifera

Gene sequence

KEGG Orthology

Encoded protein description

v1.17521; v1.02505; v1.20702; v1.05077

K02159

Apoptosis regulator BAX (BCL2-associated)

v1.05086; v1.20659

K02161

Apoptosis regulator BCL-2

v1.17522; v1.00181; v1.10817; v1.20703

K02163

Apoptosis regulator BCL-W

v1.05147 [+ 6 other sequence copies]

K12875

Apoptotic chromatin condensation inducer

v1.22264 [+ 72 other sequence copies]

K02084

Apoptotic protease-activating factor (Apaf)

v1.17326; v1.20305; v1.11586

K09555

BCL2-associated athanogene 1

v1.08601

K09558

BCL2-associated athanogene 4

v1.02839

K09559

BCL2-associated athanogene 5

v1.01518

K13087

BCL2-associated transcription factor 1

v1.20278; v1.00172; v1.07858

K14021

BCL-2 homologueous antagonist/killer

v1.09624

K02561

BCL2-related (ovarian) killer protein

v1.17749

K08573

Calpain-3

v1.00595; v1.14671; v1.00040

K08574

Calpain-5

v1.00040

K08575

Calpain-6

v1.19153; v1.17749

K08576

Calpain-7

v1.15226

K04740

Calpain-12

v1.02951

K08582

Calpain-15

v1.11167; v1.06681; v1.20230; v1.01376

K08585

Calpain, invertebrate

v1.0312 7 [+ 6 other sequence copies]

K08583

Calpain, small subunit 1

v1.17229; v1.00023; v1.09976

K02186

Caspase 2

v1.11989 [+ 5 other sequence copies]

K04397

Caspase 7

v1.02756 [+ 27 other sequence copies]

K04398

Caspase 8

v1.01818

K04399

Caspase 9

v1.00817 [+ 4 other sequence copies]

K04400

Caspase 10

v1.02005

K04741

Caspase 12

v1.00818 [+ 11 other sequence copies]

K04489

Caspase apoptosis-related cysteine protease

v1.13260

K07367

Caspase recruitment domain-containing protein 11

v1.06297 [+ 44 other sequence copies]

K02832

CASP2 and RIPK1 adaptor with death domain

v1.21531

K02308

Death-associated protein 6 (DAXX)

v1.09448; v1.15529; v1.20164

K08803

Death-associated protein kinase (DAPK)

v1.23110; v1.14222; v1.03658

K12366

Engulfment and motility protein 1 (phagocytosis/apoptosis)

v1.18448 [+ 78 other sequence copies]

K02373

Fas (TNFRSF6)-associated via death domain (FADD)

v1.24288 [+ 66 other sequence copies]

K10130

Leucine-rich repeats and death domain-containing protein

v1.20620

K04734

NF-kappa-B inhibitor alpha

v1.01706

K14214

NF-kappa-B inhibitor delta

v1.10378; v1.10729; 1.05609; v1.05609

K05872

NF-kappa-B inhibitor epsilon

v1.17893; v1.22419; v1.00700; v1.08415

K09256

NF-kappa-B inhibitor-like protein 1

v1.04158 [+ 211 other sequence copies]

K09257

NF-kappa-B inhibitor-like protein 2

v1.05320; v1.06979; v1.04467; v1.21371

K02580

Nuclear factor NF-kappa-B p105 subunit

v1.20334; v1.22743

K11970

p53-Associated parkin-like cytoplasmic protein

v1.14920; v1.11864; v1.15271; v1.11865

K06643

p53-Binding protein

v1.04289

K06708

Programmed cell death 1 ligand 2

v1.05882 [+ 7 other sequence copies]

K12200

Programmed cell death 6-interacting protein (PDCD6!P)

v1.10959; v1.04994

K04727

Programmed cell death 8 apoptosis-inducing factor

v1.16714

K06875

Programmed cell death protein 5 (PDCD-5)

v1.13112

K03171

Tnfrsf1a-associated via death domain

v1.24655; v1.12385

K10136

TP53 apoptosis effector

v1.09087

K08851

TP53 regulating kinase

v1.05030; v1.07044

K11859

Tumor necrosis factor, alpha-induced protein 3

v1.22799

K04389

Tumor necrosis factor ligand superfamily member 6

v1.05776

K05470

Tumor necrosis factor ligand superfamily member 7

v1.13754

K05472

Tumor necrosis factor ligand superfamily member 9

v1.21776 [+ 6 other sequence copies]

K04721

Tumor necrosis factor ligand superfamily member 10

v1.04001

K05473

Tumor necrosis factor ligand superfamily member 11

v1.19776

K05474

Tumor necrosis factor ligand superfamily member 12

v1.09015; v1.14041

K03158

Tumor necrosis factor receptor superfamily member 1A

v1.07010

K05141

Tumor necrosis factor receptor superfamily member 1B

v1.19735

K05142

Tumor necrosis factor receptor superfamily member 4

v1.13754

K03160

Tumor necrosis factor receptor superfamily member 5

v1.22577

K05143

Tumor necrosis factor receptor superfamily member 6B

v1.20003

K05144

Tumor necrosis factor receptor superfamily member 7

v1.23750; v1.17970; v1.19022

K05146

Tumor necrosis factor receptor superfamily member 9

v1.07527

K05148

Tumor necrosis factor receptor superfamily member 11B

v1.10221

K05151

Tumor necrosis factor receptor superfamily member 13C

v1.14826; v1.01054

K05152

Tumor necrosis factor receptor superfamily member 14

v1.09514

K05156

Tumor necrosis factor receptor superfamily member 19

v1.01640

K05161

Tumor necrosis factor receptor superfamily member 26

v1.08207; v1.16237; v1.14824

K10133

Tumor protein p53-inducible protein 3

Our genome annotation reveals 73 sequence matches for expressing the Apaf protein domain that, in conjunction with a high copy number for expressing caspase-8 (28 protein sequence matches), may enhance coral survival during embryogenesis by suppressing receptor-induced protein kinase (45 sequence matches) during early development [339]. The most conserved function of the CAPS2/RIPK adaptor (45 sequence matches) encoded in the coral genome is its essential regulation of apoptosis [340]. We find a wide repertoire of genes that additionally encode proteins that mediate apoptosis (Table 12). Amongst these are the calpain Ca+2-sensing family of proteins that initiate the signalling of apoptotic pathways [341]. There are 79 matches to sequences that encode the tumor necrosis Fas superfamily member 6 (TNFRSF6) receptor, which coupled with the death domain (FADD) protein is a cell signalling mediator for recruitment of caspase-8 that activates the apoptotic cysteine protease cascade. Coincident in the genome are 67 sequences encoding the leucine-rich repeat and death domain-containing (LRDD) adaptor that, by interacting with other p53-inducible death domain-containing (PIDD) proteins such as FADD, induces the caspase-2 pathway of apoptosis in response to DNA damage [342]. Elements of the NF-κB signalling pathway of cnidarians are highly conserved traits [343], which includes the caspase cascade and the pro-apoptotic and anti-apoptotic Bcl-2 family of proteins [344]. The coral genome of A. digitifera encodes the pleiotropic nuclear factor NF-κB p105 subunit, and astonishingly there are 212 sequence matches to the NF-κB inhibitor-like protein 2 domain with fewer matches to the NF-κB inhibitor-like protein 1 and NF-κB family inhibitors alpha, delta and epsilon. Evident in our genome annotation is the tumor necrosis factor-alpha induced protein 3 (TNFAIP3), a cytokine produced by activated (inflammatory) macrophages. Although TNF cytokines are a major extrinsic mediator of cellular apoptotic pathways, the precise function of the superfamily members of TNF ligands and receptors (Table 12) remains elusive in coral symbiology.

Microbial symbiosis and pathogenicity

It is well established that corals associate with a vast consortia of microbes, including phototrophic symbionts (Symbiodinium spp.) and other eukaryotic microbionts, cyanophytes, heterotrophic bacteria, archaea and viruses [345]. Corals harbour diverse and abundant prokaryotic communities with distinct populations residing in separate habitats of the host skeleton, tissues and surface mucus layer (reviewed in [203]). Microbial populations are dominated by a few coral-specific taxonomic traits [346], but the majority of the population comprises a high number of taxonomically diverse, low-abundance ribotypes [347] with much of the diversity within the coral microbiome belonging to the “rare” biosphere [348, 349]. The coral microbiome is vital to the nutrition and health of the holobiont [350] and contributes significantly to the protection of coral reef ecosystems against the detrimental effects of organic enrichment [351, 352]. One emerging threat to coral reefs is the outbreak of infectious diseases (reviewed in [353]). Although highly subjective and with little experimental evidence to date, the coral probiotic hypothesis [354] suggests that the coral prokaryotic microbiome can adapt to changing environmental conditions by selective microbial reorganisation to impart greater resistance to disease and pathogen-mediated bleaching [355]. Whether the coral microbiome can respond to changing environmental conditions more rapidly than by host genetic mutation and selection based on contemporary phenotypic evolution on ecological time-scales [356], is a topic of current debate [357].

Corals, like other invertebrates, have an innate immune system based on self-histocompatibility recognition (reviewed in [358]), but to date few adaptive components have been identified [359]. Corals do not produce antibodies and thus lack a true adaptive immune system. Nonetheless, corals once susceptible to infection and bleaching caused by a specific bacterial agent can become immune to the invading pathogen by a phenomenon termed “experience-mediated tolerance”, a precept of the hologenome theory of evolution [360], although how this process occurs is largely unknown. In our annotation of the genome sequence of A. digitifera we uncovered genes encoding the expression of disease resistance proteins (Table 13), two of which match the plant RPM1 and RPS2 pathogen resistance proteins that guard against disease by binding with pathogen avirulence receptors [360, 361]. Significant also is a gene to express the pathogenesis-related protein PR-1 (29 sequence domain matches) that is inducible in plants for systemic acquired resistance to pathogenic invasion [362]. We uncovered also multiple genes encoding the expression of myeloperoxidase (MPO) enzymes. MPOs produce hypochlorous acid from hydrogen peroxide and chloride ion (requiring heme as a cofactor), and it oxidizes tyrosine to the tyrosyl radical using hydrogen peroxide as an oxidizing agent. Hypochlorous acid and tyrosyl radicals are strong cytotoxic agents that in higher organisms are used as a primary defence by neutrophils to protect against invading pathogens. Phenoloxidase (tyrosinase) activity is reported to contribute to the innate defence system of A. millepora and Porites sp. [363] via activation of the melanin-signalling pathway that is induced in response to coral bleaching and localised disease [364, 365]. Three genes of A. digitifera encode tyrosinase enzymes (data not tabulated) to account for the phenoloxidase activity reported in corals.
Table 13

Microbial symbiosis and pathogenicity proteins in the predicted proteome of A. digitifera

Gene sequence

KEGG Orthology

Encoded protein description

v1.06126

K13061

Acyl homoserine lactone synthase

v1.19990

K01372

Bleomycin hydrolase

v1.00209; v1.06178

K03587

Cell division protein FtsI (penicillin-binding protein 3)

v1.18860

K13458

Disease resistance protein

v1.16231; v1.00374; v1.08191

K13457

Disease resistance protein RPM1

v1.13482 [+ 4 other sequence copies]

K13459

Disease resistance protein RPS2

v1.07889

K12090

Cag pathogenicity island protein 5

v1.24345

K12091

Cag pathogenicity island protein 6

v1.18924; v1.17622

K12093

Cag pathogenicity island protein 8

v1.05278

K12096

Cag pathogenicity island protein 11

v1.02083

K12104

Cag pathogenicity island protein 19

v1.12907

K12109

Cag pathogenicity island protein 24

v1.00209; v1.06178

K03587

Cell division protein FtsI (penicillin-binding protein 3)

v1.13874

K07259

Carboxy/endopeptidase (penicillin-binding protein 4)

v1.12514; v1.09758

K04127

Isopenicillin-N epimerase

v1.21332

K04126

Isopenicillin-N synthase

v1.07742

K02547

Methicillin resistance protein

v1.17478; v1.16977; v1.10289; v1.19907

K04079

Molecular chaperone HtpG (anti-bacterial)

v1.08255

K13651

Motility quorum-sensing regulator, GCU-specific toxin

v1.14792 [+ 7 other sequence copies]

K10789

Myeloperoxidase

v1.02333 [+ 26 other sequence copies]

K13449

Pathogenesis-related protein 1

v1.05017

K03693

Penicillin-binding protein

v1.17507

K12556

Penicillin-binding protein 2X

v1.13874

K07259

Penicillin-binding protein 4

v1.16655

K02171

Penicillinase repressor

v1.14688

K15126

Type III secretion system cytotoxic effector protein

v1.20647

K03980

Virulence factor, integral membrane protein

v1.18964

K03810

Virulence factor, oxidoreductase domain

The genome of A. digitifera also reveals homologues of genes that promote bacterial pathogenicity (Table 13), including virulence factors that are expressed and excreted by invading pathogens (bacteria, viruses, fungi and protozoa) to inhibit certain protective functions of the host. Such are the bacterial Type III cytotoxic effector protein and multiple Type IV Cag pathogenicity island proteins encoded in the coral genome. Many Gram-negative bacteria utilize Type III secretion proteins, which are regulated by quorum sensing, to deliver cytotoxic effector proteins into eukaryote host cells during infection. Cag (cytotoxin-associated) pathogenicity island (PAI) proteins are encoded by mobile genetic elements of the Type IV system secreting both proteins and large nucleoprotein complexes [366] that may be transferred between prokaryotes to enhance selected traits of virulence [367]. Our annotation reveals genes encoding six pathogenicity island proteins (Table 13) with similarity to the Cag PAI proteins of the human Heliobacter pylori, an infectious bacterium causing peptic ulcers that may lead to the development of stomach cancer. While many properties of Type III and IV secretion system proteins have been well characterized in bacteria, the functional purpose of homologous genes in A. digitifera, if expressed, are unknown.

The genome of A. digitifera contains genes of bacterial origin that encode the motility quorum-sensing regulator of the GCU-specific mRNA interferase toxin and acyl homoserine lactone synthesis used for the communication of quorum sensing between bacteria to enable the coordination of group behaviour based on collective population density. Apparent in our annotation (Table 13) is a wide array of microbial penicillin-binding proteins (PBPs) that have an affinity for β-lactam antibiotics that by binding to PBPs prevent bacteria from constructing a cell wall. There are genes also to enhance antibiotic resistance, including potential expression of a penicillinase repressor, a methicillin resistance protein and bleomycin hydrolase (cysteine peptidase). Additionally, isopenicillin-N synthase and an isopenicillin-N epimerase, both of which catalyse key steps in the biosynthesis of penicillin and cephalosporin antibiotics, are encoded in the coral genome. Taken as a whole, we demonstrate an extensive presence of ancient non-metazoan genes that are maintained in the genome of A. digitifera, as is reported in the genomes of A. millepora and the anemone N. vectensis[368]. Recent thought on genome evolution places these ancestral conserved domains as ‘orphan’ or ‘taxonomically restricted’ genes [352, 369, 370], rather than acquired later by horizontal gene transfer. There is, of course, little knowledge of how or when, if at all, these non-metazoan genes are expressed or even their function to mediate pathogenicity in the coral holobiont.

Proteins of viral pathogenicity

Marine viruses were of minor interest until 1989, when it was realised that virus-like particles (VLPs) are the most abundant biological entities to occupy aquatic environments with variable numbers reaching ~108 VLPs ml-1[371]. Typically, VLPs surpass the number of marine bacteria by an order of magnitude in coastal waters [372]; their diversity is extremely high and many are specific to the marine environment [373, 374]. Significant VLP numbers are reported from the surrounding waters of oceanic coral reef atolls [375], in waters flowing across the reef substratum [376] and in samples taken within the close vicinity of coral colonies [377, 378]. The viral load within the surface microlayer of scleractinian corals is enumerated as being 107-108 VLPs mL-1[379] and, based on VLP morphological diversity, is attributed to infecting various microbial hosts (bacteria, archaea, cyanobacteria, fungi and algae) residing within the coral mucus [380]. VLPs have been observed in the epidermal and gastrodermal tissues of corals and occasionally occur in the mesogloea [381]. Latent viruses were found to infect Symbiodinium isolated from several scleractinian corals [382384] with a preponderance of eukaryotic algae-infecting phycodnaviruses suggested [385]. A wide range of bacteriophage and eukaryotic virus families have been identified within scleractinians using metagenomic analyses [207, 386388], with bacteriophages being by far the most abundant entities (Wood-Charlson EM, Weynberg KD, Suttle CA, Roux S, van Oppen MJH: Methodological biases in coral viromics, submitted).

The importance of the coral-virus interactome in bleaching and disease (reviewed in [185, 389]) is founded on reports showing that VLP abundances are higher in the seawater immediately surrounding diseased compared to that of healthy corals [378], that latent viruses are induced by heat stress in symbiotic dinoflagellates of the sea anemone Anemonia virdis[382] and the coral Pavona danai[383], and that UV exposure induces a latent virus-like infection in cultured Symbiodinium[187]. Quantitative 454 pyrosequence analysis of the coral Porites compressa on exposure to reduced pH, elevated nutrients or thermal stress showed that the abundance of its viral consortia varied across treatments, but notably a novel herpes-like virus increased by up to 6 orders of magnitude on exposure to abiotic stress [387], although some caution may be warranted in assessing the reliability of such determinations [Wood-Charlson et al., submitted]. Unexpectedly, the proteome of an endosymbiont-enriched fraction of the coral Stylophora pistillata showed a significant 114-fold increase in a viral replication protein on thermal bleaching [39], which is consistent with the finding of VLP induction in P. compressa by similar treatment [387].

General aspects of histocompatibility [390393] and the genetic structure of innate immune receptors of the Cnidaria [363, 394401], including the immune response effected by coral disease and bleaching [364, 402], have been examined extensively, hence further elaboration here is unnecessary. Instead, we focus on proteins that directly regulate the pathogenicity of coral-associated microbes and viruses. The A. digitifera genome encodes protein homologues having either putative antiviral and virus-promoting activities (Table 14). These homologues include the antiviral “superkiller” helicase SKI2 protein that acts by blocking viral mRNA translation [403] and, together with the superkiller proteins SKI3 (69 sequence alignments) and SKI8 of the exosome complex, function in a 3′-mRNA degradation pathway [404]. The coral genome encodes also three exoribonuclease (RNase) enzymes (XRN, XRN2 and RNB) with antiviral RNA-degrading properties [405, 406]. Annotation of the coral genome reveals homologues to four interferon proteins (IFNB, IFNG, IFNW1 and IFNT1). Interferons are potent and selective antiviral cytokines [407], which are induced by viral infection or by sensing dsRNA, a by-product of viral replication, leading to the transcription of interferon-stimulated genes whose products have antiviral activities and others having antimicrobial, antiprolifera-tive/antitumor or immumomodulatory effects [408, 409]. Included in the coral antivirus defence system are three members of the interferon regulatory transcription factor (IRF1, IRF2 and IRF8) family proteins. IRF1 and IRF2 are transcriptional activators of cytokines and other target genes [410]; IRF1 is known to trans-activate the tumor suppressor protein p53 [411] while IRF2 regulates post-transcriptional induction of NO synthase [412]. Conversely, IRF8 is an interferon consensus sequence-binding protein that is a negative (interference) regulator of enhancer elements common to interferon-inducible genes [413]. The coral genome additionally includes an interferon-stimulated 20 kDa protein (ISG20) RNase specific to deactivation of singled-stranded RNA viruses [414]. The coral genome encodes several interferon-inducible proteins, notably interferon gamma induced GTPase (IGTP) that accumulates in response to IFNB [415], the interferon-induced GTP-binding protein Mx1 that is a key element of host antiviral defence [416], the interferon-induced helicase C domain-containing protein1 (aka MDA-5), which is an immune receptor that senses viral dsRNA to activate the interferon antiviral-response cascade [417] and the interferon-induced transmembrane protein (IFITM1) that suppresses cell growth [418]. The coral genome encodes the interferon-gamma receptor 2 (IFNGR2) transmembrane protein that activates downstream signal transduction cascades that control cell proliferation and apoptosis [419]. Encoded also is a homologue of the human bone marrow stromal cell antigen 2 (BST2) that inhibits retrovirus infection by preventing VLP release from infected cells [420]. Additionally encoded is a mitochondrial antiviral-signalling protein (MAVS) that triggers the host immune response by activation of the nuclear transcription factor NF-kB and the interferon regulatory transcription factor IRF3 which coordinates the expression of type-1 interferons such as IFNB [421].
Table 14

Regulatory and related proteins of viral pathogenicity in the predicted proteome of A. digitifera

Gene sequence

KEGG Orthology

Encoded protein description

v1.20647; v1.06188; v1.21287

K12599

Antiviral helicase SKI2

v1.18443 [+ 40 other sequence copies]

K12807

Baculoviral IAP repeat-containing protein 1 (BIRC1)

v1.06263 [+ 6 other sequence copies]

K04725

Baculoviral IAP repeat-containing protein 2/3/4 (BIRC2/3/4)

v1.14355

K08731

Baculoviral IAP repeat-containing protein 5 (BIRC5)

v1.04171 [+ 7 other sequence copies]

K10586

Baculoviral IAP repeat-containing protein 6 (BIRC6)

v1.12348; v1.01945; v1.16612

K06731

Bone marrow stromal cell antigen 2 (antiviral BST2)

v1.01539 [+ 7 other sequence copies]

K04012

Complement component receptor 2 (CR2)

v1.17305

K04462

Ecotropic virus integration site 1 protein (EVI1)

v1.1496 [+ 4 other sequence copies]

K12618

5′-3′ Exoribonuclease 1 (antiviral XRN1)

v1.22746; v1.19002; v1.12850; v1.21216

K12619

5′-3′ Exoribonuclease 2 (antiviral XRN2)

v1.09005

K01147

Exoribonuclease II (antiviral RNB)

v1.22793; v1.12978; v1.19008; v1.20838

K09239

HIV virus type I enhancer-binding protein (HIVEP)

v1.02776 [+ 7 other sequence copies]

K15046

Influenza virus NS1A-binding protein (NS1A-BP)

v1.09829; v1.13077

K05415

Interferon beta (IFNB)

v1.11946; v1.21512; v1.11221; v1.11927

K04687

Interferon gamma (IFNG)

v1.21512

K14140

Interferon gamma induced GTPase (ITGP)

v1.11946

K05133

Interferon gamma receptor 2 (IFNGR2)

v1.01539 [+ 4 other sequence copies]

K04012

Interferon-induced GTP-binding protein Mx1

v1.10782; v1.23797; v1.17119; v1.03221

K12647

Interferon-induced helicase C domain-containing protein 1

v1.06274; v1.15849; v1.05943

K06566

Interferon induced transmembrane protein (IFITM1)

v1.21327; v1.24081

K05440

Interferon, omega 1 (IFNW1)

v1.11817

K09444

Interferon regulatory factor 1 (IRF1)

v1.11816; v1.07639

K10153

Interferon regulatory factor 2 (IRF2)

v1.11421

K10155

Interferon regulatory factor 8 (IRF8)

v1.02158

K12579

Interferon-stimulated gene 20 kDa protein (ISG20)

v1.15947

K05442

Interferon tau-1 (IFNT1)

v1.22825; v1.08034; v1.08520

K05788

Integration host factor subunit beta (IHFB)

v1.14899

K08220

MFS transporter, FLVCR family virus subgroup C receptor

v1.04514; v1.04513; v1.16929

K12648

Mitochondrial antiviral-signalling protein (MAVS)

v1.17718; v1.08002; v1.08001; v1.22382

K06081

Poliovirus receptor-related protein 1 (PVRL1)

v1.21413; v1.06637

K06531

Poliovirus receptor-related protein 2 (PVRL2)

v1.11740; v1.21467; v1.11410; v1.17135

K06592

Poliovirus receptor-related protein 3 (PVRL3)

v1.15077

K06593

Poliovirus receptor-related protein 4 (PVRL4)

v1.04158 [+ 68 other sequence copies]

K12600

Superkiller protein 3 (antiviral SHI3)

v1.18238 [+ 4 other sequence copies]

K12601

Superkiller protein 8 (antiviral SHI8)

The coral genome encodes a full set of baculoviral IAP repeat-containing proteins BIRC 1-6 (Table 14). The IAP (inhibitor of apoptosis) family proteins were first identified secreted by baculovirus to protect infected cells from death in the progression of viral replication [422]. Expressed by most eukaryotic organisms (reviewed in [423]), their IAP function is presumably conserved in corals. The coral genome encodes a full set of poliovirus receptor-related proteins (PVRL1-4) of the immunoglobulin superfamily, which bind and transport herpesviruses at the cellular membrane in the establishment of latent infections (reviewed in [424]). Encoded also is a complement component (3d/Epstein Barr virus) receptor 2 (CR2) protein that binds to the Epstein-Barr virus Herpes viridae with antigenic activity for disease prevention [425]. Another encoded protein is a homologue of the human immunodeficiency virus type 1 (HIV-1) enhancer-binding protein (HIVEP; aka EBP1) that attaches to the HIV long terminal repeat (LTR) region to activate transcription via the HIV LTR [426]. Present in the coral genome is also a homologue of the influenza virus non-structural binding protein NS1A-BP that interacts with the NS1 virulence factor of the influenza A virus Orthomyxoviridae to interfere with NS1-inhibition of pre-mRNA splicing within the host nucleosome [427]. NS1A-BP inhibits NS1A-mediated disruption of the host immune response caused by restricting interferon production and the antiviral effects of IFN-induced proteins [428]. The genome of A. digitifera encodes an integration host factor subunit beta (IHFB), first discovered as a host factor for bacteriophage λ integration of mobile genetic elements, that in E. coli is involved in multiple processes of DNA replication, site-specific recombination and gene expression [429]. A homologue of the MFS transporter feline leukemia virus subgroup C receptor (FLVCR) cell surface protein is encoded in the coral genome, which in cats confers susceptibility to FeLV-C infection [430]. Encoded also is a viral integration site 1 (EVI1) that in humans is an oncogenic transcription factor, often activated by viral infection, to cause proliferation of invasive tumours [431]. Arguably, these homologue proteins typically expressed in such distantly related species may have similar relevance in viral interactions of the coral holobiome.

How these regulatory proteins and viral receptors interact and respond to viral infection in corals is yet to be realised. The absence of virion-specific sequences (e.g. for nucleic acid replication or capsid structure) suggests that proviral DNA is absent from the coral genome, or it may be an artefact of the limited number of marine viral sequences deposited in public databases. Discovery of viral activity through proteomics [39] may, therefore, suggest that viral proteins are synthesised from a lytic infection, but this requires confirmation.

Toxins and venom

A review of protein sequences deposited in the UniProt database in October 2012 shows that there are 150 known cnidarian toxins. These toxins have diverse biological activities (neurotoxins, pore-forming cytolysins and venom phospholipases) used to capture prey and for protection against predators [432] that are best characterised in sea anemones (Actiniaria) with 141 sequences deposited [433, 434]. The cytotoxin MCTx-1 isolated from the Net Fire Coral Millepora dichotoma is the only toxin from a coral deposited in Uniprot (accession number A8QZJ5). However, our initial examination of the predicted proteome of A. digitifera shows 18 proteins with similarity to bacterial toxins and associated regulatory proteins (Table 15). Unlike reports from proteomic examination of the coral S. pistillata[39] and nematocysts (stinging organelles) of the jellyfish Olindias samba-quiensis[435], Tamoya haplonema, Chiropsalmus quadrumanus, Chrysaora lactea (PF Long et al., pers comm), by sea anemones [434] and by the highly dangerous box jellyfish Chironex fleckeri[436, 437], no venoms typical of higher animals were found in the A. digitifera genome. This was because our annotation was carried out using the KEGG database (release v58 [53]) to relate A. digitifera protein sequences to KEGG orthologues. The KEGG database is a collection of proteins from well characterised and ubiquitous biochemical pathways. Animal venoms, however, are highly specialised proteins for which this release of the KEGG database does not contain any described orthologues.
Table 15

Proteins homologous to bacterial toxins in the predicted proteome of A. digitifera

Gene sequence

KEGG Orthology

Encoded protein description

v1.20214

K11029

Anthrax edema toxin adenylate cyclase (CyaA)

v1.17686

K10921

Cholera toxin transcriptional activator (ToxR)

v1.13017

K11020

Exotoxin A (ToxA)

v1.23507

K13655

HTH-type transcriptional regulator (MsqA) antitoxin for MqsR

v1.21184

K11009

Murine toxin (Ymt)

v1.04313

K11033

Non-hemolytic enterotoxin A (NheA)

v1.08011

K11034

Non-hemolytic enterotoxin B/C (NheBC)

v1.08255

K13651

Motility quorum-sensing regulator (MqsR) interferase toxin

v1.15986

K11059

Probable enterotoxin A (EntA)

v1.13046

K04392

Ras-related C3 botulinum toxin substrate 1 (Rac1)

v1.13966

K11007

Shiga toxin subunit B (StxB)

v1.23958

K11063

Toxin A/B (TcdAB)

v1.21174

K10930

Toxin co-regulated pilin (TCP)

v1.05802

K10961

Toxin co-regulated pilus biosynthesis protein I (TcpI)

v1.21783

K10964

Toxin co-regulated pilus biosynthesis protein S (TcpS)

v1.14688

K15126

Type III secretion system cytotoxic effector protein (BteA)

v1.05520

K11028

Vacuolating cytotoxin (VacA)

v1.06590

K10954

Zona occludens toxin (Zot)

KEGG orthology-based annotation of the A. digitifera genome reveals genes encoding protein homologues of 10 bacterial toxins, 7 regulatory toxin proteins and a botulinum protein substrate (Table 15). Of the 9 toxin homologues, one with similarity to anthrax edema factor (EF) adenylate cyclase (CyaA) is one of three proteins that comprise the anthrax toxin of Bacillus anthracis, the other two being a protective antigen (PA) and lethal factor (LF). Without the LF protein, anthrax CyaA has no known toxic effects in animals [438], although the EF protein does play an important role in disabling cellular functions vital for microbial host defences [439]. The A. digitifera genome encodes a secretion virulence factor exotoxin A-like protein produced by Pseudomonas aeruginosa, which for this bacterium affects local tissue damage, bacterial invasion and immunosuppression within their eukaryote host [440] with pathogenicity similar to that of the diphtheria toxin [441]. Another encoded protein is a murine-like toxin (Ymt) produced by the enterobacterium Yersinia pestis, which is the causative agent responsible for transmission of the notorious bubonic plague [442]. Additionally, two hemolytic enterotoxins similar to NheA and NheBC produced by Bacillus cereus[443], an enterotoxin (EntA) similar to that of Staphylococcus aureus[444], a Shiga-like enterotoxin (StxB) produced by Shigella dysenteria, the diarrhoea-causing toxin A/B (TcdAB) such as that secreted by Clostridium difficile[445], and a protein similar to the zonula occludens (tight junction) enterotoxin (Zot) secreted by Vibrio cholera[446] are encoded in the A. digitifera genome. Within the predicted proteome is also a homologue of the vacuolating cytotoxin (VacA) produced by Helicobacter pylori that colonises the gastric mucosa of the human stomach epithelium [447].

Although a direct homologue of the cholera toxin (CT) was not found encoded in the A. digitifera genome (Table 15), a protein similar to its transcriptional activator ToxR was. ToxR not only controls the expression of CT in Vibrio cholera[448], but also a co-regulated pilin (TcpA) protein that is under control of the ToxR regulon cascade [449]. Bacterial TcpA protein is assembled into toxin-coregulated pili that induce the transfer of DNA by horizontal exchange of genetic material during conjugation [450]. TcpA and two toxin co-regulated biosynthetic proteins (TcpI and Tcps) of the bacterial virulence-associated pilus appendage [451] are encoded in the coral genome. Entrained also are the motility quorum-sensing interference regulator MsqR and its transcriptional regulator MsqA that in Eschericia coli controls biofilm formation by inhibiting quorum-sensing motility, and together the MqsR/MqsA complex represses the lethal cold shock-like protein cspD gene [452] that on expression impairs DNA replication [453]. The A. digitifera genome likewise encodes a Type III secretion system T3SS cytotoxic effector (BteA) protein [454] that in Gram-negative invasive bacteria is translocated into host cells to suppress innate immunity to enhance virulence [455, 456]. However, the ecophysiological significance of these toxigenic proteins and allied regulators, if indeed expressed by the coral genome, is unknown.

In addition to using the KEGG database, we undertook a BLAST search of the predicted proteome of A. digitifera against peptide sequences for all animal venoms using the annotated UniProtKB/Swiss-Prot Tox-Prot program [457]. This search revealed a large number of accession hits from the predicted proteome, although these are unlikely to be true multiple copies given that the genome sequence has yet to be completely assembled. However, just taking a single accession number from each annotation reveals a complex array of 83 toxins that represents the predicted venom of A. digitifera (Table 16); UniProt BLAST E-values are given in Additional file 1: Table S16b. These venoms are highly diverse and are significantly homologous to toxins from a wide variety of venomous marine and terrestrial creatures such as fish, reptiles, other cnidarians, cone-snails, stinging insects and even a venomous mammal (Shrew), covering the complete range of pharmacological properties known in venoms, including cytolytic, neurotoxic, haemotoxic, phospholipase, proteinase and proteinase inhibitor activities. Both the number of toxins predicted in the venom of A. digitifera and the degree of homology to such widely divergent phyla is remarkable. Accordingly, cnidarian venoms may possess unique biological properties that might generate new leads in the discovery of novel pharmacologically active drugs. Gene duplication followed by mutation and natural selection is widely held as the key mechanism whereby the large diversity of toxins found within a single venom could have evolved [458, 459]. Conversely, primary mRNA splicing patterns have been shown to account for the diversity of metalloproteinases in the pit viper Bothrops neuwiedi[460]. Variations in peptide processing have also been shown by proteomics and transcriptomics to explain how a limited set of gene transcripts could generate thousands of toxins in a single species of cone snail [461]. Despite these various processes that could account for the evolution of toxin diversity, it has never been demonstrated how gene duplications or variations in transcript or peptide processing could have radiated across the very different poisonous creatures found on Earth. Our data (Table 16) reveal that the predicted toxins of A. digitifera venom are orthologues to all of the most important superfamilies of peptide/protein venoms found in diverse taxa. We posit that the origins of toxins in the venoms of higher organisms may have arisen from deep eumetazoan innovations and that the molecular evolution of these venom super gene families can now be addressed taking an integrated venomics approach using Cnidaria such as the jellyfish as model systems [462].
Table 16

UniProt homologues of animal venom proteins in the predicted proteome of A. digitifera

Gene sequence

UniProt toxin accession

Animal with closest homology

v1.01916 [+ 5 other sequence copies]

Q92035; Acetylcholinesterase

Bungarus fasciatus (Banded Krait)

v1.06761; v1.08075; v1.09840; v1.20323

Q9IAM1; Agkisacutacin (subunit anticoagulant protease)

Deinagkistrodon acutus (Sharp-nosed Viper)

v1.04809

A8QL52; L-Amino acid oxidase

Bungarus fasciatus (Banded Krait)

v1.06380

Q4JHE1; L-Amino acid oxidase

Pseudechis australis (Mulga Snake)

v1.10291

P81383; L-Amino acid oxidase

Ophiophagus hannah (King Cobra)

v1.14412

A6MFL0; L-Amino acid oxidase

Demansia vestigiata (Lesser Black Whipsnake)

v1.16469

P81383; L-Amino acid oxidase

Ophiophagus hannah (King Cobra)

v1.23477

P81382; L-Amino acid oxidase

Calloselasma rhodostoma (Malayan Pit Viper)

v1.16440

C5NSL2; Bandaporin (haemolysin)

Anthopleura asiatica (Sea Anemone)

v1.16571 [+ 10 other sequence copies]

Q76B45 ; Blarina toxin (vasoactive protease)

Blarina brevicauda (Northern Short-Tailed Shrew)

v1.06055 [+ 20 other sequence copies]

Q593B6; Coagulation factor V

Pseudonaja textilis (Eastern Brown Snake)

v1.07831; v1.10094 ; v1.20732

P14530; Coagulation factor IX

Protobothrops flavoviridis (Okinawa Habu Snake)

v1.01708 [+ 5 other sequence copies]

Q4QXT9; Coagulation factor X

Tropidechis carinatus (Rough-Scaled Snake)

v1.09601; v1.10410

Q93109; Equinatoxin-5 (cytolysin)

Actinia equina (Beadlet Anemone)

v1.06821

Q08169 ; Hyaluronidase

Apis mellifera (European Honey Bee)

v1.08924

I0CME7; Hyaluronidase, Conohyal-Cn1

Conus consors (Singed Cone)

v1.06189 [+ 112 other sequence copies]

Q9XZC0; α-Latrocrustotoxin Lt1a (neurotoxin)

Latrodectus tredecimguttatus (Mediterranean Black Widow Spider)

v1.02942 [+ 8 other sequence copies]

G0LXV8; α-Latrocrustotoxin Lh1a (neurotoxin)

Latrodectus hasseltii (Australian Redback Spider)

v1.00644 [+ 32 other sequence copies]

Q25338; Δ- Latroinsectotoxin Lt1a (neurotoxin)

Latrodectus tredecimguttatus (Mediterranean Black Widow Spider)

v1.07446

A7X3X3; Lectin, Lectoxin Enh4 (platelet binding)

Enhydris polylepis (Macleay’s Water Snake)

v1.20653

A7X3Y6; Lectin, Lectoxin Enh7 (platelet binding)

Enhydris polylepis (Macleay’s Water Snake)

v1.02561,v1.11493; v1.16681

A7X3Z4; Lectin, Lectoxin Lio1 (platelet binding)

Liophis poecilogyrus (Water Snake)

v1.13597; v1.08696; v1.10757; v1.20654

A7X3Z7; Lectin, Lectoxin Lio2 (platelet binding)

Liophis poecilogyrus (Water Snake)

v1.18386, v1.15479

A7X413; Lectin, Lectoxin Lio3 (platelet binding)

Liophis poecilogyrus (Water Snake)

v1.06094

A7X406; Lectin, Lectoxin Phi1 (platelet binding)

Philodryas olfersii (Green Cobra)

v1.06416; v1.16248; v1.23712

A7X3Z0; Lectin, Lectoxin Thr1 (platelet binding)

Thrasops jacksonii (Black Tree Snake)

v1.17681

Q6TPG9; Lectin, Mucrocetin (platelet binding)

Protobothrops mucrosquamatus (Brown Spotted Pit Viper)

v1.00077 [+ 14 other sequence copies]

Q66S03; Lectin, Nattectin (platelet binding)

Thalassophryne nattereri (Toad Fish)

v1.12241; v1.02332; v1.12298

Q71RQ1; Lectin, Stejaggregin-A (platelet binding)

Trimeresurus stejnegeri (Bamboo Viper)

v1.02245 [+ 19 other sequence copies]

A0FKN6; Metalloprotease, Astacin-like toxin

Loxosceles intermedia (Recluse Spider)

v1.03638; v1.14772

Q90391; Metalloprotease, Atrolysin

Crotalus atrox (Western Diamondback Rattlesnake)

v1.13106

D3TTC2; Metalloproteinase, Atragin

Naja atra (Chinese Cobra)

v1.11132

Q7T1T4; Metalloproteinase, BjussuMP-2

Bothrops jararacussu (Jararacussu Pit Viper)

v1.02168

O73795; Metalloproteinase, Disintegrin

Gloydius brevicaudus (Chinese Mamushi Snake)

v1.06910

Q7SZE0; Metalloproteinase, Disintegrin

Gloydius saxatilis (Rock Mamushi Snake)

v1.22282

P14530; Metalloproteinase, Disintegrin

Protobothrops flavoviridis (Okinawa Habu Snake)

v1.03804

Q2UXQ5; Metalloproteinase, EoVMP2

Echis ocellatus (West African Carpet Viper)

v1.02016

Q91511; Mucrofibrase-5, Hypotensive serine protease

Protobothrops mucrosquamatus (Brown Spotted Pit Viper)

v1.09026

Q7ZZN8; Natrin-2 (neurotoxin)

Naja atra (Chinese Cobra)

v1.04153; v1.04595; v1.12730; v1.04157

A0ZSK3; Neoverrucotoxin (haemolysin)

Synanceia verrucosa (Reef Stone Fish)

v1.12433 [+ 5 other sequence copies]

A2VBC4; Phospholipase A1

Polybia paulista (Neotropical Social Wasp)

v1.00019; v1.13757

Q06478; Phospholipase A1 1

Dolichovespula maculata (Bald-Faced Hornet)

v1.09322; v1.09961; v1.13629

P0CH47; Phospholipase A1, Magnifin

Vespa magnifica (Giant Hornet)

v1.03556

P53357; Phospholipase A1 2

Dolichovespula maculata (Bald-Faced Hornet)

v1.13015; v1.16921

D2X8K2; Phospholipase A2

Condylactis gigantean (Giant Caribbean Sea Anemone)

v1.18628

Q9TWL9; Phospholipase A2, Conodipine-M

Conus magus (Magical Cone)

v1.11796

Q9PUH9; Phospholipase A2, Acidic S9-53 F

Austrelaps superbus (Lowland Copperhead Snake)

v1.09883

Q8AXW7; Phospholipase A2, Basic

Micrurus corallinus (Painted Coral Snake)

v1.14874

Q90WA8; Phospholipase A2, Basic 2

Bungarus fasciatus (Banded Krait)

v1.11797

P20256; Phospholipase A2, Basic PA-12C

Pseudechis australis (Mulga Snake)

v1.07278 [+ 34 other sequence copies]

Q7SZN0; Prothrombin activator Pseutarin-C

Pseudonaja textilis (Eastern Brown Snake)

v1.11045

P83370; Prothrombin activator Hopsarin-D

Hoplocephalus stephensii (Stephen’s Branded Snake)

v1.04104 [+ 5 other sequence copies]

Q58L94; Prothrombin activator Notecarin D2

Notechis scutatus (Tiger Snake)

v1.00387 [+ 9 other sequence copies]

Q58L90; Prothrombin activator Omicarin C

Oxyuranus microlepidotus (Inland Taipan )

v1.02137 [+ 38 other sequence copies]

Q58L91; Prothrombin activator Omicarin C

Oxyuranus scutellatus (Coastal Taipan)

v1.00618 [+ 10 other sequence copies]

Q58L93; Prothrombin activator Porpharin D

Pseudechis porphyriacus (Red-Bellied Black Snake)

v1.09896

P81428; Prothrombin activator Trocarin D

Tropidechis carinatus (Rough-Scaled Snake)

v1.13726

A6MFK7; Prothrombin activator Vestarin D1

Demansia vestigiata (Lesser Black Whipsnake)

v1.02129; v1.05362; v1.20273

Q6T269; Protease inhibitor, Bitisilin-3 (neurotoxic)

Bitis gabonica (Gaboon Viper)

v1.06980; v1.09028

Q3SB05; Pseudechetoxin (neurotoxin)

Pseudonaja textilis (Eastern Brown Snake)

v1.21284 [+ 5 other sequence copies]

D8VNS7; Ryncolin-1 (haemostasis inhibitor)

Cerberus rynchops (Dog-Faced Water Snake)

v1.18895 [+ 20 other sequence copies]

D8VNS8; Ryncolin-2 (haemostasis inhibitor)

Cerberus rynchops (Dog-Faced Water Snake)

v1.14251; v1.10489; v1.14254

D8VNS9; Ryncolin-3 (haemostasis inhibitor)

Cerberus rynchops (Dog-Faced Water Snake)

v1.06759 [+ 7 other sequence copies]

D8VNT0; Ryncolin-4 (haemostasis inhinitor)

Cerberus rynchops (Dog-Faced Water Snake)

v1.01273

Q9YGN4; Salmorin toxin (haemostasis inhibitor)

Gloydius brevicaudus (Chinese Mamushi Snake)

v1.09855; v1.09856

B2DCR8; SE-Cephalotoxin

Sepia esculenta (Golden Cuttlefish)

v1.16247

O13060; Serine protease, 2A

Trimeresurus gramineus (Bamboo Viper)

v1.08397; v1.09733

Q9DF66; Serine protease, 3 (haemostasis inhibitor)

Protobothrops jerdonii (Jerdon’s Pit Viper)

v1.03275

Q9DG84; Serine protease, Serpentokallikrein-2 (haemostasis inhibitor)

Protobothrops mucrosquamatus (Brown Spotted Pit Viper)

v1.16638

Q7SYF1; Serine protease, Cerastocytin (platelet binding)

Cerastes cerastes (Saharan Horned Viper)

v1.22320

P0C5B4; Serine protease, Gloshedobin (platelet binding)

Gloydius shedaoensis (Shedao Pit Viper)

v1.15074 [+ 4 other sequence copies]

B2D0J4; Serine protease, Venom dipeptidyl peptidase 4

Apis mellifera (European Honey Bee)

v1.05361

B6RLX2; Serine protease inhibitor, TCI (neurotoxin)

Ophiophagus hannah (King Cobra)

v1.10994

B7S4N9; Serine protease inhibitor, Taicatoxin (neurotoxin)

Oxyuranus scutellatus (Coastal Taipan)

v1.11218; v1.23374

Q90WA0; Serine protease inhibitor, Textilinin-2 (thrombin inhibitor)

Pseudonaja textilis (Eastern Brown Snake)

v1.17856; v1.22256

Q8T3S7; Serine protease inhibitor, U1-aranetoxin-Av1a (neurotoxin)

Araneus ventricosus (Devil Spider)

v1.04154 [+ 4 other sequence copies]

Q98989; Stonustoxin (haemostasis inhibitor)

Synanceia horrida (Estuarine Stonefish)

v1.09427; v1.16619; v1.19446

Q76DT2; Toxin AvTX-60A (cytolysin)

Actineria villosa (Okinawan Sea Anemone)

v1.12311

Q9GV72; Toxin CrTX-A (haemolysin)

Carybdea rastonii (Jimble Jellyfish)

v1.07546 [+ 5 other sequence copies]

P58911; Toxin PsTX-60 (haemolysin)

Phyllodiscus semoni (Night Anemone)

v1.11270; v1.14265

E2IYB3; Veficolin-1 (complement activator)

Varanus komodoensis (Komodo Dragon)

v1.02115

Q98993; Verrucotoxin (cytolysin)

Synanceia verrucosa (Reef Stonefish)

Detoxification proteins of the chemical defensome

There have been considerable advancements made to better understand the effects of pollution on coral reef habitats. The three main categories of environmental pollutants from anthropogenic sources are nutrient enrichment (eutrophication), hydrocarbon pollution and heavy metal contamination. Eutrophication from terrestrial inputs are a significant threat to coral reefs stemming from the discharge of treated sewage, the runoff of agricultural fertilizers (plus herbicides and pesticides), and by sedimentation caused by the erosion of organic-rich soils [463]. Notwithstanding that eutrophication can shift coral reef communities towards macroalgae domination [19], nitrogen and phosphorus enrichment can diminish coral growth and affect the photosynthetic performance of their algal symbionts [464]. Nutrient enhancement alters multiple pathways of primary metabolism that in coral is complicated by the photosynthetic demands of its symbiotic partners. While corals respond to hypertrophic levels of nutrients by activating general stress-response proteins [465], there are no specific proteins known to mitigate the cellular effects of nutrient enrichment on corals per se, and we have not attempted to identify such in this study.

Gene families and their regulators that defend against chemical stressors comprise the chemical defensome encoding a network of detoxifying proteins that allows an organism to sense, transform and eliminate potentially toxic endogenous metabolites and xenobiotic contaminants [466]. Expressed proteins of the chemical defensome include the biotransformation cytochrome P450 (CYP) family of enzymes, conjugating enzymes, efflux transporters, heavy metal membrane pump exporters and their transcriptional activators. Annotation of the genome of A. digitifera reveals multiple genes encoding 20 hemoproteins belonging to the Phase II cytochrome P450 superfamily of monooxidase enzymes that catalyse the oxidation of diverse organic substances (Table 17). The substrates of CYP enzymes include intermediates of lipid metabolism and sterol/steroid biosynthesis, and include the detoxification of exogenous xenobiotics. Of significance are the CYP1A-type (aryl hydrocarbon hydroxylase) enzymes that have been studied widely in the hepatic response of fishes to polycyclic aromatic hydrocarbon (PAH) contamination (from crude or fuel oil) and exposure to polychlorinated biphenyl and dibenzodioxin toxicants (reviewed in [467]). CYP450 activity has been detected in the corals Favia fragum[468], Siderastrea siderea[469], Montastraea faveolata[470] and Pocillopora damicornis, [471]. Furthermore, CYP encoding sequences have been extracted from the genome of N. vectensis[472] and the transcriptome of A. millepora[29]. As well as providing chemical defence, mixed-function CYPs perform multiple endogenous tasks that are often taxon-specific. Hence, the orthology and substrate specificity of coral CYP enzymes cannot be predicted solely on homology to CYPs of known function assigned to higher metazoans. Similar to the function of CPY enzymes, there are genes encoding p-hydroxybenzoate 3-monooxygenase, an oxidoreductase catalyzing aryl oxidation and the soluble and microsomal forms of epoxide hydrolase that converts epoxides, formed by the degradation of aromatic compounds, to trans-diols that by conjugation are readily excreted. Conjugating enzymes to eliminate hydroxylated substrates are the detoxifying UDP-glucuronosyltransferase and sulfotransferase families of enzymes. Estrone sulfotransferase is significant for inactivation of exogenous (contraceptive) estrogens [473] and similar endocrine-disruptive contaminants released from treated wastewater [474]; their occurrence in marine waters are known to disrupt the reproduction and development of fish [475] and corals [476]. Glutathione S-transferase (GST) enzymes catalyse the addition of reduced glutathione to the reactive sites of electrophilic toxins [477]. Surprisingly, only two isoforms of GST were detected in the A. digitifera genome (Table 17), whereas 18 distinct GST-encoding genes (6 classes + 1 fungal-type) were classified from genome sequences of N. vectensis[472]. This unexpected genome reduction of GST elaboration in A. digitifera begs further examination.
Table 17

Proteins of the chemical defensome in the predicted proteome of A. digitifera

Gene sequence

KEGG Orthology

Encoded protein description

v1.06127; v1.06128

K01015

Alcohol sulfotransferase

v1.09267

K00537

Arsenate reductase

v1.24496; v1.24495; v1.03953

K03893

Arsenical pump membrane protein

v1.10691

K07755

Arsenite methyltransferase

v1.20443

K11811

Arsenical resistance protein ArsH

v1.14972

K01551

Arsenite-transporting ATPase

v1.17644; v1.00480; v1.08150; v1.22865

K01014

Aryl sulfotransferase

v1.21535; v1.11835; v1.02456

K01534

Cd2+/Zn2+-exporting ATPase

v1.03485; v1.21926; v1.05686

K01533

Cu2+-exporting ATPase

v1.22646 [+ 8 other sequence copies]

K07408

Cytochrome P450, family 1, subfamily A, polypeptide 1

v1.01284

K07421

Cytochrome P450, family 2, subfamily T

v1.10544; v1.02314, v1.17490

K07422

Cytochrome P450, family 2, subfamily U

v1.23039 [+ 13 other sequence copies]

K07422

Cytochrome P450, family 3, subfamily A

v1.07750

K07425

Cytochrome P450, family 4, subfamily A

v1.22798; v1.23000

K07426

Cytochrome P450, family 4, subfamily B

v1.02020 [+ 4 other sequence copies]

K07427

Cytochrome P450, family 4, subfamily V

v1.19495

K07428

Cytochrome P450, family 4, subfamily X

v1.15382

K15002

Cytochrome P450, family 6

v1.16427

K07430

Cytochrome P450, family 7, subfamily B

v1.17631

K00498

Cytochrome P450, family 11, subfamily A

v1.08074 [+ 4 other sequence copies]

K15004

Cytochrome P450, family 12

v1.02478 [+ 5 other sequence copies]

K00512

Cytochrome P450, family 17, subfamily A

v1.06713

K07435

Cytochrome P450, family 20, subfamily A

v1.22414 [+ 5 other sequence copies]

K07436

Cytochrome P450, family 24, subfamily A

v1.20153

K12665

Cytochrome P450, family 26, subfamily C

v1.08074 [+ 6 other sequence copies]

K00488

Cytochrome P450, family 27, subfamily A

v1.06537

K07439

Cytochrome P450, family 39, subfamily A

v1.22302 [+ 5 other sequence copies]

K07440

Cytochrome P450, family 46, subfamily A

v1.16335

K09832

Cytochrome P450, family 710, subfamily A

v1.18439; v1.02594; v1.02593

K01016

Estrone sulfotransferase

v1.07758 [+ 5 other sequence copies]

K00699

Glucuronosyltransferase

v1.00764

K13299

Glutathione S-transferase kappa 1

v1.17188

K00799

Glutathione S-transferase

v1.04140

K07239

Heavy-metal exporter, HME family

v1.10181

K00481

p-Hydroxybenzoate 3-monooxygenase

v1.16748; v1.07471

K08365

MerR family transcriptional regulator, mercuric resistance

v1.04382; v1.24424

K13638

MerR family transcriptional regulator, Zn(II)-responsive

v1.12760

K08363

Mercuric ion transport protein

v1.04179; v1.01891; v1.00145

K03284

Metal ion transporter, MIT family

v1.21500 [+ 5 other sequence copies]

K01253

Microsomal epoxide hydrolase

v1.08005

K08970

Nickel/cobalt exporter

v1.03484

K08364

Periplasmic mercuric ion binding protein

v1.05406

K07245

Putative copper resistance protein D

v1.14635

K08726

Soluble epoxide hydrolase

v1.01929; v1.19296

K05794

Tellurite resistance protein TerC

v1.10880; v1.15709; v1.12348

K07803

Zinc resistance-associated protein

Many toxicological studies on the effects of pollution on cnidarian fitness have focused on their response to heavy metal contamination, including copper, cadmium, mercury and zinc [478, 479]. In scleractinian corals the uptake and toxic effects of copper [480483], cadmium [482] and mercury [484, 485] have been studied at the metabolic level with specific studies to examine the effects of heavy metal toxicity on coral fertilisation [486488], settlement [487], metamorphosis [486] and in coral bleaching [489]. Yet, the identification of molecular markers to monitor the response of Cnidaria to sub-lethal levels of heavy metal exposure has been elusive [490]. We were delighted to uncover in our annotation a wide range of genes to express metal-specific (arsenic, copper, mercury, nickel/cobalt and tellurium) resistance, transportation and membrane pump exporting proteins that, together with non-specific heavy metal ion export proteins (Table 17), might prove useful for monitoring the environmental response of A. digitifera to heavy metal contamination. Included in the heavy metal defensome are the Mer-family of transcriptional regulators of Hg- and Zn-resistance proteins and a periplasmic ion-binding protein attributed to the Hg detoxification system of bacteria [491]. Enzymes specific for arsenic detoxification are an arsenate oxidoreductase for conversion of arsenate to arsenite [492] and arsenite methyltransferase for conversion of arsenite to the less toxic dimethylarsenite that is amenable to excretion [493]. Such processes may enhance the resilience of corals exposed to natural [494] and site-affected [495] levels of arsenic contamination. In contrast, there were no (organo)cyanide detoxification genes apparent in the A. digitifera genome, but one sequence (v1.01601; K10814) encodes for hydrogen cyanide synthase of unknown metabolic purpose (data not tabulated). Ancillary evidence suggests that the expression of HCN synthase could be linked to quorum sensing [496] for regulating microbial densities of the coral holobiont community.

Epigenetic and DNA-remodelling proteins

In all Kingdoms of life, DNA methylation and chromatin remodelling is pivotal to the regulation of gene transcription independent of underlying allelic variation. One such process mediated by epigenetic changes in eukaryotic biology is the all-important cellular differentiation during morphogenetic development. Epigenetic modifications cause the activation, regulation or silencing of certain genes without changing the basic DNA code. Changes in epigenetic regulation can persist during cell division and across multiple generations [497]. In addition, cytosine methylation may be associated with a higher mutation rate, because deamination of the methylated base produces thymine resulting in C/T mutations, which on reproduction may be transmitted by the germline to subsequent generations in selective processes of evolution [498]. On the other hand, environmentally induced destabilisation of the epigenome can produce gene variants (epialleles) that activate transcription and mobilization of DNA transposable elements, which may subsequently lead to stable heritable traits of environmental adaptation, as does occur by genetic imprinting in plants [499]. Transposition has thus the potential to direct increased frequencies of permanent genetic mutations for selective adaptation.

One way by which genes are regulated at the epigenome is through the remodelling of the chromatin histone-DNA complex (the nucleosome), which by post-translational modification changes the template structure of DNA associated histone proteins. These modifications are affected by histone-lysine (and histone-arginine) N-methyltransferase enzymes (Table 18) by which these proteins may be further modified by acetylation, ADP-ribosylation, ubiquination, and phosphorylation (annotation not tabulated). The methylation pattern of histone lysine residues is highly predictive of the gene expression states of transcriptional activation and repression [500]. Necessary epigenomic reprogramming of histone modification at different stages of cell development is affected by the activation of histone and lysine-specific demethylase enzymes (Table 18). Determinants for recognition of the histone code are being revealed by a growing body of experimental data providing valuable information on the molecular tractability of binding sites involved in epigenetic signalling [501], which will enhance further insight to epigenetic function.
Table 18

Epigenetic and DNA-remodelling proteins in the predicted proteome of A. digitifera

Gene sequence

KEGG Orthology

Encoded protein description

v1.04426; v1.02042

K02528

16S rRNA (adenine1518-N6/1519-N6)-dimethyltransferase

v1.22358; v1.00249

K14191

18S rRNA (adenine1779-N6/1780-N6)-dimethyltransferase

v1.19400; v1.04238

K00561

23S rRNA (adenine2085-N6)-dimethyltransferase

v1.05107; v1.05242

K01488

Adenosine deaminase

v1.04152; v1.09790

K14857

AdoMet-dependent rRNA methyltransferase SPB1

v1.00197

K13530

AraC family transcriptional regulator DNA methyltransferase

v1.12967; v1.19789; v1.07763

K14589

Cap-specific mRNA (nucleoside-2′-O-)-methyltransferase 1

v1.24281

K01489

Cytidine deaminase

v1.16211; v1.14952; v1.01094; v1.06983

K00558

DNA (cytosine-5-)-methyltransferase

v1.19683; v1.05688; v1.04223

K11324

DNA methyltransferase 1-associated protein 1

v1.14033; v1.19860; v1.19081; v1.04188

K11420

Euchromatic histone-lysine N-methyltransferase

v1.02068

K01487

Guanine deaminase

v1.02920

K05931

Histone-arginine methyltransferase CARM1

v1.17589 [+ 7 other sequence copies]

K11446

Histone demethylase JARID1

v1.07640

K06101

Histone-lysine N-methyltransferase ASH1L

v1.13515; v1.18577; v1.20187; v1.19182

K09186

Histone-lysine N-methyltransferase MLL1

v1.08381

K09187

Histone-lysine N-methyltransferase MLL2

v1.24258; v1.19182

K09188

Histone-lysine N-methyltransferase MLL3

v1.07992; v1.10302; v1.13829

K09189

Histone-lysine N-methyltransferase MLL5

v1.06939; v1.15255; v1.15254

K11424

Histone-lysine N-methyltransferase NSD1/2

v1.05552

K11422

Histone-lysine N-methyltransferase SETD1

v1.07744

K11423

Histone-lysine N-methyltransferase SETD2

v1.03190

K11431

Histone-lysine N-methyltransferase SETD7

v1.21867

K11428

Histone-lysine N-methyltransferase SETD8

v1.18700 [+ 8 other sequence copies]

K11421

Histone-lysine N-methyltransferase SETDB

v1.07557; v1.11409

K11419

Histone-lysine N-methyltransferase SUV39H

v1.24733; v1.13497

K11429

Histone-lysine N-methyltransferase SUV420H

v1.15405; v1.10291; v1.17601; v1.02845; v1.08629

K11450

Lysine-specific histone demethylase 1

v1.23155; v1.09394; v1.17624; v1.05370

K14835

Ribosomal RNA methyltransferase Nop2

v1.18460 [+ 6 other sequence copies]

K03500

Ribosomal RNA small subunit methyltransferase B

v1.07407; v1.03110

K08316

Ribosomal RNA small subunit methyltransferase D

v1.12193

K02427

Ribosomal RNA large subunit methyltransferase E

v1.11499

K11392

Ribosomal RNA small subunit methyltransferase F

v1.16053; v1.12676

K03437

RNA methyltransferase, TrmH family

v1.12453; v1.05459

K13097

Methylcytosine dioxygenase

v1.07692

K07451

5-Methylcytosine-specific restriction enzyme A

v1.21815; v1.17113

K00565

mRNA (guanine-N7-)-methyltransferase

v1.06363; v1.03360; v1.21218

K05925

mRNA (2′-O-methyladenosine-N6-)-methyltransferase

v1.09661

K07442

tRNA (adenine-N1-)-methyltransferase catalytic subunit

v1.08094; v1.04036; v1.18614

K03256

tRNA (adenine-N(1)-)-methyltransferase non-catalytic subunit

v1.11456; v1.00738; v1.04577

K03439

tRNA (guanine-N7-)-methyltransferase

v1.08042

K14864

tRNA methyltransferase

v1.20501

K00557

tRNA (uracil-5-)-methyltransferase

v1.15147

K14964

Set1/Ash2 histone methyltransferase subunit ASH2

v1.08925

K00571

Site-specific DNA-methyltransferase (adenine-specific)

Direct epigenetic modification of DNA (or mRNA) occurs by methylation of cytosine, and to a lesser extent adenosine and guanine, by nucleobase-specific DNA methyltranferases (Table 18) to give 5-methylcytosine (5-meC), 3-methyladenosine (3-meA) and 3-methylguanine (3-meG) nucleotides, respectively. The principal modification product, 5-methylcytosine behaves much like regular cytosine by pairing with guanine, but in areas of high cytosine methylation, genome transcription is strongly repressed (reviewed in [502]), together with the repression of other chromatin-dependent processes, including the incorporation of transposable elements [503]. Alteration in the methylation status of the entire genome, individual chromosomes or at specific gene sites is essential for normal cellular function, but processes for reprogramming methylated DNA at different stages of cell development, unlike the reversal of histone modifications, is poorly defined [504]. While there are abundant enzymes to repair DNA damage caused by spurious N-alkylation, direct nucleotide C-demethylation (via the hypothetical “DNA demethylase” [505]) is thermodynamically infeasible. Instead, removal of epigenetic C-methylated nucleobases occurs by several base-repair pathways involving DNA excision or mismatch repair enzymes. The genome of A. digitifera encodes expression of a specific DNA glycosylase enzyme [506] for excision of 3-meA, but there are no such enzymes encoded for the excision of 5-meC and 3-meG, although there is encoded a 5-methylcytosine-specific restriction enzyme. Another pathway for DNA demethylation requires base-specific deamination by the AID/Apobec family of deaminase enzymes that, for example, converts 5-meC to thymine that is replaced subsequently by cytosine by C/T mismatch repair enzymes. These methylated nucleobases are recognized for deamination by the cytosine, adenosine and guanine deaminase enzymes [507] that are encoded in the A. digitifera genome, and their deaminated bases are subsequently removed by DNA mismatch repair enzymes. Additionally, the genome of A. digitifera encodes a methlycytosine dioxygenase enzyme that converts 5-methylcytosine to 5-hydroxymethycytosine (5-hmC), which is recognized for removal by the base excision repair pathway [508] or via its 5-hmC deaminated intermediate [507]. Combined, these DNA demethylation pathways are able to remodel epigenetic modifications at different stages of cell development.

Most current knowledge of DNA and protein methylation comes from studies of mammals and plants, while our understanding of the extent and roles of DNA methylation in invertebrates, marine invertebrates in particular, is still limited [509]. Little is known about the epigenetic potential of corals to acclimatize and adapt to the thermal and synergistic stressors that cause wide-spread coral “bleaching” [510]. Yet, given that acclimatization occurs via the generation of epiallele variants that can in some instances lead to stable heritable traits of environmental adaptation, there is growing interest in the prospect that epigenetic modifications in corals or their algal symbionts [511] may drive adaptation to defend against the damaging threat imposed by rising temperatures from global climate change. It is anticipated that this field of study will rapidly accelerate with the need to better understand epigenetic processes that may contribute to the persistence of coral reefs.

Conclusions

We offer ZoophyteBase as an unprecedented foundation to interrogate the molecular structure of the predicted A. digitifera proteome. Some key findings include proteins with relevance to host-symbiont function, dysfunction and recovery including those that direct vacuolar trafficking and proteins linking symbiont photosynthesis to coral calcification. An extensive catalogue of mammalian-like proteins essential to neural function and venoms related to distant animal phyla suggests their origins lie deep in early eumetazoan evolution. Homologues of prokaryotic genes that have not been described previously in any eukaryote genome such as flagella proteins, proteins essential for nitrogen fixation and photosynthesis point towards lateral gene transfer, perhaps mediated by viruses, that may lead to “shared” metabolic adaptations of symbiosis, and provide corals with limited ability for gene-encoded adaptation to a changing global environment. It is anticipated that understanding how the genome of a coral host interacts with that of its vast array of symbionts, and how it may regulate its metabolic quotient, for example through biochemical or epigenetic modification, will rapidly accelerate our ability to predict the fate of coral reefs.

Availability and requirements

ZoophyteBase was constructed using the Metagenome/Genome Annotated Sequence Natural Language Search Engine (MEGGASENSE). This is a general system for the annotation of sequence collections and presentation of the results in a database that can be searched using biologically intuitive search terms. In this implementation, the predicted proteome of A. digitifera (genome assembly v1.0 [48]) was used as the source of protein sequences. The annotation was carried out using the KEGG database (release v58 [51]) to relate A. digitifera protein sequences to KEGG orthologues. The homologous protein sequences were used to construct hidden Markov model (HMM) profiles using the HMMER3 package [49]. The predicted proteome sequences of A. digitifera were searched with HMM profiles to link proteins to appropriate KEGG orthologues [50, 512]. A web interface was developed with various tools. The search platform Lucene/Solr [52] was used to implement natural language searches. Protein sequences provided by the user can be used for BLAST [50] searches against the coral proteome. Selected sequences of the coral proteome can be analysed with third party software (e.g. [53]) to interrogate conserved domains. ZoophyteBase is deployed using Apache-Tomcat (version 7.0.28 for Linux ×64 [513]) on the Ubuntu Linux server of the Section of Bioinformatics at the Faculty of Food Technology and Biotechnology, University of Zagreb, Croatia and is accessible at our published web address [47].

Declarations

Acknowledgments

The authors are indebted to Professor Noriyuki Satoh and his research team at the Marine Genomics Unit, Okinawa Institute of Science and Technology, Japan for making the protein sequence of A. digitifera publically available without restriction of use. We thank the following scientists from the Australian Institute of Marine Science for helpful discussions, Dr David Bourne, Dr Andrew Negri, Dr Elisha Wood-Charlson and Dr Karen Weynberg. We are also grateful to Dr Bill Leggat of the ARC Centre of Excellence for Coral Reef Studies and School of Pharmacy and Molecular Sciences, James Cook University, Townsville, Australia for critically reviewing this manuscript. Financial support for this work has come from the Biotechnology and Biological Sciences Research Council of the United Kingdom (BBSRC grant BB/H010009/2 to WCD and PFL); from the Australian Institute of Marine Science (WCD and MvO); from a cooperation grant of the German Academic Exchange Service (DAAD) and Ministry of Science, Education and Sports, Republic of Croatia (to JC and DH) and from the Croatian Science Foundation (grant 09/5 to DH). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Initial work (in progress) was presented at the 12th International Coral Reef Symposium, Cairns, Queensland, Australia, 9-13 July 2012.

Authors’ Affiliations

(1)
Centre for Marine Microbiology and Genetics, Australian Institute of Marine Science
(2)
Institute of Pharmaceutical Science, King’s College London
(3)
Department of Chemistry King’s College London
(4)
Section for Bioinformatics, Department of Biochemical Engineering, Faculty of Food Technology and Biotechnology, University of Zagreb
(5)
Department of Genetics, University of Kaiserslautern

References

  1. Freudenthal HD: Symbiodinium gen. nov. and Symbiodinium microadriaticum sp. nov., a zooxanthella: taxonomy, life cycle, and morphology. J Eukaryot Microbiol. 1962, 9: 45-52.Google Scholar
  2. Muscatine L: The role of symbiotic algae in carbon and energy flux in reef corals. Coral Reefs: Ecosystems of the World. Edited by: Dubinsky Z. 1990, Amsterdam: Elsevier, 75-84.Google Scholar
  3. Yellowlees D, Rees TAV, Leggat W: Metabolic interactions between algal symbionts and invertebrate hosts. Plant Cell Environ. 2008, 31: 679-694. 10.1111/j.1365-3040.2008.01802.x.PubMedGoogle Scholar
  4. Johannes RE, Wiebe WJ, Crossland CJ, Rimmer DW, Smith SV: Latitudinal limits of coral reef growth. Mar Ecol Prog Ser. 1983, 11: 105-111.Google Scholar
  5. Spalding MD, Grenfell AM: New estimates of global and regional coral reef areas. Coral Reefs. 1997, 16: 225-230. 10.1007/s003380050078.Google Scholar
  6. Hatcher BG: Coral reef primary productivity. A hierarchy of pattern and process. Trends Ecol Evol. 1990, 5: 149-155. 10.1016/0169-5347(90)90221-X.PubMedGoogle Scholar
  7. Reaka-Kudla ML: The global biodiversity of coral reefs: A comparison with rainforests. Biodiversity II. Understanding and Protecting Our Natural Resources. Edited by: Reaka-Kudla ML, Wilson DE, Wilson EO. 1997, Washington DC: Joseph Henry/National Academy Press, 83-108.Google Scholar
  8. Davy SK, Allemand D, Weis VM: Cell biology of cnidarians-dinoflagellate symbiosis. Microbiol Mol Biol Rev. 2012, 76: 229-261. 10.1128/MMBR.05014-11.PubMed CentralPubMedGoogle Scholar
  9. Bellwood DR, Hughes TP, Folke C, Nyström M: Confronting the coral reef crisis. Nature. 2004, 429: 827-833. 10.1038/nature02691.PubMedGoogle Scholar
  10. Hoegh-Guldberg O: Climate change, coral bleaching and the future of the world’s coral reefs. Mar Freshwat Res. 1999, 50: 839-866. 10.1071/MF99078.Google Scholar
  11. Hoegh-Guldberg O, Mumby PJ, Hooten AJ, Steneck RS, Greenfield P, Gomez E, Harvell CD, Sale PF, Edwards AJ, Caldeira K, Knowlton N, Eakin CM, Iglesias-Prieto R, Muthiga N, Bradbury RH, Dubi A, Hatziolos ME: Coral reefs under rapid climate change and ocean acidification. Science. 2007, 318: 1737-1742. 10.1126/science.1152509.PubMedGoogle Scholar
  12. Porter JW, Dustan P, Jaap WC, Patterson KL, Kosmynin V, Meier OW, Patterson ME, Parsons M: Patterns of spread of coral disease in the Florida Keys. Hydrobiologia. 2001, 460: 1-24. 10.1023/A:1013177617800.Google Scholar
  13. Bruno JF, Selig ER, Casey KS, Page CA, Willis BL, Harvell CD, Sweatman H, Melendy AM: Thermal stress and coral cover as drivers of coral disease outbreaks. PLoS Biol. 2007, 5: e124-10.1371/journal.pbio.0050124.PubMed CentralPubMedGoogle Scholar
  14. Brandt ME, McManus JW: Disease incidence is related to bleaching extent in reef-building corals. Ecology. 2009, 90: 2859-2867. 10.1890/08-0445.1.PubMedGoogle Scholar
  15. Devlin MJ, Brodie J: Terrestrial discharge into the Great Barrier Reef Lagoon: nutrient behaviour in coastal waters. Mar Pollut Bull. 2005, 51: 9-22. 10.1016/j.marpolbul.2004.10.037.PubMedGoogle Scholar
  16. Fabricius KE: Effects of terrestrial runoff on the ecology of corals and coral reefs: review and synthesis. Mar Pollut Bull. 2005, 50: 125-146. 10.1016/j.marpolbul.2004.11.028.PubMedGoogle Scholar
  17. Orr JC, Fabry VJ, Aumont O, Bopp L, Doney SC, Feely RA, Gnanadesikan A, Gruber N, Ishida A, Joos F, Key RM, Lindsay K, Maier-Reimer E, Matear R, Monfray P, Mouchet A, Najjar RG, Plattner GK, Rodgers KB, Sabine CL, Sarmiento JL, Schlitzer R, Slater RD, Totterdell IJ, Weirig MF, Yamanaka Y, Yool A: Anthropogenic ocean acidification over the twenty-first century and its impact on calcifying organisms. Nature. 2005, 437: 681-686. 10.1038/nature04095.PubMedGoogle Scholar
  18. Anthony KRN, Kline DI, Diaz-Pulido G, Dove S, Hoegh-Guldberg O: Ocean acidification causes bleaching and productivity loss in coral reef builders. Proc Natl Acad Sci U S A. 2008, 105: 17442-17446. 10.1073/pnas.0804478105.PubMed CentralPubMedGoogle Scholar
  19. De’ath G, Fabricius K: Water quality as a regional driver of coral biodiversity and macroalgae on the Great Barrier Reef. Ecol Appl. 2010, 20: 840-850. 10.1890/08-2023.1.PubMedGoogle Scholar
  20. Carpenter KE, Abrar M, Aeby G, Aronson RB, Banks S, Bruckner A, Chiriboga A, Cortés J, Delbeek JC, Devantier L, Edgar GJ, Edwards AJ, Fenner D, Guzmán HM, Hoeksema BW, Hodgson G, Johan O, Licuanan WY, Livingstone SR, Lovell ER, Moore JA, Obura DO, Ochavillo D, Polidoro BA, Precht WF, Quibilan MC, Reboton C, Richards ZT, Rogers AD, Sanciangco J: One third of reef-building corals face elevated extinction risk from climate change and local impacts. Science. 2008, 321: 560-563. 10.1126/science.1159196.PubMedGoogle Scholar
  21. Pendleton LH: Valuing coral reef protection. Ocean Coast Manage. 1995, 26: 119-131. 10.1016/0964-5691(95)00007-O.Google Scholar
  22. Halpern BS, Walbridge S, Selkoe KA, Kappel CV, Micheli F, D’Agrosa C, Bruno JF, Casey KS, Ebert C, Fox HE, Fujita R, Heinemann D, Lenihan HS, Madin EM, Perry MT, Selig ER, Spalding M, Steneck R, Watson R: A global map of human impact on marine ecosystems. Science. 2008, 319: 948-952. 10.1126/science.1149345.PubMedGoogle Scholar
  23. Hughes TP, Baird AH, Bellwood DR, Card M, Connolly SR, Folke C, Grosberg R, Hoegh-Guldberg O, Jackson JB, Kleypas J, Lough JM, Marshall P, Nyström M, Palumbi SR, Pandolfi JM, Rosen B, Roughgarden J: Climate change, human impacts, and the resilience of coral reefs. Science. 2003, 301: 929-933. 10.1126/science.1085046.PubMedGoogle Scholar
  24. Evans TG, Hofmann GE: Defining the limits of physiological plasticity: how gene expression can assess and predict the consequences of ocean change. Philos Trans R. Soc B-Biol Sci. 2012, 367: 1733-1745. 10.1098/rstb.2012.0019.Google Scholar
  25. Weis VM: The susceptibility and resilience of corals to thermal stress: adaptation, acclimatization or both?. Mol Ecol. 2010, 19: 1515-1517. 10.1111/j.1365-294X.2010.04575.x.PubMedGoogle Scholar
  26. Edge SE, Morgan MB, Gleason DF, Snell TW: Development of a coral cDNA array to examine gene expression profiles in Montastraea faveolata exposed to environmental stress. Mar Pollut Bull. 2005, 51: 507-523. 10.1016/j.marpolbul.2005.07.007.PubMedGoogle Scholar
  27. Grasso LC, Maindonald J, Rudd S, Hayward DC, Saint R, Miller DJ, Ball EE: Microarray analysis identifies candidate genes for key roles in coral development. BMC Genomics. 2008, 9: 540-10.1186/1471-2164-9-540.PubMed CentralPubMedGoogle Scholar
  28. Bay LK, Ulstrup KE, Nielsen HB, Jarmer H, Goffard N, Willis BL, Miller DJ, van Oppen MJ: Microarray analysis reveals transcriptional plasticity in the reef building coral Acropora millepora. Mol Ecol. 2009, 18: 3062-3075. 10.1111/j.1365-294X.2009.04257.x.PubMedGoogle Scholar
  29. Meyer E, Aglyamova GV, Wang S, Buchanan-Carter J, Abrego D, Colbourne JK, Willis BL, Matz MV: Sequencing and de novo analysis of a coral larval transcriptome using 454 GSFlx. BMC Genomics. 2009, 10: 219-10.1186/1471-2164-10-219.PubMed CentralPubMedGoogle Scholar
  30. DeSalvo MK, Sunagawa S, Voolstra CR, Medina M: Transcriptomic responses to heat stress and bleaching in the elkhorn coral Acropora palmata. Mar Ecol Prog Ser. 2010, 402: 97-113.Google Scholar
  31. Portune KJ, Voolstra CR, Medina M, Szmant AM: Development and heat stress induced transcriptomic changes during embryogenesis of the scleractinian coral Acropora palmata. Mar Genom. 2010, 3: 51-62. 10.1016/j.margen.2010.03.002.Google Scholar
  32. Souter P, Bay LK, Andreakis N, Császár N, Seneca FO, van Oppen MJ: A multilocus, temperature stress-related gene expression profile in Acropora millepora, a dominant reef-building coral. Mol Ecol Resour. 2011, 11: 328-334. 10.1111/j.1755-0998.2010.02923.x.PubMedGoogle Scholar
  33. Ladner JT, Barshis DJ, Palumbi SR:Protein evolution in two co-occurring types of Symbiodinium: an exploration into the genetic basis of thermal tolerance inSymbiodiniumclade D.BMC Evol Biol. 2012, 12: 217-10.1186/1471-2148-12-217.PubMed CentralPubMedGoogle Scholar
  34. Barshis DJ, Ladner JT, Oliver TA, Seneca FO, Traylor-Knowles N, Palumbi SR: Genomic basis for coral resilience to climate change. Proc Natl Acad Sci USA. 2013, 110: 1387-1392. 10.1073/pnas.1210224110.PubMed CentralPubMedGoogle Scholar
  35. Granados-Cifuentes C, Bellantuono AJ, Ridgway T, Hoegh-Guldberg O, Rodriguez-Lanetty M: High natural gene expression variation in the reef-building coralAcropora millepora: potential for acclimative and adaptive plasticity. BMC Genomics. 2013, 14: 228-10.1186/1471-2164-14-228.PubMed CentralPubMedGoogle Scholar
  36. Traylor-Knowles N, Granger BR, Lubinski TJ, Parikh JR, Garamszegi S, Xia Y, Marto JA, Kaufman L, Finnerty JR: Production of a reference transcriptome and transcriptomic database (PocilloporaBase) for the cauliflower coral, Pocillopora damicornis. BMC Genomics. 2011, 12: 585-10.1186/1471-2164-12-585.PubMed CentralPubMedGoogle Scholar
  37. Sun J, Chen Q, Lun JC, Xu J, Qiu JW: PcarnBase: Development of a transcriptomic database for the brain coral Platygyra carnosus. Mar Biotechnol. 2013, 15: 244-451. 10.1007/s10126-012-9482-z.PubMedGoogle Scholar
  38. Cossins A, Fraser J, Hughes M, Gracey A: Post-genomic approaches to understanding the mechanisms of environmentally induced phenotypic plasticity. J Exp Biol. 2006, 209: 2328-2336. 10.1242/jeb.02256.PubMedGoogle Scholar
  39. Weston AJ, Dunlap WC, Shick JM, Klueter A, Iglic K, Vukelic A, Starcevic A, Ward M, Wells ML, Trick CG, Long PF: A profile of an endosymbiont-enriched fraction of the coral Stylophora pistillata reveals proteins relevant to microbial-host interactions. Mol Cell Proteomics. 2012, 11: M111.015487-10.1074/mcp.M111.015487.PubMed CentralPubMedGoogle Scholar
  40. Starcevic A, Dunlap WC, Cullum J, Shick JM, Hranueli D, Long PF: Gene expression in the scleractinian Acropora microphthalma exposed to high solar irradiance reveals elements of photoprotection and coral bleaching. PLoS One. 2010, 5: e13975-10.1371/journal.pone.0013975.PubMed CentralPubMedGoogle Scholar
  41. Miller DJ, Ball EE, Technau U: Cnidarians and ancestral genetic complexity in the animal kingdom. Trends Genet. 2005, 21: 536-539. 10.1016/j.tig.2005.08.002.PubMedGoogle Scholar
  42. Putnam NH, Srivastava M, Hellsten U, Dirks B, Chapman J, Salamov A, Terry A, Shapiro H, Lindquist E, Kapitonov VV, Jurka J, Genikhovich G, Grigoriev IV, Lucas SM, Steele RE, Finnerty JR, Technau U, Martindale MQ, Rokhsar DS: Sea anemone genome reveals ancestral eumetazoan gene repertoire and genomic organization. Science. 2007, 317: 86-94. 10.1126/science.1139158.PubMedGoogle Scholar
  43. Kortschak RD, Samuel G, Saint R, Miller DJ: EST analysis of the cnidarians Acropora millepora reveals extensive gene loss and rapid sequence divergence in the model invertebrates. Curr Biol. 2003, 13: 2190-2195. 10.1016/j.cub.2003.11.030.PubMedGoogle Scholar
  44. Chapman JA, Kirkness EF, Simakov O, Hampson SE, Mitros T, Weinmaier T, Rattei T, Balasubramanian PG, Borman J, Busam D, Disbennett K, Pfannkoch C, Sumin N, Sutton GG, Viswanathan LD, Walenz B, Goodstein DM, Hellsten U, Kawashima T, Prochnik SE, Putnam NH, Shu S, Blumberg B, Dana CE, Gee L, Kibler DF, Law L, Lindgens D, Martinez DE, Peng J, Wigge PA:The dynamic genome of theHydra. Nature. 2010, 464: 591-596.Google Scholar
  45. Shinzato C, Shoguchi E, Kawashima T, Hamada M, Hisata K, Tanaka M, Fujie M, Fujiwara M, Koyanagi R, Ikuta T, Fujiyama A, Miller DJ, Satoh N: Using the Acropora digitifera genome to understand coral responses to environmental change. Nature. 2011, 476: 320-323. 10.1038/nature10249.PubMedGoogle Scholar
  46. Coral genome sequence data usage policy. [http://coralbase.org/accounts/register/]
  47. ZoophyteBase coral proteome database. [http://bioserv7.bioinfo.pbf.hr/Zoophyte/index.jsp]
  48. Genome Sequencing/Annotation Projects. http://marinegenomics.oist.jp/genomes/download?%20project_id=3]
  49. Finn RD, Clements J, Eddy SR: HMMER web server: interactive sequence similarity searching. Nucleic Acids Res. 2011, 39 (Web Server Issue): W29-W37.PubMed CentralPubMedGoogle Scholar
  50. Camacho C, Coulouris G, Avagyan V, Ma N, Papadopoulos J, Bealer K, Madden TL: BLAST+: architecture and applications. BMC Bioinforma. 2009, 10: 421-10.1186/1471-2105-10-421.Google Scholar
  51. KEGG: Kyoto Encyclopedia of Genes and Genomes. [http://www.genome.jp/kegg/]
  52. Apache Solr™. [http://lucene.apache.org/solr/]
  53. Conserved Domain Database(CDD). [http://www.ncbi.nlm.nih.gov/Structure/cdd/cdd.shtml]
  54. Stanley GD: Photosymbiosis and the evolution of modern coral reefs. Science. 2006, 312: 857-858. 10.1126/science.1123701.PubMedGoogle Scholar
  55. Dubinski Z, Falkowski P: Light as a source of information and energy in zooxanthellate corals. Coral Reefs: And Ecosystem in Transition. Edited by: Dubinski Z, Stambler N. 2011, Berlin: Springer-Verlag, 107-118.Google Scholar
  56. Dunn SR, Schnitzler CE, Wies VM: Apoptosis and autophagy as mechanisms of dinoflagellate symbiont release during cnidarian bleaching: every which way you lose. Proc Biol Soc. 2007, 274: 3079-3085. 10.1098/rspb.2007.0711.Google Scholar
  57. Downs CA, Kramarsky-Winter E, Martinez J, Kushmaro A, Woodley CM, Loya Y, Ostrander GK: Symbiophagy as a cellular mechanism for coral bleaching. Autophagy. 2009, 5: 211-216. 10.4161/auto.5.2.7405.PubMedGoogle Scholar
  58. Muscatine L, Pool RR: Regulation of numbers of intracellular alage. Proc R Soc B. 1979, 204: 131-139. 10.1098/rspb.1979.0018.Google Scholar
  59. Hutagalung AH, Novick PJ: Role of Rab GTPases in membrane traffic and cell physiology. Physiol Rev. 2011, 91: 119-149. 10.1152/physrev.00059.2009.PubMed CentralPubMedGoogle Scholar
  60. Hong M-C, Huang Y-S, Lin W-W, Fang L-S, Chen M-C: ApRab3, a biosynthetic Rab protein, accumulates on the maturing phagosomes and symbiosomes in the tropical sea anemone, Aiptasia pulchella. Comp Biochem Physiol B Biochem Mol Biol. 2009, 152: 249-259. 10.1016/j.cbpb.2008.12.005.PubMedGoogle Scholar
  61. Hong M-C, Huang J-S, Song P-C, Lin W-W, Fang L-S, Chen M-C: Cloning and characterization of ApRab4, a recycling Rab protein of Aiptasia pulchella, and its implication in the symbiosome biogenesis. Mar Biotechnol. 2009, 11: 771-785. 10.1007/s10126-009-9193-2.PubMedGoogle Scholar
  62. Chen MC, Cheng YM, Hong MC, Fang LS: Molecular cloning of Rab5 (ApRab5) in Aiptasia pulchella and its retention in phagosomes harbouring live zooxanthellae. Biochem Biophys Res Commun. 2004, 324: 1024-1033. 10.1016/j.bbrc.2004.09.151.PubMedGoogle Scholar
  63. Chen MC, Cheng YM, Sung PJ, Kuo CE, Fang LS: Molecular identification of Rab7 (ApRab7) in Aiptasia pulchella and its exclusion from phagosomes harboring zooxanthellae. Biochem Biophys Res Commun. 2003, 308: 586-595. 10.1016/S0006-291X(03)01428-1.PubMedGoogle Scholar
  64. Chen MC, Hong MC, Huang YS, Liu MC, Cheng YM, Fang LS: ApRab11, a cnidarian homologue of the recycling regulatory protein Rab11, is involved in the establishment and maintenance of the Aiptasia-Symbiodinium endosymbiosis. Biochem Biophys Res Commun. 2005, 338: 1607-1616. 10.1016/j.bbrc.2005.10.133.PubMedGoogle Scholar
  65. Collins RN: Rab and ARF GTPase regulation of exocytosis. Mol Membr Biol. 2003, 20: 105-115. 10.1080/0968768031000085892.PubMedGoogle Scholar
  66. Deneka M, Neeft M, van der Sluijs P: Regulation of membrane transport by Rab GTPases. Crit Rev Biochem Mol Biol. 2003, 38: 121-142. 10.1080/713609214.PubMedGoogle Scholar
  67. Vassilieva EV, Nusrat A: Vesicular trafficking: molecular tools and targets. Methods Mol Biol. 2008, 440: 3-14. 10.1007/978-1-59745-178-9_1.PubMedGoogle Scholar
  68. Ohya T, Miaczynska M, Coskun Ü, Lommer B, Runge A, Drechsel D, Kalaidzidis Y, Zerial M: Reconstruction of Rab- and SNARE-dependent membrane fusion by synthetic endosomes. Nature. 2009, 459: 1091-1097. 10.1038/nature08107.PubMedGoogle Scholar
  69. SNARE Database. [http://bioinformatics.mpibpc.mpg.de/snare/snareQueryPage.jsp]
  70. Kloepper TH, Kienle CN, Fasshauer D: An elaborate classification of SNARE proteins sheds light on the conservation of the eukaryotic endomembrane system. Mol Biol Cell. 2007, 18: 3463-3471. 10.1091/mbc.E07-03-0193.PubMed CentralPubMedGoogle Scholar
  71. Hay JC, Chao DS, Kuo CS, Scheller RH: Protein interactions regulating vesicle transport between the endoplasmic reticulum and Golgi apparatus in mammalian cells. Cell. 1997, 89: 149-158. 10.1016/S0092-8674(00)80191-9.PubMedGoogle Scholar
  72. Masuda ES, Huang BC, Fisher JM, Luo Y, Scheller RH: Tomosyn binds t-SNARE proteins via a VAMP-like coiled coli. Neuron. 1998, 21: 479-480. 10.1016/S0896-6273(00)80559-0.PubMedGoogle Scholar
  73. Scales SJ, Hesser BA, Masuda ES, Scheller RH: Amisyn, a novel syntaxin-binding protein that may regulate SNARE complex assembly. J Biol Chem. 2002, 227: 28271-28279.Google Scholar
  74. Jahn R, Scheller RH: SNAREs – Engines for membrane fusion. Nat Rev Mol Cell Biol. 2006, 7: 631-643. 10.1038/nrm2002.PubMedGoogle Scholar
  75. Muscatine L, Pool RR, Cernichiari E: Some factors influencing selective release of soluble organic material by zooxanthellae from reef corals. Mar Biol. 1972, 13: 298-308. 10.1007/BF00348077.Google Scholar
  76. Sutton DC, Hoegh-Guldberg O: Host-zooxanthellae interactions in four temperate marine invertebrate symbioses: assessment of the effect of host extracts on symbionts. Biol Bull. 1990, 178: 175-186. 10.2307/1541975.Google Scholar
  77. Masuda K, Miyachi S, Maruyama T: Sensitivity of zooxanthellae and non-symbiotic microalgae to stimulation of photosynthate excretion by giant clam tissue homogenate. Mar Biol. 1994, 118: 687-693. 10.1007/BF00347517.Google Scholar
  78. Narayanan A, Nogueira ML, Ruyechan WT, Kristie TM: Combinatorial transcription of Herpes simplex virus and Varicella zoster virus intermediate early genes is strictly determined by cellular coactivator HCF-1. J Biol Chem. 2005, 280: 1369-1375.PubMedGoogle Scholar
  79. Lee S, Horn V, Julien E, Liu Y, Wysocka J, Bowerman B, Hengartner MO, Herr W:Epigenetic regulation of histone H3 serine 10 phosphorylation status by HCF-1 proteins inC. elegansand mammalian cells. PLoS One. 2007, 2: e1213-10.1371/journal.pone.0001213.PubMed CentralPubMedGoogle Scholar
  80. Kristie TM, Liang Y, Vogel JL: Control of α-herpesvirus IE gene expression by HCF-1 coupled chromatin modification activities. Biochem Biophys Acta. 2010, 1799: 257-265. 10.1016/j.bbagrm.2009.08.003.PubMed CentralPubMedGoogle Scholar
  81. Christensen RG, Enuameh MS, Noyes MB, Brodsky MH, Wolfe SA, Stormo GD: Recognition models to predict DNA-binding specificities of homeodomain proteins. Bioinformatics. 2012, 28: i84-i89. 10.1093/bioinformatics/bts202.PubMed CentralPubMedGoogle Scholar
  82. Mann RS, Lelli KM, Joshi R: Hox specificity: unique roles for cofactors and collaborators. Curr Top Dev Biol. 2009, 88: 66-101.Google Scholar
  83. Finnerty JR, Martindale MQ: Ancient origins of axial patterning genes: Hox genes and ParaHox genes in the Cnidaria. Evol Dev. 1999, 1: 16-23. 10.1046/j.1525-142x.1999.99010.x.PubMedGoogle Scholar
  84. Hislop NR, de Jong D, Hayward DC, Ball EE, Miller DJ: Tandem organisation of independently duplicated homeobox genes in the basal cnidarian Acropora millepora. Dev Genes Evol. 2005, 215: 268-273. 10.1007/s00427-005-0468-y.PubMedGoogle Scholar
  85. Ryan JF, Mazza ME, Pang K, Matus DQ, Baxevanis AD, Martindale MQ, Finnerty JR:Pre-bilaterian origins of the Hox cluster and the Hox code: evidence from the sea anemone,Nematostella vectensis. PLoS One. 2007, 2: e153-10.1371/journal.pone.0000153.PubMed CentralPubMedGoogle Scholar
  86. Larroux C, Fahey B, Degnan SM, Adamski M, Rokhsar DS, Degnan BM: The NK homeobox gene cluster predates the origin of Hox genes. Curr Biol. 2007, 17: 706-710.PubMedGoogle Scholar
  87. Dowid IB, Chitnis AB: LIM homeobox genes and the CNS: a close relationship. Neuron. 2001, 30: 301-303. 10.1016/S0896-6273(01)00307-5.Google Scholar
  88. Srivastava M, Larroux C, Lu CD, Mohanty K, Chapman J, Degnan BM, Rokhsar DS: Early evolution of the LIM homeobox gene family. BMC Biol. 2010, 8: 4-10.1186/1741-7007-8-4.PubMed CentralPubMedGoogle Scholar
  89. Ryan JF, Burton PM, Mazza ME, Kwong GK, Mullikin JC, Finnerty JR:The cnidarian-bilaterian ancestor possessed at least 56 homeoboxes: evidence from the starlet sea anemone,Nematostella vectensis. Genome Biol. 2006, 7: R64-10.1186/gb-2006-7-7-r64.PubMed CentralPubMedGoogle Scholar
  90. Saito T, Sawamoto K, Okano H, Anderson DJ, Mikoshiba K: Mammalian BarH homologue is a potential regulator of neural bHLH genes. Dev Biol. 1998, 199: 216-225. 10.1006/dbio.1998.8889.PubMedGoogle Scholar
  91. Habas S, Kato Y, He X: Wnt/Frazzled activation of Rho regulates vertebrate gastrulation and requires a novel Formin homology protein Daam1. Cell. 2001, 107: 843-854. 10.1016/S0092-8674(01)00614-6.PubMedGoogle Scholar
  92. Matusek T, Djiane A, Jankovics F, Brunner D, Mlodzik M, Mihály J: The Drosophila formin DAAM regulates the tracheal cuticle pattern through organizing the actin cytoskeleton. Development. 2006, 133: 957-966. 10.1242/dev.02266.PubMedGoogle Scholar
  93. Li D, Hallett MA, Zhu W, Rubart M, Liu Y, Yang Z, Chen H, Haneline LS, Chan RJ, Schwartz RJ, Field LJ, Atkinson SJ, Shou W: Dishevelled-associated activator of morphogenesis 1 (Daam1) is required for heart morphogenesis. Development. 2011, 138: 303-315. 10.1242/dev.055566.PubMed CentralPubMedGoogle Scholar
  94. Wada Y, Kitamoto K, Kanbe T, Tanaka K, Anraku Y: The SLP1 gene of Saccharomyces cerevisiae is essential for vacuolar morphogenesis and function. Mol Cell Biol. 1990, 10: 2214-2223.PubMed CentralPubMedGoogle Scholar
  95. Bragdon B, Moseychuk O, Saldanha S, King D, Julian J, Nohe A: Bone morphogenetic proteins: a critical review. Cell Signal. 2011, 23: 609-620. 10.1016/j.cellsig.2010.10.003.PubMedGoogle Scholar
  96. Martin VJ: Photoreceptors of cnidarians. Can J Zool. 2002, 80: 1703-1722. 10.1139/z02-136.Google Scholar
  97. Mason BM, Cohen JH: Long-wavelength photosensitivity in coral planula larvae. Biol Bull. 2012, 222: 88-92.PubMedGoogle Scholar
  98. Gorbunov MY, Falkowski PG: Photoreceptors in the cnidarian host allow symbiotic corals to sense blue moonlight. Limnol Oceanogr. 2002, 47: 309-315. 10.4319/lo.2002.47.1.0309.Google Scholar
  99. Levy O, Appelbaum L, Leggat W, Gothlif Y, Hayward DC, Miller DJ, Hoegh-Guldberg O: Light-responsive cryptochromes from a simple multicellular animal, the coral Acropora millepora. Science. 2007, 318: 467-470. 10.1126/science.1145432.PubMedGoogle Scholar
  100. Vize PD: Transcriptome analysis of the circadian regulatory network in the coral Acropora millepora. Biol Bull. 2009, 216: 131-137.PubMedGoogle Scholar
  101. Brady AK, Snyder KA, Vize PD:Circadian cycles of gene expression in the coral,Acropora millepora. PLoS One. 2011, 6: e25072-10.1371/journal.pone.0025072.PubMed CentralPubMedGoogle Scholar
  102. Ishiura M, Kutsuna S, Aoki S, Iwasaki H, Andersson CR, Tanabe A, Golden SS, Johnson CH, Kondo T: Expression of a gene cluster kaiABC as a circadian feedback process in cyanobacteria. Science. 1998, 281: 1519-1523.PubMedGoogle Scholar
  103. Nishiwaki T, Satomi Y, Nakajima M, Lee C, Kiyohara R, Kageyama H, Kitayama Y, Temamoto M, Yamaguchi A, Hijikata A, Go M, Iwasaki H, Takao T, Kondo T: Role of KaiC phosphorylation in the circadian clock system of Synechococcus elongatus PCC 7942. Proc Natl Acad Sci USA. 2004, 101: 13927-13932. 10.1073/pnas.0403906101.PubMed CentralPubMedGoogle Scholar
  104. Xu Y, Mori T, Qin X, Yan H, Egli M, Johnson CH: Intramolecular regulation of phosphorylation status of the circadian clock protein KaiC. PLoS One. 2009, 4: e7509-10.1371/journal.pone.0007509.PubMed CentralPubMedGoogle Scholar
  105. Nakahira Y, Katayama M, Miyashita H, Katsuna S, Iwasaki H, Oyama T, Kondo T: Global gene repression by KaiC as a master process of prokaryotic circadian system. Proc Natl Acad Sci USA. 2004, 101: 881-885. 10.1073/pnas.0307411100.PubMed CentralPubMedGoogle Scholar
  106. Kondo T, Ishiura M: The circadian clock of cyanobacteria. Bioessays. 2000, 22: 10-15. 10.1002/(SICI)1521-1878(200001)22:1<10::AID-BIES4>3.0.CO;2-A.PubMedGoogle Scholar
  107. Hardin PE, Hall JC, Rosbash M: Feedback of the Drosophila period gene product on circadian cycling of its messenger RNA levels. Nature. 1990, 343: 536-540. 10.1038/343536a0.PubMedGoogle Scholar
  108. Dunlap JC: Molecular bases for circadian clocks. Cell. 1999, 96: 271-290. 10.1016/S0092-8674(00)80566-8.PubMedGoogle Scholar
  109. Reitzel AM, Behrendt L, Tarrant AM: Light entrained rhythmic gene expression in the sea anemone Nematostella vectensis: the evolution of the animal circadian clock. PLoS One. 2010, 5: e12805-10.1371/journal.pone.0012805.PubMed CentralPubMedGoogle Scholar
  110. Gibbons IR: Cilia and flagella of eukaryotes. J Cell Biol. 1981, 91: 107s-124s. 10.1083/jcb.91.3.107s.PubMedGoogle Scholar
  111. Linck RW: Tektins and microtubules. Adv Mol Cell Biol. 1990, 3: 35-63.Google Scholar
  112. Fox LA, Sawin KE, Sale WS: Kinesin-related proteins in eukaryotic flagella. J Cell Sci. 1994, 107: 1545-1550.PubMedGoogle Scholar
  113. Amos LA: The tektin family of microtubule-stabilizing proteins. Genome Biol. 2008, 9: 229-PubMed CentralPubMedGoogle Scholar
  114. Kozminski KG, Beech PL, Rosenbaum JL: The Chlamydomonas kinesin-like protein FLA10 is involved in motility associated with the flagellar membrane. J Cell Biol. 1995, 131: 1517-1527. 10.1083/jcb.131.6.1517.PubMedGoogle Scholar
  115. Bartlett DH, Frantz BB, Matsumura P: Flagellar activators FlbB and FlaI: gene sequences and 5′ consensus sequences of operons under FlbB and FlaI control. J Bacteriol. 1988, 170: 1575-1581.PubMed CentralPubMedGoogle Scholar
  116. Aldridge P, Karlinsey J, Hughes KT: The type III secretion chaperone FlgN regulates flagellar assembly via a negative feedback loop containing its chaperone substrates FlgK and FlgL. Mol Microbiol. 2003, 49: 1333-1345. 10.1046/j.1365-2958.2003.03637.x.PubMedGoogle Scholar
  117. Ferris HU, Furukawa Y, Minamino T, Kroetz MB, Kihara M, Namba K, Macnab RM: FlhB regulates ordered export of flagellar components via autocleavage mechanisms. J Biol Chem. 2005, 280: 41236-41242. 10.1074/jbc.M509438200.PubMedGoogle Scholar
  118. Mattick JS: Type IV pili and twitching motility. Annu Rev Microbiol. 2002, 56: 289-314. 10.1146/annurev.micro.56.012302.160938.PubMedGoogle Scholar
  119. Starcevic A, Akthar S, Dunlap WC, Shick JM, Hranueli D, Cullum J, Long PF: Enzymes of the shikimate acid pathway encoded in the genome of a basal metazoan, Nematostella vectensis, have microbial origins. Proc Natl Acad Sci U S A. 2008, 105: 2533-2537. 10.1073/pnas.0707388105.PubMed CentralPubMedGoogle Scholar
  120. Rebbapragada A, Johnson MS, Harding GP, Zuccarelli AJ, Fletcher HM, Zhulin IB, Taylor BL: The Aer protein and the serine chemoreceptor Tsr independently sense intracellular energy levels and transduce oxygen, redox, and energy signals for Escherichia coli behavior. Proc Natl Acad Sci USA. 1997, 94: 10541-10546. 10.1073/pnas.94.20.10541.PubMed CentralPubMedGoogle Scholar
  121. Troemei ER, Chou JH, Dwyer ND, Colbert HA, Bargmann CI: Divergent seven transmembrane receptors are candidate chemosensory receptors in C. elegans. Cell. 1995, 83: 207-218. 10.1016/0092-8674(95)90162-0.Google Scholar
  122. Sen M, Shah A, Marsh L: Two types of alpha-factor receptor determinants for pheromone specificity in the mating-incompatible yeasts S. cerevisiae and S. kluyveri. Curr Genet. 1997, 31: 235-240. 10.1007/s002940050200.PubMedGoogle Scholar
  123. Gestwicki JE, Lamanna AC, Harshey RM, McCarter LL, Kiessling LL, Adler J: Evolutionary conservation of methyl accepting chemotaxis protein location in Bacteria and Archaea. J Bacteriol. 2000, 182: 6499-6502. 10.1128/JB.182.22.6499-6502.2000.PubMed CentralPubMedGoogle Scholar
  124. Capra EJ, Laub MT: Evolution of two-component signal transduction systems. Annu Rev Microbiol. 2012, 66: 325-347. 10.1146/annurev-micro-092611-150039.PubMed CentralPubMedGoogle Scholar
  125. Koretke KK, Lupas AN, Warren PV, Rosenberg M, Brown JR: Evolution of two-component signal transduction. Mol Biol Evol. 2000, 17: 1956-1970. 10.1093/oxfordjournals.molbev.a026297.PubMedGoogle Scholar
  126. DeVries ME, Kelvin AA, Xu L, Ran L, Robertson J, Kelvin DJ: Defining the origins and evolution of the chemokine/chemokine receptor system. J Immunol. 2006, 176: 401-415.PubMedGoogle Scholar
  127. Holland ND: Early central nervous system evolution: an era of skin brains?. Nat Rev Neurosci. 2003, 4: 617-627. 10.1038/nrn1175.PubMedGoogle Scholar
  128. Galliot B, Quiquand M, Ghila L, de Rosa R, Miljkovic-Licina M, Chera S: Origins of neurogenesis, a cnidarian view. Dev Biol. 2009, 332: 2-24. 10.1016/j.ydbio.2009.05.563.PubMedGoogle Scholar
  129. Nakanishi N, Renfer E, Technau U, Rentzsch F: Nervous systems of the sea anemone Nematostella vectensis are generated by ectoderm and endoderm and shaped by distinct mechanisms. Development. 2012, 139: 347-357. 10.1242/dev.071902.PubMedGoogle Scholar
  130. Sakarya O, Armstrong KA, Adamska M, Adamski M, Wang IF, Tidor B, Degnan BM, Oakley TH, Kosik KS: A post-synaptic scaffold at the origin of the animal kingdom. PLoS One. 2007, 2: e506-10.1371/journal.pone.0000506.PubMed CentralPubMedGoogle Scholar
  131. Miljkovik-Licina M, Gauchat D, Galliot B: Neuronal evolution: analysis of regulatory genes in a first-evolved nervous system, the hydra nervous system. Biosystems. 2004, 76: 75-87. 10.1016/j.biosystems.2004.05.030.Google Scholar
  132. Marlow HQ, Srivastava M, Matus DQ, Rokhsar D, Martindale MQ: Anatomy and development of the nervous system of Nematostella vectensis, an anthozoan cnidarian. Dev Neurobiol. 2009, 69: 235-254. 10.1002/dneu.20698.PubMedGoogle Scholar
  133. Kass-Simon G, Pierobon P: Cnidarian chemical neurotransmission, an updated overview. Comp Biochem Physiol A Mol Intergr Physiol. 2007, 146: 9-25. 10.1016/j.cbpa.2006.09.008.Google Scholar
  134. Grimmelikhuijzen CJP, Westfall JA: The nervous systems of cnidarians. The Nervous Systems of Invertebrates. An Evolutionary and Comparative Approach. Edited by: Breidbach O, Kutch W. 1995, Basel: Brikhäuser Verlag, 7-24.Google Scholar
  135. Sleeman MW, Anderson KD, Lambert PD, Yancopoulos GD, Wiegand SJ: The ciliary neurotrophic factor and its receptor, CNFRTA alpha. Pharm Acta Helv. 2000, 74: 265-272. 10.1016/S0031-6865(99)00050-3.PubMedGoogle Scholar
  136. Kass-Simon G, Scappaticci AA: The behavioural and developmental physiology of nematocysts. Can J Zool. 2002, 80: 1772-1794. 10.1139/z02-135.Google Scholar
  137. Plachetzki DC, Fong CR, Oakley TH: Cnidocyte discharge is regulated by light and opsin-mediated phototransduction. BMC Biol. 2012, 10: 17-10.1186/1741-7007-10-17.PubMed CentralPubMedGoogle Scholar
  138. Coates MM: Visual ecology and functional morphology of Cubozoa (Cnidaria). Integr Comp Biol. 2003, 43: 542-548. 10.1093/icb/43.4.542.PubMedGoogle Scholar
  139. Koyanagi M, Takano K, Tsukamoto H, Ohtsu K, Tokunaga F, Terakita A: Jellyfish vision starts with cAMP signalling mediated by opsin-GS cascade. Proc Natl Acad Sci U S A. 2008, 105: 15576-15580. 10.1073/pnas.0806215105.PubMed CentralPubMedGoogle Scholar
  140. Shichida Y, Matsuyama T: Evolution of opsins and phototransduction. Philos Trans R Soc Lond B Biol Sci. 2009, 364: 2881-2895. 10.1098/rstb.2009.0051.PubMed CentralPubMedGoogle Scholar
  141. Beliaev A, Learmonth DA, Soares-da-Silva P: Synthesis and biological evaluation of novel, peripherally selective chromanyl imidazolethione-based inhibitors of dopamine beta-hydroxylase. J Med Chem. 2006, 49: 1191-1197. 10.1021/jm051051f.PubMedGoogle Scholar
  142. Grimmelikhuijzen CJP, Williamson M, Hansen GN: Neuropeptides in cnidarians. Can J Zool. 2002, 80: 1690-1702. 10.1139/z02-137.Google Scholar
  143. Attenborough RM, Hayward DC, Kitahara MV, Miller DJ, Ball EE: A “neural” enzyme in nonbilaterian animals and algae: preneural origins for peptidylglycine α-amindating monooxygenase. Mol Biol Evol. 2012, 29: 3095-3109. 10.1093/molbev/mss114.PubMedGoogle Scholar
  144. Allemand D, Tambutté E, Zoccola D, Tambutté S: Coral calcification, cells to reefs. Coral Reefs: An Ecosystem in Transition. Edited by: Dubinsky Z, Stambler N. 2011, Dordrecht: Springer, 119-150.Google Scholar
  145. Bénazet-Tambutté S, Allemand D, Joubert J: Permeability of the oral epithelial layers in cnidarians. Mar Biol. 1996, 126: 43-53. 10.1007/BF00571376.Google Scholar
  146. Al-Horani FA, Al-Moghrabi SM, de Beer D: The mechanism of calcification and its relation to photosynthesis and respiration in the scleractinian coral Galaxea fascicularis. Mar Biol. 2003, 142: 419-429.Google Scholar
  147. Zoccola D, Tambutté E, Sénégas-Balas F, Michiels JF, Failla JP, Jaubert J, Allemand D: Cloning of a calcium channel α1 subunit from the reef-building coral, Stylophora pistillata. Gene. 1999, 227: 157-167. 10.1016/S0378-1119(98)00602-7.PubMedGoogle Scholar
  148. Zoccola D, Tambutté E, Kulhanek E, Puverel S, Scimeca JC, Allemand D, Tambutté S: Molecular cloning and localisation of a PMCA P-type calcium ATPase from the coral Stylophora pistillata. Biochim Biophys Acta. 2004, 1663: 117-126. 10.1016/j.bbamem.2004.02.010.PubMedGoogle Scholar
  149. Moya A, Tambutté S, Bertucci A, Tambutté E, Lotto S, Vullo D, Supuran CT, Allemand D, Zoccola D: Carbonic anydrase in the scleractinian coral Stylophora pistillata: characterisation, localisation, and role in biomineralisation. J Biol Chem. 2008, 283: 25475-25484. 10.1074/jbc.M804726200.PubMedGoogle Scholar
  150. Gattuso J-P, Allemand D, Frankignoulle M: Photosynthesis and calcification at cellular, organismal and community levels in coral reefs: A review on interactions and control by carbonate chemistry. Amer Zool. 1999, 39: 160-183.Google Scholar
  151. Allemand D, Ferrier-Pagè C, Furla P, Houlbrèque F, Puverel S, Reynaud S, Tambutté É, Tambutté S, Zoccola D: Biomineralisation in reef-building corals: from molecular mechanisms to environmental control. C R Palevol. 2004, 3: 453-467. 10.1016/j.crpv.2004.07.011.Google Scholar
  152. Venn A, Tambutté E, Holcomb M, Allemand D, Tambutté S: Live tissue imaging shows corals elevate pH under their calcifying tissues relative to seawater. PLoS One. 2011, 6: e20013-10.1371/journal.pone.0020013.PubMed CentralPubMedGoogle Scholar
  153. Kaniewska P, Campbell PR, Kline DI, Rodriguez-Lanetty M, Miller DJ, Dove S, Hoegh-Guldberg O: Major cellular and physiological impacts of ocean acidification on a reef building coral. PLoS One. 2012, 7: e34659-10.1371/journal.pone.0034659.PubMed CentralPubMedGoogle Scholar
  154. Catterall WA: Voltage-gated calcium channels. Cold Spring Harb Perspect Biol. 2011, 3: 003947-10.1101/cshperspect.a003947.Google Scholar
  155. Bertucci A, Tembutté S, Supuran CT, Allemand D, Zoccola D: A new coral carbonic anhydrase in Stylophora pistillata. Mar Biotechnol. 2011, 13: 992-1002. 10.1007/s10126-011-9363-x.PubMedGoogle Scholar
  156. Morita M, Iguchi A, Takamura A: Roles of calmodulin and calcium/calmodulin-dependent protein in flagellate motility regulation in the coral Acropora digitifera. Mar Biotechnol. 2009, 11: 118-123. 10.1007/s10126-008-9127-4.PubMedGoogle Scholar
  157. Lane CE, Archibald JM: The eukaryotic tree of life: endosymbionts takes its TOL. Trends Ecol Evol. 2008, 32: 268-275.Google Scholar
  158. Keeling PJ: Functional and ecological impacts of horizontal gene transfer in eukaryotes. Curr Opin Genet Dev. 2009, 19: 613-619. 10.1016/j.gde.2009.10.001.PubMedGoogle Scholar
  159. Bock R: The give-and-take of DNA: horizontal gene transfer in plants. Trends Plant Sci. 2010, 15: 11-22.PubMedGoogle Scholar
  160. Balskus EP, Walsh CT: The genetic and molecular basis for sunscreen biosynthesis in cyanobacteria. Science. 2010, 329: 1653-1656. 10.1126/science.1193637.PubMed CentralPubMedGoogle Scholar
  161. Waller RF, Stamovits CH, Keeling PJ: Lateral gene transfer of a multigene region from cyanobacteria to dinoflagellates resulting in a novel plastid-targeted fusion protein. Mol Biol Evol. 2006, 23: 1437-1443. 10.1093/molbev/msl008.PubMedGoogle Scholar
  162. Richards TA, Dacks JB, Campbell SA, Blanchard JL, Foster PG, McLeod R, Roberts CW: Evolutionary origins of the eukaryotic shikimate pathway: gene fusions, horizontal transfer, and endosymbiotic replacement. Eukaryot Cell. 2006, 5: 1517-1531. 10.1128/EC.00106-06.PubMed CentralPubMedGoogle Scholar
  163. Habetha M, Bosch TC: Symbiotic Hydra express a plant-like peroxidase gene during oogenesis. J Exp Biol. 2005, 208: 2157-2165. 10.1242/jeb.01571.PubMedGoogle Scholar
  164. Technau U, Miller MA, Bridge D, Steele RE: Arrested apoptosis of nurse cells during Hydra oogenesis and embryogenesis. Dev Biol. 2003, 260: 191-206. 10.1016/S0012-1606(03)00241-0.PubMedGoogle Scholar
  165. Rumpho ME, Worful JM, Lee J, Kannan K, Tyler MS, Bhattacharya D, Moustafa A, Manhart JR: Horizontal gene transfer of the algal nuclear gene psbO to the photosynthetic sea slug Elysia chlorotica. Proc Natl Acad Sci U S A. 2008, 105: 17867-17871. 10.1073/pnas.0804968105.PubMed CentralPubMedGoogle Scholar
  166. Rumpho ME, Pelletreau KN, Moustafa A, Bhattacharya D: The making of a photosynthetic animal. J Exp Biol. 2011, 214: 303-311. 10.1242/jeb.046540.PubMed CentralPubMedGoogle Scholar
  167. Pierce SK, Curtis NE: Cell biology of the chloroplast symbiosis in sacaglossan sea slugs. Int Rev Cell Mol Biol. 2012, 293: 123-148.PubMedGoogle Scholar
  168. Wägele H, Deusch O, Händeler K, Martin R, Schmitt V, Christa G, Pinzger B, Gould SB, Dagan T, Klussmann-Kolb A, Martin W: Transcriptomic evidence that longevity of acquired plastids in the photosynthetic slugs Elysia timida and Plakobranchus ocellatus does not entail lateral transfer of algal nuclear genes. Mol Biol Evol. 2011, 28: 699-706. 10.1093/molbev/msq239.PubMed CentralPubMedGoogle Scholar
  169. Pierce SK, Fang X, Schwartz JA, Jiang X, Zhao W, Curtis NE, Kocot KM, Yang B, Wang J: Transcriptomic evidence for the expression of horizontally transferred algal nuclear genes in the photosynthetic sea slug, Elysia chlorotica. Mol Biol Evol. 2012, 29: 1545-1556. 10.1093/molbev/msr316.PubMedGoogle Scholar
  170. Lang BF, O’Kelly C, Nerad T, Gray MW, Burger G: The closest unicellular relatives of animals. Curr Biol. 2002, 12: 1773-1778. 10.1016/S0960-9822(02)01187-9.PubMedGoogle Scholar
  171. Ruiz-Trillo I, Roger AJ, Burger G, Gray MW, Lang BF: A phylogenomic investigation into the origins of metazoa. Mol Biol Evol. 2008, 25: 664-672. 10.1093/molbev/msn006.PubMedGoogle Scholar
  172. Sun G, Yang Z, Ishwar A, Huang J: Algal genes in the closest relatives of animals. Mol Biol Evol. 2010, 27: 2879-2889. 10.1093/molbev/msq175.PubMedGoogle Scholar
  173. Ohad I, Dal Bosco C, Herrmann RG, Meurer J: Photosysytem II proteins PsbL and PsbJ regulate electron flow to the plastoquinone pool. Biochemistry. 2004, 43: 2297-2308. 10.1021/bi0348260.PubMedGoogle Scholar
  174. Yamazaki S, Nomata J, Fujita Y: Differential operation of dual protochlorophyllide reductases for chlorophyll biosynthesis in response to environmental oxygen levels in the cyanobacterium Leptolyngbya boryana. Plant Physiol. 2006, 142: 911-922. 10.1104/pp.106.086090.PubMed CentralPubMedGoogle Scholar
  175. Pruzinská A, Anders I, Aubry S, Schenk N, Tapernoux-Lüthi E, Müller T, Kräutler B, Hörtensteiner S: In vivo participation of red chlorophyll catabolite reductase in chlorophyll breakdown. Plant Cell. 2007, 19: 369-387. 10.1105/tpc.106.044404.PubMed CentralPubMedGoogle Scholar
  176. Harada J, Saga Y, Yaeda Y, Oh-Oka H, Tamiaki H: In vitro activity of C-20 methyltransferase, BchU, involved in bacteriochlorophyll c biosynthetic pathway in green sulfur bacteria. FEBS Lett. 2005, 579: 1983-1987. 10.1016/j.febslet.2005.01.087.PubMedGoogle Scholar
  177. Long H, King PW, Ghirardi ML, Kim K: Hydrogenase/ferredoxin charge-transfer complexes: effect of hydrogenase mutations on the complex association. J Phys Chem A. 2009, 113: 4060-4067. 10.1021/jp810409z.PubMedGoogle Scholar
  178. Spence E, Dunlap WC, Shick JM, Long PF: Redundant pathways of sunscreen biosynthesis in a cyanobacterium. ChemBioChem. 2012, 13: 531-533. 10.1002/cbic.201100737.PubMedGoogle Scholar
  179. Wegkamp A, van Oorschot W, de Vos WM, Smid EJ: Characterization of the role of para-aminobenzoic acid biosynthesis in folate production by Lactococcus lactis. Appl Environ Microbiol. 2007, 73: 2673-2681. 10.1128/AEM.02174-06.PubMed CentralPubMedGoogle Scholar
  180. Sharon I, Tzahor S, Williamson S, Shmoish M, Man-Aharonovich D, Rusch DB, Yooseph S, Zeidner G, Golden SS, Mackey SR, Adir N, Weingart U, Horn D, Venter JC, Mandel-Gutfreund YM, Béjà O: Viral photosynthetic reaction center genes and transcripts in the marine environment. ISME J. 2007, 1: 492-501. 10.1038/ismej.2007.67.PubMedGoogle Scholar
  181. Wang Q, Jantaro S, Lu B, Majeed W, Bailey M, He Q: The high light-inducible polypeptides stabilize trimeric photosystem I complex under high light conditions in Synechocystis PCC 6803. Plant Physiol. 2008, 147: 1239-1250. 10.1104/pp.108.121087.PubMed CentralPubMedGoogle Scholar
  182. Mann NH, Cook A, Millard A, Bailey S, Clokie M: Marine ecosystems: bacterial photosynthesis genes in a virus. Nature. 2003, 424: 741-PubMedGoogle Scholar
  183. Lindell D, Sullivan MB, Johnson ZI, Tolonen AC, Rohwer F, Chisholm SW: Transfer of photosynthesis genes to and from Prochlorococcus viruses. Proc Natl Acad Sci U S A. 2004, 101: 11013-11018. 10.1073/pnas.0401526101.PubMed CentralPubMedGoogle Scholar
  184. Mann NH, Clokie MRJ, Millard A, Cook A, Wilson WH, Wheatley PJ, Letarov A, Krisch HM: The genome of S-PM2, a “photosynthetic” T4-type bacteriophage that infects marine Synechococcus strains. J Bacteriol. 2005, 187: 3188-3200. 10.1128/JB.187.9.3188-3200.2005.PubMed CentralPubMedGoogle Scholar
  185. van Oppen MJH, Leong J-A, Gates RD: Coral-virus interactions: a double-edged sword?. Symbiosis. 2009, 47: 1-8. 10.1007/BF03179964.Google Scholar
  186. Nagasaki K, Tomaru Y, Shirai Y, Takano Y, Mizumoto H: Dinoflagellate-infecting viruses. J Mar Biol Assoc UK. 2006, 86: 469-474. 10.1017/S0025315406013361.Google Scholar
  187. Lohr J, Munn CB, Wilson WH: Characterization of a latent virus-like infection of symbiotic zooxanthellae. Appl Environ Microbiol. 2007, 73: 2976-2981. 10.1128/AEM.02449-06.PubMed CentralPubMedGoogle Scholar
  188. Moran NA: Symbiosis as an adaptive process and source of phenotypic complexity. Proc Natl Acad Sci U S A. 2007, 104 (Suppl 1): 8627-8633.PubMed CentralPubMedGoogle Scholar
  189. Falkowski PG, Dubinsky Z, Muscatine L, Porter JW: Light and the bioenergentics of a symbiotic coral. Bioscience. 1984, 34: 705-709. 10.2307/1309663.Google Scholar
  190. Muscatine L, D’Elia CF: The uptake, retention and release of ammonium by reef corals. Limnol Oceanogr. 1978, 23: 725-734. 10.4319/lo.1978.23.4.0725.Google Scholar
  191. Rahav O, Dubinsky Z, Achituv Y, Falkowski PG: Ammonium metabolism in the zooxanthellate coral, Stylophora pistillata. Proc R Soc Lond B. 1989, 236: 325-337. 10.1098/rspb.1989.0026.Google Scholar
  192. Kawaguti S: Ammonium metabolism of the reef corals. Biol J Okayama Univ. 1953, 1: 171-176.Google Scholar
  193. Kühl M, Cohen Y, Dalsgaard T, Jørgensen BB, Revsbech NP: Microenvironment and photosynthesis of zooxanthellae in scleractinian corals studied with microsensors for O2, pH and light. Mar Ecol Prog Ser. 1995, 117: 159-172.Google Scholar
  194. Gallon JR: The oxygen sensitivity of nitrogenase: a problem for biochemists and micro-organisms. Trends Biochem Sci. 1981, 6: 19-23.Google Scholar
  195. Webb KL, DuPaul WD, Wiebe W, Sottile W, Johannes RE: Enewetak (Eniwetok) atoll: aspects of the nitrogen cycle on a coral reef. Limnol Oceanogr. 1975, 20: 198-210. 10.4319/lo.1975.20.2.0198.Google Scholar
  196. Crossland CJ, Barnes DJ: Dissolved nutrients and organic particulates in water flowing over coral reefs at Lizard Island. Aust J Mar Freshwat Res. 1983, 34: 835-844. 10.1071/MF9830835.Google Scholar
  197. Wilkinson CR, Williams DMB, Sommarco PW, Hogg RW, Trott LA: Rates of nitrogen fixation on coral reefs across the continental shelf of the central Great Barrier Reef. Reef Mar Biol. 1984, 80: 255-262. 10.1007/BF00392820.Google Scholar
  198. Capone DG, Dunham SE, Horrigan SG, Duguay LE: Microbial nitrogen transformations in unconsolidated coral reef sediments. Mar Ecol Prog Ser. 1992, 80: 75-88.Google Scholar
  199. Hewson I, Moisander PH, Morrison AE, Zehr JP: Diazotrophic bacterioplankton in a coral reef lagoon: phylogeny, diel nitrogenase expression and response to phosphate enrichment. ISME J. 2007, 1: 78-91. 10.1038/ismej.2007.5.PubMedGoogle Scholar
  200. Larkum AWD, Kennedy IR, Muller WJ: Nitrogen fixation on a coral reef. Mar Biol. 1988, 98: 143-155. 10.1007/BF00392669.Google Scholar
  201. Shashar N, Cohen Y, Loya Y, Sar N: Nitrogen fixation (acetylene reduction) in stony corals: evidence for coral-bacteria interactions. Mar Ecol Prog Ser. 1994, 111: 259-264.Google Scholar
  202. Lesser MP, Mazel CH, Gorbunov MY, Falkowski PG: Discovery of symbiotic nitrogen-fixing cyanobacteria in corals. Science. 2004, 305: 997-1000. 10.1126/science.1099128.PubMedGoogle Scholar
  203. Mouchka ME, Hewson I, Harvell CD: Coral-associated bacterial assemblages: current knowledge and the potential for climate-driven impacts. Integr Comp Biol. 2010, 50: 662-674. 10.1093/icb/icq061.PubMedGoogle Scholar
  204. Olson ND, Ainsworth TD, Gates RD, Takabayashi M: Diazotrophic bacteria associated with Hawaiian Montipora coral: diversity and abundance in correlation with symbiotic dinoflagellates. J Exp Mar Biol Ecol. 2009, 371: 140-146. 10.1016/j.jembe.2009.01.012.Google Scholar
  205. Lema KA, Willis BL, Bourne DG: Corals form characteristic associations with symbiotic nitrogen-fixing bacteria. Appl Environ Microbiol. 2012, 78: 3136-3144. 10.1128/AEM.07800-11.PubMed CentralPubMedGoogle Scholar
  206. Kellogg CA: Tropical Archaea: diversity associated with the surface microlayer of corals. Mar Ecol Prog Ser. 2004, 273: 81-88.Google Scholar
  207. Wegley L, Edwards R, Rodriguez-Brito B, Liu H, Rohwer F: Metagenomic analysis of the microbial community associated with the coral Porites astreoides. Environ Microbiol. 2007, 9: 2707-2719. 10.1111/j.1462-2920.2007.01383.x.PubMedGoogle Scholar
  208. Siboni N, Ben-Dov E, Silvan A, Kushmaro A: Global distribution and diversity of coral-associated Archaea and their possible role in the coral holobiont nitrogen cycle. Environ Microbiol. 2008, 10: 2979-2990. 10.1111/j.1462-2920.2008.01718.x.PubMedGoogle Scholar
  209. Rubio LM, Ludden PW: Biosynthesis of the iron-molybdenum cofactor of nitrogenase. Annu Rev Microbiol. 2008, 62: 93-111. 10.1146/annurev.micro.62.081307.162737.PubMedGoogle Scholar
  210. Fujita Y, Bauer CE: Reconstitutuion of the light-independent protochlorophyllide reductase from purified Bchl and BchN-BchB subunits. In vitro confirmation of nitrogenase features of a bacteriochlorophyll biosynthesis enzyme. J Biol Chem. 2000, 275: 23583-23588. 10.1074/jbc.M002904200.PubMedGoogle Scholar
  211. Sarma R, Barney BM, Hamilton TL, Jones A, Seefeld LC, Peters JW: Crystal structure of the L protein of Rhodobacter sphaeroides light-independent protochlorophyllide reductase with MgADP bound: a homologue of the nitrogenase Fe protein. Biochemistry. 2008, 47: 13004-13015. 10.1021/bi801058r.PubMedGoogle Scholar
  212. Muraki N, Nomata J, Ebata K, Mizoguchi T, Shiba T, Tamiaki H, Kurisu G, Fujita Y: X-ray crystal structure of the light-independent protochlorophyllide reductase. Nature. 2010, 465: 110-114. 10.1038/nature08950.PubMedGoogle Scholar
  213. Heyes DJ, Hunter CN: Making light work of enzyme catalysis: protochlorophyllide oxidoreductase. Trends Biochem Sci. 2005, 30: 642-649. 10.1016/j.tibs.2005.09.001.PubMedGoogle Scholar
  214. Dean DR, Bolin JT, Zheng L: Nitrogenase metalloclusters: structure, organisation, and synthesis. J Bacteriol. 1993, 175: 6737-6744.PubMed CentralPubMedGoogle Scholar
  215. Zheng L, Dean DR: Catalytic formation of a nitrogenase iron-sulfur cluster. J Biol Chem. 1994, 269: 18723-18726.PubMedGoogle Scholar
  216. Shah VK, Stacey G, Brill WJ: Electron transport to nitrogenase. J Biol Chem. 1983, 258: 12064-12068.PubMedGoogle Scholar
  217. Hoover TR, Santero E, Porter S, Kustu S: The integration host factor stimulates interaction of RNA polymerase with FIFA, the transcriptional activator for nitrogen fixation operons. Cell. 1990, 63: 11-22. 10.1016/0092-8674(90)90284-L.PubMedGoogle Scholar
  218. Merrick MJ, Edwards RA: Nitrogen control in bacteria. Microbiol Mol Biol Rev. 1995, 59: 604-622.Google Scholar
  219. Kneip C, Lockhart P, Voss C, Maier UG: Nitrogen fixation in eukaryotes – new models for symbiosis. BMC Evol Biol. 2007, 7: 55-10.1186/1471-2148-7-55.PubMed CentralPubMedGoogle Scholar
  220. Beman JM, Roberts KJ, Wegley L, Rohwer F, Francis CA: Distribution and diversity of archaeal ammonia momooxygenase genes associated with corals. Appl Environ Microbiol. 2007, 73: 5642-5647. 10.1128/AEM.00461-07.PubMed CentralPubMedGoogle Scholar
  221. Campbell WH: Nitrate reductase structure, function and regulation: Bridging the gap between biochemistry and physiology. Annu Rev Plant Physiol Mol Biol. 1999, 50: 277-303. 10.1146/annurev.arplant.50.1.277.Google Scholar
  222. Einsle O: Structure and function of formate-dependent cytochrome c nitrite reductase, NrfA. Methods Enzymol. 2011, 496: 399-422.PubMedGoogle Scholar
  223. Holden HM, Thoden JB, Raushel FM: Carbamoyl phosphate synthetase: an amazing biochemical odyssey from substrate to product. Cell Mol Life Sci. 1999, 56: 507-522. 10.1007/s000180050448.PubMedGoogle Scholar
  224. Crandall JB, Teece MA: Urea is a dynamic pool of bioavailable nitrogen in coral reefs. Coral Reefs. 2012, 31: 207-214. 10.1007/s00338-011-0836-1.Google Scholar
  225. Catmull J, Yellowlees D, Miller DJ: NADP+-dependent glutamate dehydrogenase from Acropora formosa: purification and properties. Mar Biol. 1987, 95: 559-563. 10.1007/BF00393099.Google Scholar
  226. Clode PL, Saunders M, Maker G, Ludwig M, Atkins CA: Uric acid deposits in symbiotic marine algae. Plant Cell Environ. 2009, 32: 170-177. 10.1111/j.1365-3040.2008.01909.x.PubMedGoogle Scholar
  227. Lancaster JR: Simulation of the diffusion and reaction of endogenously produced nitric oxide. Proc Natl Acad Sci USA. 1994, 91: 8137-8141. 10.1073/pnas.91.17.8137.PubMed CentralPubMedGoogle Scholar
  228. Trapido-Rosenthal H, Zielke S, Owen R, Buxon L, Boeing B, Bhagooli R, Archer J: Increased zooxanthellae nitric oxide synthase activity is associated with coral bleaching. Biol Bull. 2005, 208: 3-6. 10.2307/3593094.PubMedGoogle Scholar
  229. Bouchard JN, Yamasaki H: Heat stress stimulates nitric oxide production in Symbiodinium microadriaticum: a possible linkage between nitric oxide and the coral bleaching phenomenon. Plant Cell Physiol. 2008, 49: 641-652. 10.1093/pcp/pcn037.PubMedGoogle Scholar
  230. Perez S, Weis V: Nitric oxide and cnidarians bleaching: an eviction notice mediates breakdown of a symbiosis. J Exp Biol. 2006, 209: 2804-2810. 10.1242/jeb.02309.PubMedGoogle Scholar
  231. Safavi-Hemami H, Young ND, Doyle J, Llewellyn L, Klueter A: Characterisation of nitric oxide synthase in three cnidarian-dinoflagellate symbioses. PLoS One. 2010, 5: e10379-10.1371/journal.pone.0010379.PubMed CentralPubMedGoogle Scholar
  232. Dreyer J, Schleicher M, Tappe A, Schilling K, Kuner T, Kusumawidijaja G, Müller-Esterl W, Oess S, Kuner R: Nitric oxide synthase (NOS)-interacting protein interacts with neuronal NOS and regulates its distribution and activity. J Neurosci. 2004, 24: 10454-10465. 10.1523/JNEUROSCI.2265-04.2004.PubMedGoogle Scholar
  233. Siebeck O: Photoactivation and depth-dependent UV tolerance in reef coral in the Great Barrier Reef/Australia. Naturwissenschaften. 1981, 68: 426-428. 10.1007/BF01079713.Google Scholar
  234. Reef R, Dunn S, Levy O, Dove S, Shemesh E, Brickner I, Leggat W, Hoegh-Guldberg O: Photoreactivation is the main repair pathway for UV-induced DNA damage in coral plenulae. J Exp Biol. 2009, 212: 2760-2766. 10.1242/jeb.031286.PubMedGoogle Scholar
  235. Anderson S, Zepp R, Machula J, Santavy D, Hansen L, Mueller E: Indicators of UV exposure in corals and their relevance to global climate change and coral bleaching. Hum Ecol Risk Assess. 2001, 7: 1271-1782. 10.1080/20018091094998.Google Scholar
  236. Torregiani JH, Lesser MP: The effects of short-term exposures to ultraviolet radiation in the Hawaiian coral Montipora verrucosa. J Exp Mar Biol Ecol. 2007, 340: 194-203. 10.1016/j.jembe.2006.09.004.Google Scholar
  237. Baruch R, Avishai N, Rabinowitz C: UV incites diverse levels of DNA breaks in different cellular compartments of a branching coral species. J Exp Biol. 2005, 208: 843-848. 10.1242/jeb.01496.PubMedGoogle Scholar
  238. Hudson CL, Ferrier MD: Assessing ultraviolet radiation-induced DNA damage and repair in field-collected Aiptasia pallida using the comet assay. Proceedings of the 11th International Coral Reef Symposium. 2008, Florida, 7-11.Google Scholar
  239. Nesa B, Baird AH, Harii S, Yakovleva I, Hidaka M: Algal symbionts increase DNA damage in coral plenulae exposed to sunlight. Zool Stud. 2012, 51: 112-117.Google Scholar
  240. Vijayavel K, Downs CA, Ostrander GK, Richmond RH: Oxidative DNA damage induced by iron chloride in the larvae of the lace coral Pocillopora damicornis. Comp Biochem Physiol C Toxicol Pharmacol. 2012, 155: 275-280. 10.1016/j.cbpc.2011.09.007.PubMedGoogle Scholar
  241. Lesser MP, Farrell JH: Exposure to solar radiation increases damage to both host tissues and algal symbionts of corals during thermal stress. Coral Reefs. 2004, 23: 367-377. 10.1007/s00338-004-0392-z.Google Scholar
  242. Polato NR, Vera JC, Baums IB: Gene discovery in the threatened elkhorn coral: 454 sequencing of the Acropora palmata transcriptome. PLoS One. 2011, 6: e28634-10.1371/journal.pone.0028634.PubMed CentralPubMedGoogle Scholar
  243. Lindquist S, Craig EA: The heat-shock proteins. Annu Rev Genet. 1988, 22: 631-677. 10.1146/annurev.ge.22.120188.003215.PubMedGoogle Scholar
  244. Åkerfelt M, Morimoto RI, Sistonen L: Heat shock factors: Integrators of cell stress, development and lifespan. Nat Rev Mol Cell Biol. 2010, 11: 545-555. 10.1038/nrm2938.PubMed CentralPubMedGoogle Scholar
  245. Bosch TCG, Praetzel G: The heat shock response in hydra: immunological relationship of hsp60, the major heat shock protein of Hydra vulgaris, to the ubiquitous hsp70 family. Hydrobiologia. 1991, 216–217: 513-517.Google Scholar
  246. Choresh O, Ron E, Loya Y: The 60-kDa heat shock protein (HSP60) of the sea anemone Anemonia virdis; a potential early warning system for environmental change. Mar Biotechnol (NY). 2001, 3: 501-508. 10.1007/s10126-001-0007-4.Google Scholar
  247. Bromage E, Carpenter L, Kaattari S, Patterson M: Quantification of coral heat shock proteins from individual coral polyps. Mar Ecol Prog Ser. 2009, 376: 123-132.Google Scholar
  248. Chow AR, Ferrier-Pagès C, Khalouei S, Reynaud S, Brown IR: Increased light intensity induces heat shock protein Hsp60 in coral species. Cell Stress Chaperones. 2009, 14: 469-476. 10.1007/s12192-009-0100-6.PubMed CentralPubMedGoogle Scholar
  249. Venn AA, Quinn J, Jones R, Bodnar A: P-glycoprotein (multi-xenobiotic resistance) and heat shock protein gene expression in the coral Montastraea franksi in response to environmental toxicants. Aquat Toxicol. 2009, 93: 188-195. 10.1016/j.aquatox.2009.05.003.PubMedGoogle Scholar
  250. Nakamura M, Morita M, Kurihara H, Mitarai S: Expression of hsp 70, hsp 90 and hsf 1 in the reef coral Acropora digitifera under prospective acidified conditions over the next several decades. Biol Open. 2012, 1: 75-81. 10.1242/bio.2011036.PubMed CentralPubMedGoogle Scholar
  251. Hayes RL, King CM: Induction of 70-kD heat shock protein in scleractinian corals by elevated temperature: significance for coral bleaching. Mol Mar Biol Biotechnol. 1995, 4: 36-42.PubMedGoogle Scholar
  252. Fang L-S, Huang S-P, Lin K-L: High temperature induces the synthesis of heat-shock proteins and the elevation on intracellular calcium in the coral Acropora grandis. Coral Reefs. 1997, 16: 127-131. 10.1007/s003380050066.Google Scholar
  253. Carpenter LW, Patterson MR, Bromage ES: Water flow influences the spatiotemporal distribution of heat shock protein 70 within colonies of the scleractinian coral Montastrea annularis (Ellis and Solander, 1786) following heat stress: Implications for coral bleaching. J Exp Mar Biol Ecol. 2010, 387: 52-59. 10.1016/j.jembe.2010.02.019.Google Scholar
  254. Rosic NN, Pernice M, Dove S, Dunn S, Hoegh-Guldberg O: Gene expression profiles of cytosolic heat shock proteins Hsp70 and Hsp90 from symbiotic dinoflagellates in response to thermal stress: possible implications for coral bleaching. Cell Stress Chaperones. 2011, 16: 69-80. 10.1007/s12192-010-0222-x.PubMed CentralPubMedGoogle Scholar
  255. Roth MS, Goericke R, Deheyn DD: Cold induces acute stress but heat is ultimately more deleterious for the reef-building coral Acropora yongei. Sci Rep. 2012, 2: 240-PubMed CentralPubMedGoogle Scholar
  256. Aravind L, Anantharaman V, Koonin EV: Monophyly of class I aminoacyl tRNA synthase, USPA, ETFP, photolyase and the PP-ATPase nucleotide-binding domains: Implications for protein evolution in the RNA world. Proteins. 2002, 48: 1-14. 10.1002/prot.10064.PubMedGoogle Scholar
  257. Kültz D: Molecular and evolutionary basis of the cellular stress response. Annu Rev Physiol. 2005, 67: 225-257. 10.1146/annurev.physiol.67.040403.103635.PubMedGoogle Scholar
  258. Kvint K, Nachin L, Diez A, Nyström T: The bacterial universal stress protein: functions and regulation. Curr Opin Microbiol. 2003, 6: 140-145. 10.1016/S1369-5274(03)00025-0.PubMedGoogle Scholar
  259. Kerk D, Bulgrien J, Smith DW, Gribskov M: Arabidopsis proteins containing similarity to the universal stress protein domain of bacteria. Plant Physiol. 2003, 131: 1209-1219. 10.1104/pp.102.016006.PubMed CentralPubMedGoogle Scholar
  260. Forêt S, Seneca F, de Jong D, Bieller A, Hemmrich G, Augustin R, Hayward DC, Ball EE, Bosch TC, Agata K, Hassel M, Miller DJ: Phylogenomics reveals an anomalous distribution of USP genes in metazoans. Mol Biol Evol. 2011, 28: 153-161. 10.1093/molbev/msq183.PubMedGoogle Scholar
  261. DeSalvo MK, Voolstra CR, Sunagawa S, Schwartz JA, Stillman JH, Coffroth MA, Szmant AM, Medina M: Differential gene expression during thermal stress and bleaching in the Caribbean coral Montastraea faveolata. Mol Ecol. 2008, 17: 3952-3971. 10.1111/j.1365-294X.2008.03879.x.PubMedGoogle Scholar
  262. Voolstra CR, Schnetzer J, Penshkin L, Randall CJ, Szmant AM, Medina M: Effects of temperature on gene expression in embryos of the coral Montastraea faveolata. BMC Genomics. 2009, 10: 627-10.1186/1471-2164-10-627.PubMed CentralPubMedGoogle Scholar
  263. Sone H, Akanuma H, Fukuda T: Oxygenomics in environmental stress. Redox Rep. 2010, 15: 98-114. 10.1179/174329210X12650506623843.PubMedGoogle Scholar
  264. Pronk TE, Van Someren EP, Stierum RH, Ezendam J, Pennings JLA: Unraveling toxicological mechanisms and predicting toxicity classes with gene dysregulation networks. J Appl Toxicol. 2012, 10.1002/jat.2800. [Epub ahead of print]Google Scholar
  265. Hernández MP, Sullivan WP, Toft DO: The assembly and intermolecular properties of the hsp70-Hop-hsp90 molecular complex. J Biol Chem. 2002, 277: 38294-38304. 10.1074/jbc.M206566200.PubMedGoogle Scholar
  266. Song Y, Masison DC: Independent regulation of Hsp70 and Hsp90 chaperones by Hsp70/Hsp90-organising protein Sti (Hop1). J Biol Chem. 2005, 280: 34178-34185. 10.1074/jbc.M505420200.PubMed CentralPubMedGoogle Scholar
  267. Ellis RJ, van der Vies SM: Molecular chaperones. Annu Rev Biochem. 1991, 60: 321-347. 10.1146/annurev.bi.60.070191.001541.PubMedGoogle Scholar
  268. Takayama S, Reed JC, Homma S: Heat-shock proteins as regulators of apoptosis. Oncogene. 2003, 2003 (22): 9041-9047.Google Scholar
  269. Qiu XB, Shao YM, Miao S, Wang L: The diversity of the DnaJ/Hsp40 family, the crucial partners for Hsp70 chaperones. Cell Mol Life Sci. 2006, 63: 2560-2570. 10.1007/s00018-006-6192-6.PubMedGoogle Scholar
  270. Ke Q, Costa M: Hypoxia-inducible factor-1 (HIF-1). Mol Pharmacol. 2006, 70: 1469-1480. 10.1124/mol.106.027029.PubMedGoogle Scholar
  271. Rankin EB, Biju MP, Liu Q, Unger TL, Rha J, Johnson RS, Simon MC, Keith B, Haase VH: Hypoxia-inducible factor-2 (HIF-2) regulates erythropoietin in vivo. J Clin Invest. 2007, 117: 1068-1077. 10.1172/JCI30117.PubMed CentralPubMedGoogle Scholar
  272. Levy O, Kaniewska P, Alon S, Eisenberg E, Karako-Lampert S, Bay LK, Reef R, Rodriguez-Lanetty M, Miller DJ, Hoegh-Guldberg O: Complex diel cycles of gene expression in coral-algal symbiosis. Science. 2011, 331: 175-10.1126/science.1196419.PubMedGoogle Scholar
  273. Glickman MH, Ciechanover A: The ubiquitin-proteasome proteolytic pathway: destruction for the sake of construction. Physiol Rev. 2002, 82: 373-428.PubMedGoogle Scholar
  274. Parsell DA, Lindquist S: The function of heat-shock proteins in stress tolerance: degradation and reactivation of damaged proteins. Annu Rev Genet. 1993, 27: 437-496. 10.1146/annurev.ge.27.120193.002253.PubMedGoogle Scholar
  275. Imai J, Yashiroda H, Maruya M, Yahara I, Tanaka K: Proteasomes and molecular chaperones: cellular machinery responsible for folding and destruction of unfolded proteins. Cell Cycle. 2003, 2: 585-590.PubMedGoogle Scholar
  276. Rosenzweig R, Glickman MH: Forging a proteasome α-ring with dedicated proteasome chaperones. Nat Sruct Mol Biol. 2008, 15: 218-220. 10.1038/nsmb0308-218.Google Scholar
  277. Murata S, Yashiroda H, Tanaka K: Molecular mechanisms of proteasome assembly. Nat Rev Mol Cell Biol. 2009, 10: 104-115. 10.1038/nrm2630.PubMedGoogle Scholar
  278. Bügl H, Fauman EB, Staker BL, Zheng F, Kushner SR, Saper MA, Bardwell JC, Jakob U: RNA methylation under heat shock control. Mol Cell. 2000, 6: 349-360. 10.1016/S1097-2765(00)00035-6.PubMedGoogle Scholar
  279. Caldas T, Binet E, Bouloc P, Costa A, Desgres J, Richarme G: The FtsJ/RrmJ heat shock protein of Escherichia coli is a 23S ribosomal RNA methyl transferase. J Biol Chem. 2000, 275: 16414-16419. 10.1074/jbc.M001854200.PubMedGoogle Scholar
  280. Shimuta T, Nakano K, Yamaguchi Y, Ozaki S, Fujimitsu K, Matsunaga C, Noguchi K, Emoto A, Katayama T: Novel heat shock protein HspQ stimulates degradation of mutant DnaA protein in Escherichia coli. Genes Cells. 2004, 9: 1151-1166. 10.1111/j.1365-2443.2004.00800.x.PubMedGoogle Scholar
  281. Kitagawa M, Wada C, Yoshioka S, Yura T: Expression of ClpB, an analog of the ATP-dependent protease regulatory subunit in Escherichia coli, is controlled by a heat shock sigma factor (sigma 32). J Bacteriol. 1991, 173: 4247-4253.PubMed CentralPubMedGoogle Scholar
  282. Squires C, Squires CL: The Clp proteins: proteolysis or molecular chaperones?. J Bacteriol. 1992, 174: 1081-1085.PubMed CentralPubMedGoogle Scholar
  283. Lupas AN, Koretke KK: Bioinformatic anatysis of ClpS, a protein module involved in prokaryotic and eukaryotic protein degradation. J Struct Biol. 2003, 141: 77-83. 10.1016/S1047-8477(02)00582-8.PubMedGoogle Scholar
  284. Maillard RA, Chistol G, Sen M, Righini M, Tan J, Kaiser CM, Hodges C, Martin A, Bustamante C: ClpX(P) generates mechanical force to unfold and translocate its protein substrates. Cell. 2011, 145: 459-469. 10.1016/j.cell.2011.04.010.PubMed CentralPubMedGoogle Scholar
  285. Horwitz J: Alpha-crystallin can function as a molecular chaperone. Proc Natl Acad Sci U S A. 1992, 89: 10449-10453. 10.1073/pnas.89.21.10449.PubMed CentralPubMedGoogle Scholar
  286. Rao PV, Horwitz J, Zigler JS: α-Crystallin, a molecular chaperone, forms a stable complex with carbonic anhydrase upon heat denaturation. Biochem Biophys Res Commun. 1993, 190: 786-793. 10.1006/bbrc.1993.1118.PubMedGoogle Scholar
  287. Carricart-Ganivet JP, Cabanillas-Terán N, Cruz-Ortega I, Blanchon P: Sensitivity of calcification to thermal stress varies among genera of massive reef-building corals. PLoS One. 2012, 7: e32859-10.1371/journal.pone.0032859.PubMed CentralPubMedGoogle Scholar
  288. Taylor JL, Wieczorek A, Keyser AR, Grover A, Flinkstrom R, Karls RK, Bielefeldt-Ohmann H, Dobos KM, Izzo AA: HspX-mediated protection against tuberculosis depends on the chaperoning of a mycobacterial molecule. Immunol Cell Biol. 2012, 90: 945-954. 10.1038/icb.2012.34.PubMed CentralPubMedGoogle Scholar
  289. Santo Ede O, Alves N, Dias GM, Mazotto AM, Vermelho A, Vora GJ, Wilson B, Beltran VH, Bourne DG, Le Roux F, Thompson FL: Genomic and proteomic analysis of the pathogen Vibrio coralliilyticus reveals a diverse virulence repertoire. ISME J. 2011, 5: 1471-1483. 10.1038/ismej.2011.19.Google Scholar
  290. Kimes NE, Grim CJ, Johnson WR, Hasan NA, Tall BD, Kothary MH, Kiss H, Munk AC, Tapia R, Green L, Detter C, Bruce DC, Brettin TS, Colwell RR, Morris PJ: Temperature regulation of virulence factors in the pathogen Vibrio coralliilyticus. ISME J. 2012, 6: 835-846. 10.1038/ismej.2011.154.PubMed CentralPubMedGoogle Scholar
  291. Zhang T, Kraus WL: SIRT1-dependent regulation of chromatin and transcription: linking NAD+ metabolism and signaling to the control of cellular functions. Biochim Biophys Acta. 1804, 2010: 1666-1675.Google Scholar
  292. Liu TF, Yoza BK, El Gazzar M, Vachharajani VT, McCall CE: NAD+-dependent SIRT1 deacetylase participates in epigenetic reprogramming during endotoxin tolerance. J Biol Chem. 2011, 286: 9856-9864. 10.1074/jbc.M110.196790.PubMed CentralPubMedGoogle Scholar
  293. Katada S, Imhof A, Sassone-Corsi P: Connecting threads: epigenetics and metabolism. Cell. 2012, 148: 24-28. 10.1016/j.cell.2012.01.001.PubMedGoogle Scholar
  294. Lim JH, Lee YM, Chun YS, Chen J, Kim JE, Park JW: Sirtuin 1 modulates cellular responses to hypoxia by deacetylating hypoxia-inducible Factor 1α. Mol Cell. 2010, 38: 864-878. 10.1016/j.molcel.2010.05.023.PubMedGoogle Scholar
  295. Lee JH, Song MY, Song EK, Kim EK, Moon WS, Han MK, Park JW, Kwon KB, Park BH: Overexpression of SIRT1 protects pancreatic beta-cells against cytokine toxicity by suppressing the nuclear factor-kappaB signalling pathway. Diabetes. 2009, 58: 344-351.PubMed CentralPubMedGoogle Scholar
  296. Wang F, Nguyen M, Qin FX, Tong Q: SIRT2 deacetylates FOXO3a in response to oxidative stress and caloric restriction. Aging Cell. 2007, 6: 505-514. 10.1111/j.1474-9726.2007.00304.x.PubMedGoogle Scholar
  297. Shi T, Wang F, Stieren E, Tong Q: SIRT3, a mitochondrial sirtuin deacetylase, regulates mitochondrial function and thermogenesis in brown adipocytes. J Biol Chem. 2005, 280: 13560-13567. 10.1074/jbc.M414670200.PubMedGoogle Scholar
  298. Nasrin N, Wu X, Fortier E, Feng Y, Baré OC, Chen S, Ren X, Wu Z, Streeper RS, Bordone L: SIRT4 regulates fatty acid oxidation and mitochondrial gene expression in liver and muscle cells. J Biol Chem. 2010, 285: 31995-32002. 10.1074/jbc.M110.124164.PubMed CentralPubMedGoogle Scholar
  299. Du J, Zhou Y, Su X, Yu JJ, Khan S, Jiang H, Kim J, Woo J, Kim JH, Choi BH, He B, Chen W, Zhang S, Cerione RA, Auwerx J, Hao Q, Lin H: Sirt5 is a NAD-dependent protein lysine demalonylase and desuccinylase. Science. 2011, 334: 806-809. 10.1126/science.1207861.PubMed CentralPubMedGoogle Scholar
  300. Mostoslavsky R, Chua KF, Lombard DB, Pang WW, Fisher MR, Gellon L, Liu P, Mostoslavsky G, Franco S, Murphy MM, Mills KD, Patel P, Hsu JT, Hong AL, Ford E, Cheng HL, Kennedy C, Nunez N, Bronson R, Frendewey D, Auerbach W, Valenzuela D, Karow M, Hottiger MO, Hursting S, Barrett JC, Guarente L, Mulligan R, Demple B, Yancopoulos GD: Genomic instability and aging-like phenotype in the absence of mammalian SIRT6. Cell. 2006, 124: 315-329. 10.1016/j.cell.2005.11.044.PubMedGoogle Scholar
  301. Luo J, Nikolaev AY, Imai S, Chen D, Su F, Shiloh A, Guarente L, Gu W: Negative control of p53 by Sir2alpha promotes cell survival under stress. Cell. 2001, 107: 137-148. 10.1016/S0092-8674(01)00524-4.PubMedGoogle Scholar
  302. Vakhrusheva O, Smolka C, Gajawada P, Kostin S, Boettger T, Kubin T, Braun T, Bober E: Sirt7 increases stress resistance of cardiomyocytes and prevent apoptosis and inflammatory cardiomyopathy in mice. Circ Res. 2008, 102: 703-710. 10.1161/CIRCRESAHA.107.164558.PubMedGoogle Scholar
  303. O’Halloran TV, Culotta VC: Metallochaperones, an intracellular shuttle service for metal ions. J Biol Chem. 2000, 275: 25057-25060. 10.1074/jbc.R000006200.PubMedGoogle Scholar
  304. Lin SJ, Culotta VC: The ATX1 gene of Saccharomyces cerevisiae encodes a small metal homeostasis factor that protects cells against reactive oxygen toxicity. Proc Natl Acad Sci U S A. 1995, 92: 3784-3788. 10.1073/pnas.92.9.3784.PubMed CentralPubMedGoogle Scholar
  305. Culotta VC, Klomp LWJ, Strain J, Casareno RLB, Krems B, Gitlin JD: The copper chaperone for superoxide dismutase. J Biol Chem. 1997, 272: 23469-23472. 10.1074/jbc.272.38.23469.PubMedGoogle Scholar
  306. Wong PC, Waggoner D, Subramaniam JR, Tessarollo L, Bartnikas TB, Culotta VC, Price DL, Rothstein J, Gitlin JD: Copper chaperone for superoxide dismutase is essential to activate mammalian Cu/Zn superoxide dismutase. Proc Natl Acad Sci USA. 2000, 97: 2886-2891. 10.1073/pnas.040461197.PubMed CentralPubMedGoogle Scholar
  307. Shick JM, Lesser MP, Dunlap WC, Stochaj WR, Chalker BE, Wu Won J: Depth-dependent responses to solar ultraviolet radiation and oxidative stress in the zooxanthellate coral Acropora microphthalma. Mar Biol. 1995, 122: 41-51. 10.1007/BF00349276.Google Scholar
  308. Lesser MP, Stochaj WR, Tapley DW, Shick JM: Bleaching in coral-reef anthozoans: effects of irradiance, ultraviolet radiation, and temperature on the activities of protective enzymes against active oxygen. Coral Reefs. 1990, 8: 225-232. 10.1007/BF00265015.Google Scholar
  309. Crowe JH, Carpenter JF, Crowe LM: The role of vitrification in anhydrobiosis. Annu Rev Physiol. 1998, 60: 73-103. 10.1146/annurev.physiol.60.1.73.PubMedGoogle Scholar
  310. Sola-Penna M, Meyer-Fernandes JR: Stabilization against thermal inactivation promoted by sugars on enzyme structure and function: Why is trehalose more effective than other sugars?. Arch Biochem Biophys. 1998, 360: 10-14. 10.1006/abbi.1998.0906.PubMedGoogle Scholar
  311. Veron JEN: Corals of Australia and the Indo-Pacific. 1993, Singapore: University of Hawaii Press, 644-Google Scholar
  312. Shick JM: Solar UV and oxidative stress in algal-animal symbioses. Frontiers of Photobiology. Edited by: Shima A, Ichihashi M, Fujiwara Y, Takebe H. 1993, Amsterdam: Elsevier Science Publishers, 561-564.Google Scholar
  313. Turrens JF: Mitochondrial formation of reactive oxygen species. J Physiol. 2003, 552: 335-344. 10.1113/jphysiol.2003.049478.PubMed CentralPubMedGoogle Scholar
  314. Lesser MP: Oxidative stress causes coral bleaching during exposure to elevated temperatures. Coral Reefs. 1997, 16: 187-192. 10.1007/s003380050073.Google Scholar
  315. Brown BE, Downs CA, Dunn RP, Gibb SW: Exploring the basis of thermotolerance in the reef coral Goniastrea aspera. Mar Ecol Prog Ser. 2002, 242: 119-120.Google Scholar
  316. Halliwell B: Reactive species and antioxidants. Redox biology is a fundamental theme of aerobic life. Plant Physiol. 2006, 141: 312-322. 10.1104/pp.106.077073.PubMed CentralPubMedGoogle Scholar
  317. Liñán-Cabello MA, Lesser MP, Flores-Ramírez LA, Zenteno-Savín T, Reyes-Bonilla H: Oxidative stress in coral-photobiont communities. Oxidative Stress in Aquatic Ecosystems. Edited by: Abele D, Vázquez-Medina JP, Zenteno-Savín T. 2011, Chichester: Wiley-Blackwell, 127-138.Google Scholar
  318. Jönsson TJ, Lowther WT: The peroxiredoxin repair proteins. Subcell Biochem. 2007, 44: 115-141. 10.1007/978-1-4020-6051-9_6.PubMed CentralPubMedGoogle Scholar
  319. Cheng Z, Zhang J, Ballou DP, Williams CH: Reactivity of thioredoxin as a protein thiol-disulfide oxidoreductase. Chem Rev. 2011, 111: 5768-5783. 10.1021/cr100006x.PubMed CentralPubMedGoogle Scholar
  320. Arnér ESJ, Holmgren A: Physiological function of thioredoxin and thioredoxin reductase. Eur J Biochem. 2000, 267: 6102-6109. 10.1046/j.1432-1327.2000.01701.x.PubMedGoogle Scholar
  321. Ainsworth TD, Hoegh-Guldberg O: Cellular processes of bleaching in the Mediterranean coral Oculina patagonica. Coral Reefs. 2008, 27: 593-597. 10.1007/s00338-008-0355-x.Google Scholar
  322. Kenkel CD, Aglyamova G, Alamaru A, Bhagooli R, Capper R, Cunning R, De Villers A, Haslun JA, Hédouin L, Keshavmurthy S, Kuehl KA, Mahmoud H, McGinty ES, Montoya-Maya PH, Palmer CV, Pantile R, Sánchez JA, Schils T, Silverstein RN, Squiers LB, Tang PC, Goulet TL, Matz MV: Development of gene expression markers of acute heat-light stress in reef-building corals of the genus Porites<