Comparative transcriptome analysis of obligately asexual and cyclically sexual rotifers reveals genes with putative functions in sexual reproduction, dormancy, and asexual egg production
© Hanson et al.; licensee BioMed Central Ltd. 2013
Received: 16 March 2013
Accepted: 31 May 2013
Published: 19 June 2013
Sexual reproduction is a widely studied biological process because it is critically important to the genetics, evolution, and ecology of eukaryotes. Despite decades of study on this topic, no comprehensive explanation has been accepted that explains the evolutionary forces underlying its prevalence and persistence in nature. Monogonont rotifers offer a useful system for experimental studies relating to the evolution of sexual reproduction due to their rapid reproductive rate and close relationship to the putatively ancient asexual bdelloid rotifers. However, little is known about the molecular underpinnings of sex in any rotifer species.
We generated mRNA-seq libraries for obligate parthenogenetic (OP) and cyclical parthenogenetic (CP) strains of the monogonont rotifer, Brachionus calyciflorus, to identify genes specific to both modes of reproduction. Our differential expression analysis identified receptors with putative roles in signaling pathways responsible for the transition from asexual to sexual reproduction. Differential expression of a specific copy of the duplicated cell cycle regulatory gene CDC20 and specific copies of histone H2A suggest that such duplications may underlie the phenotypic plasticity required for reproductive mode switch in monogononts. We further identified differential expression of genes involved in the formation of resting eggs, a process linked exclusively to sex in this species. Finally, we identified transcripts from the bdelloid rotifer Adineta ricciae that have significant sequence similarity to genes with higher expression in CP strains of B. calyciflorus.
Our analysis of global gene expression differences between facultatively sexual and exclusively asexual populations of B. calyciflorus provides insights into the molecular nature of sexual reproduction in rotifers. Furthermore, our results offer insight into the evolution of obligate asexuality in bdelloid rotifers and provide indicators important for the use of monogononts as a model system for investigating the evolution of sexual reproduction.
KeywordsEvolution of sexual reproduction Differential expression analysis Gene ontology analysis Meiosis Gametogenesis Resting eggs Mixis induction
Understanding the origins and persistence of sexual reproduction have long been recognized as challenges in evolutionary biology [1–3]. Although sexual reproduction is pervasive in eukaryotes —suggesting it provides critical benefits to an organism—it is also frequently and sporadically lost [e.g. [5, 6]]—suggesting it can be dispensable. In fact, loss of sexual reproduction appears to provide several immediate benefits to a lineage, including a two-fold increase in the production of females, the preservation of beneficial combinations of alleles , and the loss of risky and energetically expensive processes and behaviors associated with mating. Dubbed “the queen of problems in evolutionary biology” , a comprehensive explanation for the prevalence of sexual reproduction in nature has remained elusive [7–9].
Rotifers—a diverse phylum of aquatic invertebrates—are an interesting example of the diversity in reproductive modes found in Metazoa: they include obligate sexual (seisonids and acanthocephalans), facultative sexual (monogononts), and obligate asexual (bdelloids) lineages. Indeed, bdelloid rotifers have persisted and diversified into hundreds of species over tens of millions of years without the canonical forms of sexual reproduction [10, 11]. However, it has been difficult to establish clear expectations for any potential cryptic forms of sex in bdelloids without first understanding the molecular nature of sex in relatives such as the facultatively sexual monogononts, from which bdelloids diverged over 40 million years ago .
Intriguingly, the sensitivity of monogononts to the chemical signal and transition to sexual reproduction is highly polymorphic and can be lost entirely. Sex loss has been observed in several monogonont species subjected to long-term laboratory culture [16, 17] and is mediated by a single locus in the monogonont Brachionus calyciflorus. These B. calyciflorus strains reproduce exclusively by obligate parthenogenesis (OP) and no longer respond to the chemical signal that induces mixis in cyclical parthenogenetic (CP) strains.
OP and CP strains of B. calyciflorus provide a powerful means through which the evolution of sexual reproduction can be directly tested by allowing direct competition between sexually and asexually reproducing populations . However, little is known about the molecular underpinnings of sexual reproduction in monogononts generally, or the underlying mechanism for the loss of sex in B. calyciflorus specifically. Furthermore, several key processes are linked exclusively with sexual reproduction in monogononts and are therefore lost in the OP strains (Figure 1). First, the signaling pathway resulting in the induction of sexual reproduction has apparently been disrupted in the OP strains. Although progesterone signaling contributes to sex induction in the monogonont Brachionus manjavacas[20, 21], B. calyciflorus does not appear to respond to the same cues . Comparisons between OP and CP strains may therefore help elucidate the pathway in B. calyciflorus, allowing an examination of the divergence of this signaling pathway in monogononts. Second, production of haploid eggs through a reductive division (meiosis) and the presence of males are unique to sexual populations. Identification of genes specific to meiosis or the production of males through OP and CP expression patterns will offer insights into these processes in monogononts, which will provide an important framework for the study of the asexual bdelloids, as purifying selection should no longer be acting on these loci once sex has been lost. Finally, resting eggs—which are capable of surviving harsh environmental conditions and are used for overwintering—are produced only by fertilization. Previous studies have established gene expression associated specifically with resting eggs in Brachionus plicatilis[23, 24]; however, genes involved in the early stages of oogenesis and egg development have not been examined quantitatively.
In order to elucidate the gene expression changes associated with sexual reproduction in monogononts, we generated transcriptome data from OP and CP strains of B. calyciflorus. We observed increased expression of genes associated with steroid signaling, meiosis, gametogenesis and dormancy in the CP strains, implicating specific pathways in processes related to sex in B. calyciflorus. Further, several genes with increased expression in the OP strains were identified, suggesting pathways specific to asexual egg production. Finally, we analyzed previously published bdelloid gene expression data in the context of the observed monogonont expression patterns in order to identify genes present in the bdelloids that have putative roles in monogonont reproduction. Our findings provide molecular insights into sexual reproduction in monogononts and apply an initial comparative approach to understanding the nature of reproduction in bdelloids.
RNA extraction and library preparation
OP and CP clones of Brachionus calyciflorus homozygous for the op and wild-type alleles, respectively, were generated from the same heterozygous mother by selfing . Six B. calyciflorus females were added to 600 mL cultures of 5 × 105 cells/mL Chlamydomonas reinhardtii in COMBO medium . The cultures grew at 25 C and with 24 hour light exposure for ~6 days. Density of females was determined every day by removal of ~10 mL of culture and counting individual females under a microscope at low magnification. When cultures reached a density of ~25-40 females/mL, they were harvested for RNA. To confirm the presence of sexually-reproducing females in the CP cultures, a sample of females was used to determine rate of mixis induction as described previously , and the absence of males and resting eggs was confirmed for the OP cultures. The rotifer cultures were filtered through 20 μm Millipore nylon filters to remove algal cells, the rotifers were washed in COMBO medium, and rinsed into 15 mL conical tubes. Total RNA was isolated using Trizol reagent (Ambion, Life Technologies, Grand Island, NY) according to manufacturer’s instructions. RNA quality was determined by Experion Automated Electrophoresis (Bio-Rad Laboratories, Hercules, CA) according to manufacturer’s instructions before construction of mRNA-seq libraries using the mRNA-seq Sample Prep kit (Illumina, Inc., San Diego, CA). Seventy-six base pair single end reads were sequenced from each library on an Illumina Genome Analyzer IIx platform at the Iowa State University DNA Facility (http://www.dna.iastate.edu/nextgen.html).
Illumina sequence analysis
Analysis of Illumina sequence reads was performed in part using the Galaxy server (http://galaxyproject.org/) [27–29]. Adapter sequences were trimmed from the ends of the reads, and a blastn search revealed reads that matched C. reinhardtii transcripts . Reads with an alignment length ≥ 20 base pairs matching an algal sequence were removed from the libraries. Transcriptome assembly was performed using the Tuxedo pipeline, where reads were mapped to a partial assembly of the B. calyciflorus genome via Tophat , assembled into transcripts using Cufflinks (assembly for individual samples with 0.1 minimum isoform fraction and 0.1 pre-mRNA fraction) and Cuffmerge (combined assembly from all samples). Differential expression was determined between OP and CP samples with Cuffdiff using a minimum alignment count of 500 [33, 34]. Differential expression between OP and CP samples was also determined with edgeR using a table of counts constructed with the fragments per kilobase locus length per million reads mapped (FPKM) values determined by Cufflinks . Gene ontology mapping and enrichment analyses were performed with BLAST2GO (http://www.blast2go.com/b2ghome). The transcriptome assembly projects have been deposited at GenBank under the accessions GACQ00000000 and GACL00000000. The versions described in this paper are the first versions, GACQ01000000 and GACL01000000. Assembled transcripts shorter than 200 base pairs are given in Additional files 1, 2, 3 and 4.
OP and CP RNA samples used for Illumina library construction were treated with DNase I prior to first-strand cDNA synthesis using Superscript II reverse transcriptase (Invitrogen, Life Technologies, Grand Island, NY). Quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) was performed with Platinum SYBR Green quantitative PCR (qPCR) Supermix (Invitrogen, Life Technologies, Grand Island, NY) and a 1:25 dilution of the cDNA. Primers designed for amplification of thirteen genes from cDNA in the qPCR reaction (200 nM final concentration) are listed in Additional file 5. Relative quantification of gene expression was determined for each gene, with actin transcript amplification used for normalization.
For analysis of histone H2A, amino acid sequences of metazoan and fungal homologs were obtained from the National Center for Biotechnology Information (NCBI) database by keyword search and aligned with B. calyciflorus sequences by ClustalX version 2.1 . Alignments were manually curated in MacClade (http://macclade.org/index.html) and phylogenetic analysis was performed using the Bioportal parallel computational resource . Maximum likelihood analysis was performed using PhyML  with Whelan and Goldman (WAG) substitution model, best tree topology search, and one thousand bootstrap replicates.
Bdelloid sequence analysis
Transcript sequences for Adineta ricciae were obtained from GenBank (Accession numbers: HE687510 to HE716431). A. ricciae transcripts were used as queries against the assembled B. calyciflorus transcriptome using tblastx  with a Bit Score cutoff of 50. Gene ontology term enrichment was performed using BLAST2GO .
Results and discussion
Assembled genes with highest expression values in OP and CP libraries
Sequence (top hit)
MRP motif repeat 3 protein precursor
MRP motif repeat 4 protein precursor
60S ribosomal protein L4
Pediculus humanus corporis
MRP motif repeat 3 protein precursor
MRP motif repeat 3 protein precursor
MRP repeat motif 5 protein precursor
tropomyosin isoform 5
ribosomal protein L4
60S ribosomal protein L23a
ubiquitin/ribosomal S27A fusion protein 3
60S ribosomal protein L38
ribosomal protein rps23
60S ribosomal protein L35
60S ribosomal protein L15
60S ribosomal L7a-like protein
60S acidic ribosomal protein P1
ribosomal protein S18
MRP motif repeat 3 protein precursor
MRP motif repeat 4 protein precursor
MRP motif repeat 3 protein precursor
3.E + 00
MRP motif repeat 3 protein precursor
MRP repeat motif 5 protein precursor
MRP motif repeat 3 protein precursor
60S ribosomal protein L35
Gram-negative binding protein 7, partial
60S ribosomal protein L38
tropomyosin isoform 5
ubiquitin/ribosomal S27A fusion protein
ribosomal protein S18
60S ribosomal protein L12
ribosomal protein rps23
voltage-dependent calcium channel beta subunit
Notably, we found that a large number of the genes assembled by Cuffmerge (45%) did not have identifiable homologs by BLAST search (e value < 1). In many cases this is likely due to the short length of the assembled sequences, as only 24% of the genes >400 base pairs in length did not have significant BLAST hits. This proportion of unique sequences is consistent with that observed for expression data from the monogonont B. plicatilis[42, 43]. Furthermore, a recently published transcriptome for the bdelloid rotifer Adineta ricciae found that 53% of transcripts did not have a significant BLAST hit , which may indicate that the lack of identifiable homologs may be due to limited taxonomic sampling in the NCBI database.
Comparison of FPKM distributions between OP and CP libraries
MI-Chromosome Struc.& Int.
We next examined the expression of genes involved in several aspects of monogonont reproductive biology in our expression libraries for the OP and CP samples. These include genes with putative roles in mixis induction, meiosis and gametogenesis, dormancy, and asexual egg production.
The trigger for the transition from asexual to sexual reproduction in many species of monogononts is a secreted protein signal that accumulates in their environment . Although little is known about the signal or the pathway(s) it activates to induce the transition, several lines of evidence indicate that it is mediated by steroid hormone signaling. First, isolation of the protein signal and sequencing of its N-terminus revealed that its sequence in the species B. plicatilis is similar to the mammalian steroidogenesis-inducing protein . Second, exposure to hormones, including estradiol-17 and progesterone, resulted in increased production of sexual females in populations of B. plicatilis[21, 45]. Third, RNAi knockdown of the progesterone receptor in B. manjavacas reduced production of sexual females . Intriguingly, there appears to be species-specificity in the response to steroid hormones, as sexual reproduction is not induced by progesterone treatment alone in B. calyciflorus cultures [21, 22].
Steroid hormone signaling is mediated by members of the nuclear receptor superfamily, a large family of proteins that are well-conserved across Metazoa . Nuclear receptors have critical functions in developmental processes, including the induction of meiosis during oogenesis in mammals [47, 48]. Members of the nuclear receptor superfamily are transcription factors whose activity is modulated through direct interactions with steroid ligands. Nuclear receptors frequently act as homodimers or heterodimers when modulating transcription of target sequences via histone acetylation or deacetylation, including auto-regulating and cross-regulating the transcription of other nuclear receptors [48, 49].
We identified 73 genes in our transcriptome analysis with significant sequence identity to the DNA-binding domains of the nuclear receptor superfamily, of which five have significantly higher expression levels in CP populations (Additional file 12). Although we were unable to obtain phylogenetic resolution to confirm the orthology of the receptors (data not shown), these data are consistent with the role of steroid signaling in mixis induction.
We also identified differentially expressed genes with functions and GO term designations associated with steroid metabolic and signaling pathways (Additional file 9). CBP/p300 is a transcription factor with histone acetyltransferase activity that acts as a co-regulator of transcription with nuclear receptor superfamily members . Protein Kinase A modulates nuclear receptor activity via phosphorylation and has roles during gametogenesis . Bcl-X mediates apoptosis and is regulated through steroid hormone signaling pathways . These genes provide additional candidates for roles in the transition from asexual to sexual reproduction in B. calyciflorus.
Meiosis, recombination, and gametogenesis
Genes with known roles in meiosis based on evidence in well-studied model systems (e.g. yeast, mouse, and nematodes) were previously annotated in monogononts . The expression levels of these genes are shown in Additional file 11. The distribution of expression values for these genes as a group is significantly different in CP than in OP strains, with a greater median expression value in the CP strain (Table 2, Additional file 10). This difference in expression value distribution is also found in more specific categories of gene function within the meiotic gene inventory (Table 2, Additional files 10 and 11).
Histone H2A genes identified in OP and CP libraries
# A. ricciae transcripts
A. ricciae accession
Xenopus (Silurana) tropicalis
Mustela putorius furo
Progression through mitosis and meiosis is mediated by the anaphase-promoting complex/cyclosome (APC/C), which acts as a ubiquitin ligase targeting proteins for proteasomal degradation. APC/C activity is mediated by CDC20, which has paralogs (Fizzy and Fizzy-Related) functioning in both mitosis and meiosis . In addition, an arthropod-specific paralog, Cortex, functions exclusively in meiosis .
We previously identified four copies of CDC20 in the genome of B. calyciflorus with phylogenetic analysis placing three copies in the Fizzy clade (CDC20A, B, and C) and one copy in the Fizzy-related clade (CDC20D) . One Fizzy copy (CDC20A) is differentially expressed, with ~18-fold higher expression in the CP populations (Additional file 11). Its higher expression in CP populations suggests it may function in a process specific to sexual reproduction in B. calyciflorus, such as resting egg production, spermatogenesis, or meiosis, or is involved in a developmental process specific to the sexual cycle.
Histone H2A variants
Histone octamers comprised of the histones H2A, H2B, H3, and H4 are the primary protein component of chromatin. Histones are globular proteins that have a highly conserved core sequence and a C-terminal tail sequence that is post-translationally modified to alter chromatin structure and transcriptional activity. Canonical histones may be replaced by variants in a spatial or temporal manner in response to internal or external cues . In particular, variants of histone H2A are associated with DNA repair and gametogenesis. Histone H2A.X localizes to DNA double-strand breaks, where it recruits components of repair machinery. This occurs when breaks are induced through exposure to endogenous or exogenous agents, as well as in meiosis, when double-strand breaks are created by the topoisomerase Spo11 (Fernandez, 2004). Another histone variant specific to mammals, H2A.bbd, localizes to transcriptionally active sites within mouse testes, where genes with meiotic expression patterns are located (Soboleva, 2011).
Bdelloid rotifers, which apparently reproduce solely through ameiotic parthenogenesis, possess three histone H2A variants, some with multiple identified alleles. These H2A sequences have long C-terminal tail sequences that do not correspond to the tails of canonical or known variants of H2A, including the well-conserved SQE/D sequence in the tail of H2A.X . Although functionally uncharacterized, because of the importance of H2A variants to double-strand break repair in other systems, it is hypothesized that these novel variants may have evolved to aid the bdelloids in DNA repair following bouts of desiccation, which the rotifers can survive at any life stage. This same report identified three apparently canonical histone H2A sequences in the monogonont rotifer B. plicatilis.
Our transcriptome analysis indicated that three of the histone H2A sequences are differentially expressed between OP and CP populations, with higher expression in the CP strains (Table 3). Differential expression of these sequences suggests a role for these H2A genes in processes related to sexual reproduction in monogononts, perhaps similar to the functions performed by H2A.X or H2A.bbd in other systems.
In addition to these novel H2A sequences, we identified two histone interacting genes with significantly higher expression in the CP populations (Additional file 9). Tonsoku interacts with both histone H2A and H2B, and has a role in resolution of stalled replication forks and double strand breaks . Pax-Interacting Protein 1 recruits the H3K4 methyltransferase complex to trimethylate specific sites and activate transcription , and is required for both embryonic and somatic tissue differentiation . H3K4 trimethylation is also required during meiosis for pairing of homologs and progression through pachytene .
We identified 39 genes in our study with significant sequence identity to genes with roles in spermatogenesis or sperm morphology (Additional file 13). Several structural components of sperm flagellar tails were identified in the assembled genes from B. calyciflorus: Outer Dense Fiber of Sperm Tails 2 [64, 65], Sperm-Associated Antigen 6 [66, 67], Sperm-Associated Antigen 16 , Sperm-Associated Antigen 17 , Sperm Flagellar Protein 1  and Sperm Flagellar Protein 2 . We also identified genes with regulatory roles for transcription or cell cycle progression in the testes: AMY-1-associated protein expressed in testis-1 (AAT-1) , Nuclear autoantigenic sperm protein , Round Spermatid Basic Protein 1 , Sperm-Associated Antigen 1 [74, 75], Sperm-Associated Antigen 8 , Testis Expressed gene 14 [77, 78], Testis expressed gene 15, and Testis specific ser/thr protein kinase 2 . Finally, we found several genes whose gene expression and/or localization patterns are specific to testes or sperm in various developmental stages in Metazoa, including Granulin , PolyA Polymerase Beta , and Spermatogenesis Associated Genes [82–88].
Intriguingly, these genes were expressed in both OP and CP strains and while the distribution of expression of the genes as a group was significantly different between OP and CP with a higher median expression value in CP samples (Table 2, Additional file 10), none of the genes were significantly differentially expressed individually (Additional file 13). The expression of these genes in the presence and absence of males is consistent with a previous study in B. plicatilis showing that genes with putatively male-specific functions were also expressed in females . Indeed, several of these genes have been implicated in processes outside of spermatogenesis, such as roles in cilia development and in more generalized differentiation processes including oncogenesis (e.g. [70, 78]).
Finally, several genes with other roles in gametogenesis have significantly higher expression in the CP populations (Additional file 9). Brain Tumor Protein is a homolog of Mei-P26, which maintains germline stem cells and promotes germline differentiation during oogenesis in Drosophila melanogaster through interactions with the miRNA pathway protein Argonaute 1 [89, 90]. HSP70, in addition to an implicated role in monogonont dormancy , also plays a role in chromatin restructuring during spermatogenesis [91, 92]. Furthermore, Sacsin integrates the HSP70 pathway with proteasomal degradation during gametogenesis  and also has higher expression in CP strains.
Gene ontology enrichment analysis for B. calyciflorus genes with higher expression in CP strains
structural constituent of cuticle
chitin metabolic process
aminoglycan metabolic process
polysaccharide metabolic process
serine-type endopeptidase activity
transition metal ion binding
Further, enrichment for other terms likely relating to resting egg components/structure was identified, such as chitin binding (GO:0008061) and chitin metabolic process (GO:0006030). Asexually produced eggs are encased in one chitin-containing shell, while resting eggs have two shells, with the inner shell of the resting eggs—also composed of chitin—thicker than the asexual egg shell , providing a likely explanation for the enrichment of these terms in the differentially expressed genes. Interestingly, these GO terms were not identified as enriched in the previous resting egg transcriptome analyses , which focused on transcripts exclusively in the eggs. This discrepancy may indicate the maternal production of these transcripts during early stages of egg development, prior to egg extrusion.
In addition to significantly different distribution of expression values for dormancy genes as a group in CP samples (Table 2, Additional file 10), we identified several individual genes with significantly higher expression in the CP strains that were previously associated with dormancy in the monogonont B. plicatilis (Additional file 11). Aquaporin 3 is involved in water and small molecule transport . Genes involved in lipid metabolism, including cathepsin L, fatty acid binding genes, and lipases may be involved in yolk processing that differs between resting and amictic eggs in B. calyciflorus[95, 96]. Heat shock proteins are molecular chaperones that prevent the aggregation of misfolded proteins and have been implicated in dormancy in rotifers and other systems . Trehalose is a likely energy source for dormant organisms, suggesting a role for trehalose metabolic enzymes in monogononts [98, 99]. Late embryogenesis-abundant genes have been associated with embryonic development and dormancy in several organisms, including rotifers [100, 101].
The number of genes with significantly higher expression in the OP strain was much lower than that for the CP strains (Additional file 9). This is expected given that asexually reproducing females cannot be morphologically distinguished from sexually reproducing females in the CP strains. These females were therefore also included in CP samples, which likely decreased the fold change in expression of any gene specific to asexual females and/or their eggs. However, several interesting genes were found to have significantly higher expression in the OP strains.
Several genes with significant sequence similarity to genes associated with transposable elements show significantly higher expression in OP strains. Transposable element replication is highly regulated through multiple pathways in mammals and Drosophila, including transcriptional repression through epigenetic modifications, RNA editing, transcript degradation by RNA interference, and DNA repair pathways . Further, specific developmental stages during gametogenesis and embryonic development in mammals allow windows of opportunity for transposable elements to replicate and integrate into the host genome [102, 103]. Increased expression of transposons in OP strains suggests developmental differences that result in reduced suppression of these elements in B. calyciflorus.
Septins are cytoskeletal components that are flexibly combinatorial in the formation of filaments and rings. In ring form, septins act as scaffolds that aid in cell signaling, diffusion barriers between cells during cytokinesis, and components of cilia and flagella . The increased expression of a gene with significant sequence similarity to Septin 10 in OP strains could indicate a role for cytoskeletal restructuring during asexual female development or oogenesis. This is supported by previous transcriptome data from asexual eggs in B. plicatilis, in which several other cytoskeletal components had increased expression .
Two genes with sequence similarity to pancreatic triacylglycerol lipases were expressed at significantly higher levels in OP strains, while one pancreatic triacylglycerol lipase had significantly higher expression in the CP strains (Additional file 11). Combined with the significantly increased expression of other metabolic genes in CP described above this further suggests a differential role for lipid metabolism in the development of both resting eggs and asexually-produced eggs, as has been suggested previously by gene expression studies specifically examining resting and amictic eggs .
Gene expression in bdelloid rotifers
Using a published gene expression dataset containing 28,922 assembled transcripts from the bdelloid rotifer A. ricciae, we identified transcripts with significant sequence similarity to the B. calyciflorus sequences. Using tblastx with a bit score cutoff of 50, a total of 12,512 (~31%) of the B. calyciflorus sequences had at least one significant A. ricciae transcript hit (Additional file 14).
We further examined how the proportion of B. calyciflorus sequences with significant sequence similarity to A. ricciae transcripts changed with the difference in expression observed between CP and OP strains of B. calyciflorus. If genes with increased expression in the CP strain represent those that function in a process specific to sexual reproduction, it is likely that no identifiable homolog is present in bdelloids, as selection would no longer maintain the gene for that function. In contrast, genes that show no difference in expression likely have constitutive functions and would be more likely to be conserved in bdelloids. Finally, the presence of genes with higher expression in the OP strain in the A. ricciae transcripts would be more likely given that they may function in ameiotic egg production.
Gene ontology enrichment analysis for genes with significant similarity to bdelloid transcript
oxidoreductase activity, acting on peroxide as acceptor
ferric iron binding
response to stress
response to oxidative stress
A decrease in the proportion of B. calyciflorus genes with significant sequence similarity to A. ricciae transcripts was also observed for genes with increased expression in the OP strain (Figure 4, Additional file 6). While this could suggest a larger trend for more highly regulated genes in monogononts having faster rates of evolution or species-specificity, it may also suggest that the genes associated with asexual reproduction have roles in the suppression of sexual processes; these genes would be unnecessary in the bdelloids, as they are not known to undergo the reproductive switch that occurs in monogononts.
Finally, we examined the identified genes in B. calyciflorus with known roles in mixis induction, meiosis, and gametogenesis that had significant sequence similarity to A. ricciae transcripts. Of the seventy-three nuclear receptor superfamily genes identified in B. calyciflorus, twenty-eight had at least one significant A. ricciae transcript hit (Additional file 12). Three of the receptors with A. ricciae hits are differentially expressed between CP and OP strains of B. calyciflorus. However, as mentioned above, the highly conserved DNA binding domain of nuclear receptors makes it difficult to differentiate members within the superfamily, so additional studies will be required to determine the roles of these genes in both monogononts and bdelloids.
For genes with potential roles in meiosis, seven of the histone H2A genes identified in B. calyciflorus have significant hits in the A. ricciae dataset (Table 3). No significant hits were found for H2A.Z, consistent with a previous study suggesting this variant is absent in bdelloids . The three H2A genes that have significant differential expression in B. calyciflorus have at least one A. ricciae transcript with significant sequence similarity. Of the 122 monogonont genes with known roles in meiosis in model systems, 73 have A. ricciae transcripts, including 31 genes with roles in chromosome structure and interaction, 16 genes involved in recombination, and 26 genes involved in cell cycle progression (Additional file 11). CDC20A, which is differentially expressed in B. calyciflorus CP and OP strains, does not have a significant A. ricciae transcript hit, although A. ricciae transcripts were identified for two other copies, CDC20C and CDC20D. For “meiosis-specific” genes, no A. ricciae transcripts were identified with the exception of the MutS homologs Msh4 and Msh5. These genes act as a heterodimer in the resolution of the double-Holliday junction during meiotic recombination , and have also previously been identified in bdelloids via degenerate PCR (Schurko et al., unpublished).
Of the 39 genes with putative functions in male gametogenesis, twenty-two have significant hits in the A. ricciae transcripts (Additional file 13). The presence and expression of these genes in bdelloids combined with the lack of significant differential expression between CP and OP strains of B. calyciflorus suggests that roles for these genes are not exclusive to spermatogenesis in rotifers.
We identified a large set of genes with significantly higher expression in either cyclically parthenogenetic (CP) or obligately parthenogenetic (OP) populations of the monogonont rotifer, Brachionus calyciflorus. Included in this set of genes are those with potential roles in processes specific to sexual reproduction in monogononts such as mixis induction, meiosis and gametogenesis, and dormancy. We further identified several genes with increased expression in OP populations that suggest developmental and metabolic differences during asexual egg production. The samples used in this study included entire populations of monogonont rotifers, and future studies focusing on specific life cycle stages will clarify and enhance the data presented here.
Notably, none of the identified “meiosis-specific” genes were differentially expressed between OP and CP populations (Additional file 11). Although the expression levels were low for some of these genes—potentially due to restricted temporal and spatial expression to meiosis, which is likely only occurring in a subset of embryos in the population —this is consistent with observations made in previous studies suggesting that these genes are not always robust markers for sex, as their transcription is not specific to meiosis in all organisms. For example, in the cyclical parthenogen Daphnia pulex, these genes are expressed in the ovaries of both sexual and asexual females . Furthermore, oogenesis in asexual D. pulex females is an abortive meiotic division, suggesting that some of these genes may be required for egg production regardless of reproductive status .
Examples such as D. pulex highlight the importance of an unbiased examination of global gene expression patterns in establishing markers for sexual reproduction in rotifers. Our transcriptome analysis of B. calyciflorus reveals differential expression of genes that have roles in sexual processes in other systems as well as many genes that have no identifiable homologs in NCBI that may represent fast evolving or rotifer-specific genes. With this more informed frame of reference, we have provided an initial analysis of bdelloid transcriptome data that provides us with unique insights relating to the molecular nature of long-term asexual egg production and/or any potential for meiosis or the inclusion of meiotic processes during oogenesis in bdelloids.
Special thanks to Cynthia Toll and the Roy J. Carver Center for Genomics at the University of Iowa for technical assistance. Thanks also to Danielle Beekman and Elizabeth Savelkoul for helpful discussion and comments on the manuscript. This work was funded by National Institutes of Health Institute of General Medical Sciences (grant number 5R01GM079484, to JML and DMW).
- Weismann A: On the significance of the polar globules. Nature. 1887, 36: 607-609.Google Scholar
- Bell G: The masterpiece of nature: the evolution and genetics of sexuality. 1982, Berkeley: University of California PressGoogle Scholar
- Maynard Smith J: The evolution of sex. 1978, London, New York, Melbourne: Cambridge University Press, 1Google Scholar
- Ramesh MA, Malik S-B, Logsdon JM: A phylogenomic inventory of meiotic genes: evidence for sex in Giardia and an early eukaryotic origin of meiosis. Curr Biol. 2005, 15: 185-191.PubMedGoogle Scholar
- Bode SNS, Adolfsson S, Lamatsch DK, Martins MJF, Schmit O, Vandekerkhove J, Mezquita F, Namiotko T, Rossetti G, Schön I, et al: Exceptional cryptic diversity and multiple origins of parthenogenesis in a freshwater ostracod. Molecular Phylogenetics and Evolution. 2010, 54 (2): 542-552. 10.1016/j.ympev.2009.08.022.PubMedGoogle Scholar
- Fernãndez R, Almodóvar A, Novo M, Simancas B, Cosín DJD: Adding complexity to the complex: new insights into the phylogeny, diversification and origin of parthenogenesis in the Aporrectodea caliginosa species complex (Oligochaeta, Lumbricidae). Molecular Phylogenetics and Evolution. 2012, 64 (2): 368-379. 10.1016/j.ympev.2012.04.011.PubMedGoogle Scholar
- West SA, Lively CM, Read AF: A pluralist approach to sex and recombination. J Evol Biol. 1999, 12: 1003-1012. 10.1046/j.1420-9101.1999.00119.x.Google Scholar
- Kondrashov AS: Classification of hypotheses on the advantage of amphimixis. J Hered. 1993, 84: 372-387.PubMedGoogle Scholar
- Meirmans S, Strand R: Why are there so many theories for sex, and what do we do with them?. J Hered. 2010, 101 (Supplement 1): S3-S12. 10.1093/jhered/esq021.PubMedGoogle Scholar
- Mark Welch D, Meselson M: Evidence for the evolution of bdelloid rotifers without sexual reproduction or genetic exchange. Science. 2000, 288: 1211-1215. 10.1126/science.288.5469.1211.PubMedGoogle Scholar
- Mark Welch DB, Meselson M: Rates of nucleotide substitution in sexual and anciently asexual rotifers. Proc Natl Acad Sci USA. 2001, 98 (12): 6720-6724. 10.1073/pnas.111144598.PubMedGoogle Scholar
- Gilbert JJ: Environmental and endogenous control of sexuality in a rotifer life cycle: developmental and population biology. Evol Dev. 2003, 5 (1): 19-24. 10.1046/j.1525-142X.2003.03004.x.PubMedGoogle Scholar
- Snell TW, Boyer EM: Thresholds for mictic female production in the rotifer Brachionus plicatilis (Muller). J Exp Mar Biol Ecol. 1988, 124: 73-85. 10.1016/0022-0981(88)90112-8.Google Scholar
- Snell TW, Kubanek J, Carter W, Payne AB, Kim J, Hicks MK, Stelzer C-P: A protein signal triggers sexual reproduction in Brachionus plicatilis (Rotifera). Mar Biol. 2006, 149: 763-773. 10.1007/s00227-006-0251-2.Google Scholar
- Gilbert JJ: Dormancy in rotifers. Trans Amer Micros Soc. 1974, 93 (4): 490-513. 10.2307/3225154.Google Scholar
- Stelzer C-P: Obligate asex in a rotifer and the role of sexual signals. J Evol Biol. 2008, 21: 287-293.PubMedGoogle Scholar
- Fussmann GF, Ellner SP, Hairston NG: Evolution as a critical component of plankton dynamics. Proc R Soc B. 2003, 270: 1015-1022. 10.1098/rspb.2003.2335.PubMed CentralPubMedGoogle Scholar
- Stelzer C-P, Schmidt J, Wiedlroither A, Riss S: Loss of sexual reproduction and dwarfing in a small metazon. PLoS One. 2010, 5 (9): e12854-10.1371/journal.pone.0012854.PubMed CentralPubMedGoogle Scholar
- Stelzer C-P: The cost of sex and competition between cyclical and obligate parthenogenetic rotifers. Am Nat. 2011, 177 (2): E43-E53. 10.1086/657685.PubMedGoogle Scholar
- Stout EP, La Clair JJ, Snell TW, Shearer TL, Kubanek J: Conservation of progesterone hormone function in invertebrate reproduction. Proc Natl Acad Sci USA. 2010, 107 (26): 11859-11864. 10.1073/pnas.1006074107.PubMed CentralPubMedGoogle Scholar
- Snell TW, DesRosiers NJD: Effect of progesterone on sexual reproduction of Brachionus manjavacas (Rotifera). J Exp Mar Biol Ecol. 2008, 363: 104-109. 10.1016/j.jembe.2008.06.031.Google Scholar
- Yang J, Snell TW: Effects of progesterone, testosterone, and estrogen on sexual reproduction of the rotifer Brachionus calyciflorus. Int Rev Hydrobiol. 2010, 95 (6): 441-449. 10.1002/iroh.201011267.Google Scholar
- Denekamp NY, Thorne MAS, Clark MS, Kube M, Reinhardt R, Lubzens E: Discovering genes associated with dormancy in the monogonont rotifer Brachionus plicatilis. BMC Genomics. 2009, 10 (108): 1-17.Google Scholar
- Clark MS, Denekamp NY, Thorne MAS, Reinhardt R, Drungowski M, Albrecht MW, Klages S, Beck A, Kube M, Lubzens E: Long-term survival of hydrated resting eggs from Brachionus plicatilis. PLoS One. 2012, 7 (1): e29365-10.1371/journal.pone.0029365.PubMed CentralPubMedGoogle Scholar
- Kilham SS, Kreeger DA, Lynn SG, Goulden CE, Herrera L: COMBO: A defined freshwater culture medium for algae and zooplankton. Hydrobiologia. 1998, 377: 147-159. 10.1023/A:1003231628456.Google Scholar
- Stelzer C-P, Snell TW: Induction of sexual reproduction in Brachionus plicatilis (Monogononta, Rotifera) by a density-dependent chemical cue. Limnol Oceanogr. 2003, 48 (2): 939-943. 10.4319/lo.2003.48.2.0939.Google Scholar
- Goecks J, Nekrutenko A, Taylor J, Team TG: Galaxy: a comprehensive approach for supporting accessible, reproducible, and transparent computational research in the life sciences. Genome Biol. 2010, 11 (8): R86-10.1186/gb-2010-11-8-r86.PubMed CentralPubMedGoogle Scholar
- Blankenberg D, Von Kuster G, Coraor N, Ananda G, Lazarus R, Mangan M, Nekrutenko A, Taylor J: Galaxy: a web-based genome analysis tool for experimentalists. Current Protocols in Molecular Biology. 2010, 19.10: 11-21.Google Scholar
- Giardine B, Riemer C, Hardison RC, Burhans R, Elnitski L, Shah P, Zhang Y, Blankenberg D, Albert I, Taylor J, et al: Galaxy: a platform for interactive large-scale genome analysis. Genome Res. 2005, 15 (10): 1451-1455. 10.1101/gr.4086505.PubMed CentralPubMedGoogle Scholar
- Merchant SS, Prochnik SE, Vallon O, Harris EH, Karpowicz SJ, Witman GB, Terry A, Salamov A, Fritz-Laylin LK, Marechal-Drouard L, et al: The Chlamydomonas genome reveals the evolution of key animal and plant functions. Science. 2007, 318 (5848): 245-250. 10.1126/science.1143609.PubMed CentralPubMedGoogle Scholar
- Hanson SJ, Schurko AM, Hecox-Lea B, Mark Welch D, Stelzer C-P, Logsdon JM: Inventory and phylogenetic analysis of meiotic genes in monogonont rotifers. J Hered. 2013, 104 (3): 357-370. 10.1093/jhered/est011.PubMed CentralPubMedGoogle Scholar
- Trapnell C, Pachiter L, Salzber SL: TopHat: discovering splice junctions with RNA-Seq. Bioinformatics. 2009, 25 (9): 1105-1111. 10.1093/bioinformatics/btp120.PubMed CentralPubMedGoogle Scholar
- Trapnell C, Roberts A, Goff L, Pertea G, Kim D, Kelley DR, Pimentel H, Salzberg SL, Rinn JL, Pachter L: Differential gene and transcript expression analysis of RNA-seq experiments with TopHat and Cufflinks. Nat Protoc. 2012, 7: 562-578.PubMed CentralPubMedGoogle Scholar
- Robinson MD, Mccarthy DJ, Smyth GK: EdgeR: a Bioconductor package for differential expression analysis of digital gene expression data. Bioinformatics. 2010, 26 (1): 139-140. 10.1093/bioinformatics/btp616.PubMed CentralPubMedGoogle Scholar
- Larkin MA, Blackshields G, Brown NP, Chenna R, Mcgettigan PA, Mcwilliam H, Valentin F, Wallace IM, Wilm A, Lopez R, et al: Clustal W and Clustal X version 2.0. Bioinformatics. 2007, 23 (21): 2947-2948. 10.1093/bioinformatics/btm404.PubMedGoogle Scholar
- Kumar S, Skjæveland Å, Orr RJ, Enger P, Ruden T, Mevik B-H, Burki F, Botnen A, Shalchian-Tabrizi K: AIR: a batch-oriented web program package for construction of supermatrices ready for phylogenomic analyses. BMC Bioinforma. 2009, 10 (1): 357-10.1186/1471-2105-10-357.Google Scholar
- Guindon SXEP, Gascuel O: A simple, fast, and accurate algorithm to estimate large phylogenies by maximum likelihood. Syst Biol. 2003, 52 (5): 696-704. 10.1080/10635150390235520.PubMedGoogle Scholar
- Camacho C, Coulouris G, Avagyan V, Ma N, Papadopoulos J, Bealer K, Madden TL: BLAST+: architecture and applications. BMC Bioinforma. 2009, 10 (1): 421-10.1186/1471-2105-10-421.Google Scholar
- Conesa A, Götz S: Blast2GO: a comprehensive suite for functional analysis in plant genomics. International Journal of Plant Genomics. 2008, 2008: 1-12.Google Scholar
- Snell TW, Shearer TL, Smith HA, Kubanek J, Gribble KE, Mark Welch DB: Genetic determinants of mate recognition in Brachionus manjavacas (Rotifera). BMC Biol. 2009, 7 (60): 1-12.Google Scholar
- Gomez A, Serra M: Mate choice in male Brachionus plicatilis rotifers. Funct Ecol. 1996, 10 (6): 681-687. 10.2307/2390502.Google Scholar
- Suga K, Mark Welch D, Tanaka Y, Sakakura Y, Hagiwara A: Analysis of expressed sequence tags of the cyclically parthenogenetic rotifer Brachionus plicatilis. PLoS One. 2007, 2 (8): e671-10.1371/journal.pone.0000671.PubMed CentralPubMedGoogle Scholar
- Denekamp NY, Suga K, Hagiwara A, Reinhardt R, Lubzens E: A role for molecular studies in unveiling the pathways for formation of rotifer resting eggs and their survival during dormancy. Dormancy and Resistance in Harsh Environments, Topics in Current Genetics. 2010, 21: 109-132. 10.1007/978-3-642-12422-8_7.Google Scholar
- Boschetti C, Carr A, Crisp A, Eyres I, Wang-Koh Y, Lubzens E, Barraclough TG, Micklem G, Tunnacliffe A: Biochemical diversification through foreign gene expression in bdelloid rotifers. PLoS Genetics. 2012, 8 (11): e1003035-10.1371/journal.pgen.1003035.PubMed CentralPubMedGoogle Scholar
- Gallardo W, Hagiwara A, Tomita Y, Soyano K, Snell T: Effect of some vertebrate and invertebrate hormones on the population growth, mictic female production, and body size of the marine rotifer Brachionus plicatilis Muller. Hydrobiologia. 1997, 358: 113-120. 10.1023/A:1003124205002.Google Scholar
- Bridgham JT, Eick GN, Larroux C, Deshpande K, Harms MJ, Gauthier MEA, Ortlund EA, Degnan BM, Thornton JW: Protein evolution by molecular tinkering: diversification of the nuclear receptor superfamily from a ligand-dependent ancestor. PLoS Biol. 2010, 8 (10): e1000497-10.1371/journal.pbio.1000497.PubMed CentralPubMedGoogle Scholar
- Bowles J, Koopman P: Retinoic acid, meiosis and germ cell fate in mammals. Development. 2007, 134 (19): 3401-3411. 10.1242/dev.001107.PubMedGoogle Scholar
- Bagamasbad P, Denver RJ: Mechanisms and significance of nuclear receptor auto- and cross-regulation. Gen Comp Endocrinol. 2011, 170 (1): 3-17. 10.1016/j.ygcen.2010.03.013.PubMed CentralPubMedGoogle Scholar
- Gronemeyer H, Gustafsson J-Å, Laudet V: Principles for modulation of the nuclear receptor superfamily. Nat Rev Drug Discov. 2004, 3 (11): 950-964. 10.1038/nrd1551.PubMedGoogle Scholar
- Shi Y, Mello C: A CBP/p300 homolog specifies multiple differentiation pathways in Caenorhabditis elegans. Genes Dev. 1998, 12: 943-955. 10.1101/gad.12.7.943.PubMed CentralPubMedGoogle Scholar
- Kovo M, Kandli-Cohen M, Ben-Haim M, Galiani D, Carr D, Dekel N: An active protein kinase A (PKA) is involved in meiotic arrest of rat growing oocytes. Reproduction. 2006, 132: 33-43. 10.1530/rep.1.00824.PubMedGoogle Scholar
- Viegas LR, Vicent GP, Baranao JL, Beato M, Pecci A: Steroid hormones induce bcl-x gene expression through direct activation of distal promoter P4. J Biol Chem. 2003, 279 (11): 9831-9839. 10.1074/jbc.M312402200.PubMedGoogle Scholar
- Schurko AM, Logsdon JM: Using a meiosis detection toolkit to investigate ancient asexual “scandals” and the evolution of sex. BioEssays. 2008, 30: 579-589. 10.1002/bies.20764.PubMedGoogle Scholar
- Pesin J, Orr-Weaver T: Regulation of APC/C activators in mitosis and meiosis. Annual Review of Cell and Developmental Biology. 2008, 24: 475-499. 10.1146/annurev.cellbio.041408.115949.PubMed CentralPubMedGoogle Scholar
- Swan A, Schupbach T: The Cdc20 (Fzy)/Cdh1-related protein, Cort, cooperates with Fzy in cyclin destruction and anaphase progression in meiosis I and II in Drosophila. Development. 2007, 134 (5): 891-899. 10.1242/dev.02784.PubMed CentralPubMedGoogle Scholar
- Talbert PB, Henikoff S: Histone variants — ancient wrap artists of the epigenome. Nat Rev Mol Cell Biol. 2010, 11 (4): 264-275. 10.1038/nrm2861.PubMedGoogle Scholar
- Van Doninck K, Mandigo M, Hur J, Wang P, Guglielmini J, Milinkovitch M, Lane W, Meselson M: Phylogenomics of unusual histone H2A variants in bdelloid rotifers. PLoS Genetics. 2009, 5 (3): e1000401-10.1371/journal.pgen.1000401.PubMed CentralPubMedGoogle Scholar
- Ehinger A, Denison SH, May GS: Sequence, organization and expression of the core histone genes of Aspergillus nidulans. Mol Gen Genet. 1990, 222: 416-424. 10.1007/BF00633848.PubMedGoogle Scholar
- Buschbeck M, Uribesalgo I, Wibowo I, Rué P, Martin D, Gutierrez A, Morey L, Guigó R, López-Schier H, Croce LD: The histone variant macroH2A is an epigenetic regulator of key developmental genes. Nat Struct Mol Biol. 2009, 16 (10): 1074-1079. 10.1038/nsmb.1665.PubMedGoogle Scholar
- O’Donnell L, Panier S, Wildenhain J, Tkach JM, Al-Hakim A, Landry M-C, Escribano-Diaz C, Szilard RK, Young JTF, Munro M, et al: The MMS22L-TONSL complex mediates recovery from replication stress and homologous recombination. Molecular Cell. 2010, 40 (4): 619-631. 10.1016/j.molcel.2010.10.024.PubMed CentralPubMedGoogle Scholar
- Schwab KR, Patel SR, Dressler GR: Role of PTIP in class switch recombination and long-range chromatin interactions at the immunoglobulin heavy chain locus. Mol Cell Biol. 2011, 31 (7): 1503-1511. 10.1128/MCB.00990-10.PubMed CentralPubMedGoogle Scholar
- Stein AB, Jones TA, Herron TJ, Patel SR, Day SM, Noujaim SF, Milstein ML, Klos M, Furspan PB, Jalife J, et al: Loss of H3K4 methylation destabilizes gene expression patterns and physiological functions in adult murine cardiomyocytes. J Clin Invest. 2011, 121 (7): 2641-2650. 10.1172/JCI44641.PubMed CentralPubMedGoogle Scholar
- Kota SK, Feil R: Epigenetic transitions in germ cell development and meiosis. Developmental Cell. 2010, 19 (5): 675-686. 10.1016/j.devcel.2010.10.009.PubMedGoogle Scholar
- Petersen C, Fuzesi L, Hoyer-Fender S: Outer dense fibre proteins from human sperm tail: molecular cloning and expression analyses of two cDNA transcripts encoding proteins of 70 kDa. Mol Hum Reprod. 1999, 5 (7): 627-635. 10.1093/molehr/5.7.627.PubMedGoogle Scholar
- Soung N-K, Kang YH, Kim K, Kamijo K, Yoon H, Seong Y-S, Kuo Y-L, Miki T, Kim SR, Kuriyama R, et al: Requirement of hCenexin for proper mitotic functions of Polo-Like Kinase 1 at the centrosomes. Mol Cell Biol. 2006, 26 (22): 8316-8335. 10.1128/MCB.00671-06.PubMed CentralPubMedGoogle Scholar
- Kiselak EA, Shen X, Song J, Gude DR, Wang J, Brody SL, Strauss JF, Zhang Z: Transcriptional regulation of an axonemal central apparatus gene, Sperm-associated Antigen 6, by a SRY-related high mobility group transcription factor, S-SOX5. J Biol Chem. 2010, 285 (40): 30496-30505. 10.1074/jbc.M110.121590.PubMed CentralPubMedGoogle Scholar
- Zhang Z, Jones BH, Tang W, Moss SB, Wei Z, Ho C, Pollack M, Horowitz E, Bennett J, Baker ME, et al: Dissecting the axoneme interactome: The mammalian orthologue of Chlamydomonas PF6 interacts with Sperm-Associated Antigen 6, the mammalian orthologue of Chlamydomonas PF16. Molecular & Cellular Proteomics. 2005, 4 (7): 914-923. 10.1074/mcp.M400177-MCP200.Google Scholar
- Zhang Z, Kostetskii I, Moss SB, Jones BH, Ho C, Wang H, Kishida T, Gerton GL, Radice GL, Strauss JF: Haploinsufficiency for the murine orthologue of Chlamydomonas PF20 disrupts spermatogenesis. Proc Natl Acad Sci. 2004, 101 (35): 12946-12951. 10.1073/pnas.0404280101.PubMed CentralPubMedGoogle Scholar
- Chan SW, Fowler KJ, Choo KHA, Kalitsis P: Spef1, a conserved novel testis protein found in mouse sperm flagella. Gene. 2005, 353: 189-199. 10.1016/j.gene.2005.04.025.PubMedGoogle Scholar
- Sironen A, Kotaja N, Mulhern H, Wyatt TA, Sisson JH, Pavlik JA, Miiluniemi M, Fleming MD, Lee L: Loss of SPEF2 function in mice results in spermatogenesis defects and primary ciliary dyskinesia. Biol Reprod. 2011, 85 (4): 690-701. 10.1095/biolreprod.111.091132.PubMed CentralPubMedGoogle Scholar
- Yukitake H, Furusawa M, Taira T, Iguchi-Ariga SMM, Ariga H: AAT-1, a novel testis-specific AMY-1-binding protein, forms a quaternary complex with AMY-1, A-kinase Anchor Protein 84, and a regulatory subunit of cAMP-dependent protein kinase and is phosphorylated by its kinase. J Biol Chem. 2002, 277 (47): 45480-45492. 10.1074/jbc.M206201200.PubMedGoogle Scholar
- Alekseev OM, Richardson RT, O’rand MG: Linker histones stimulate HSPA2 ATPase activity through NASP binding and inhibit CDC2/Cyclin B1 complex formation during meiosis in the mouse. Biol Reprod. 2009, 81 (4): 739-748. 10.1095/biolreprod.109.076497.PubMed CentralPubMedGoogle Scholar
- Takahashi T, Tanaka H, Iguchi N, Kitamura K, Chen Y-F, Maekawa M, Nishimura H, Ohta H, Miyagawa Y, Matsumiya K, et al: Rosbin: A novel homeobox-like protein gene expressed exclusively in round spermatids. Biol Reprod. 2004, 70 (5): 1485-1492. 10.1095/biolreprod.103.026096.PubMedGoogle Scholar
- Kanazawa R, Komori S, Sakata K, Tanaka H, Sawai H, Tsuji Y, Koyama K: Isolation and characterization of a human sperm antigen gene h-Sp-1. Int J Androl. 2003, 26: 226-235. 10.1046/j.1365-2605.2003.00418.x.PubMedGoogle Scholar
- Liu N, Qiao Y, Cai C, Lin W, Zhang J, Miao S, Zong S, Koide SS, Wang L: A sperm component, HSD-3.8 (SPAG1), interacts with G-protein beta 1 subunit and activates extracellular signal-regulated kinases (ERK). Front Biosci. 2006, 11: 1679-1689. 10.2741/1913.PubMedGoogle Scholar
- Wu H, Chen Y, Miao S, Zhang C, Zong S, Koide SS, Wang L: Sperm associated antigen 8 (SPAG8), a novel regulator of activator of CREM in testis during spermatogenesis. FEBS Lett. 2010, 584 (13): 2807-2815. 10.1016/j.febslet.2010.05.016.PubMedGoogle Scholar
- Greenbaum MP, Yan W, Wu M, Lin Y, Agno J, Sharma M, Braun RE, Rajkovic A, Matzuk MM: TEX14 is essential for intercellular bridges and fertility in male mice. Proc Natl Acad Sci. 2006, 103 (13): 4982-4987. 10.1073/pnas.0505123103.PubMed CentralPubMedGoogle Scholar
- Mondal G, Ohashi A, Yang L, Rowley M, Couch Fergus J: Tex14, a Plk1-regulated protein, is required for kinetochore-microtubule attachment and regulation of the spindle assembly checkpoint. Molecular Cell. 2012, 45 (5): 680-695. 10.1016/j.molcel.2012.01.013.PubMed CentralPubMedGoogle Scholar
- Xu B, Hao Z, Jha KN, Zhang Z, Urekar C, Digilio L, Pulido S, Strauss JF, Flickinger CJ, Herr JC: Targeted deletion of Tssk1 and 2 causes male infertility due to haploinsufficiency. Dev Biol. 2008, 319: 211-222. 10.1016/j.ydbio.2008.03.047.PubMed CentralPubMedGoogle Scholar
- Daniel R, Daniels E, He Z, Bateman A: Progranulin (acrogranin/PC cell-derived growth factor/granulin-epithelin precursor) is expressed in the placenta, epidermis, microvasculature, and brain during murine development. Dev Dyn. 2003, 227 (4): 593-599. 10.1002/dvdy.10341.PubMedGoogle Scholar
- Lee YJ, Lee Y, Chung JH: An intronless gene encoding a poly(A) polymerase is specifically expressed in testis. FEBS Lett. 2000, 487: 287-292. 10.1016/S0014-5793(00)02367-X.PubMedGoogle Scholar
- Liu Y, Black J, Kisiel N, Kulesz-Martin MF: SPAF, a new AAA-protein specific to early spermatogenesis and malignant conversion. Oncogene. 2000, 19: 1579-1588. 10.1038/sj.onc.1203442.PubMedGoogle Scholar
- Zhang X, Liu H, Zhang Y, Qiao Y, Miao S, Want L, Zhang J, Zong S, Koide SS: A novel gene, RSD-3/HSD-3.1, encodes a meiotic-related protein expressed in rat and human testis. J Mol Med. 2003, 81 (81): 380-387.PubMedGoogle Scholar
- Deng Y, Hu L-S, Lu G-X: Expression and identification of a novel apoptosis gene Spata17 (MSRG-11) in mouse spermatogenic cells. Acta Biochim Biophys Sinica. 2006, 38 (1): 37-45. 10.1111/j.1745-7270.2006.00125.x.Google Scholar
- Bornstein C, Brosh R, Molchadsky A, Madar S, Kogan-Sakin I, Goldstein I, Chakravarti D, Flores ER, Goldfinger N, Sarig R, et al: SPATA18, a Spermatogenesis-Associated Gene, is a novel transcriptional target of p53 and p63. Mol Cell Biol. 2011, 31 (8): 1679-1689. 10.1128/MCB.01072-10.PubMed CentralPubMedGoogle Scholar
- Senoo M, Hoshino S, Mochida N, Matsumura Y, Habu S: Identification of a Novel Protein p59scr, Which Is Expressed at Specific Stages of Mouse Spermatogenesis. Biochem Biophys Res Commun. 2002, 292 (4): 992-998. 10.1006/bbrc.2002.6769.PubMedGoogle Scholar
- Song X, Li Y, Shi Y, Hu X, Hu Z, Han C, Liu Y: Cloning and characterization of a novel spermiogenesis-related gene, T6441, in rat testis. Front Biosci. 2006, 11: 143-150. 10.2741/1787.PubMedGoogle Scholar
- Oh C, Aho H, Shamsadin R, Nayernia K, Muller C, Sancken U, Szpirer C, Engel W, Adham IM: Characterization, expression pattern and chromosomal localization of the spermatogenesis associated 6 gene (Spata6). Mol Hum Reprod. 2003, 9 (6): 321-330. 10.1093/molehr/gag047.PubMedGoogle Scholar
- Li Y, Maines JZ, Tastan OY, Mckearin DM, Buszczak M: Mei-P26 regulates the maintenance of ovarian germline stem cells by promoting BMP signaling. Development. 2012, 139 (9): 1547-1556. 10.1242/dev.077412.PubMed CentralPubMedGoogle Scholar
- Liu N, Han H, Lasko P: Vasa promotes Drosophila germline stem cell differentiation by activating mei-P26 translation by directly interacting with a (U)-rich motif in its 3′ UTR. Genes Dev. 2009, 23 (23): 2742-2752. 10.1101/gad.1820709.PubMed CentralPubMedGoogle Scholar
- Quenet D, Mark M, Govin J, van Dorsselear A, Schreiber V, Khochbin S, Dantzer F: Parp2 is required for the differentiation of post-meiotic germ cells: identification of a spermatid-specific complex containing Parp1, Parp2, TP2 and HSPA2. Experimental Cell Research. 2009, 315 (16): 2824-2834. 10.1016/j.yexcr.2009.07.003.PubMedGoogle Scholar
- Chi MN, Auriol J, Jegou B, Kontoyiannis DL, Turner JMA, De Rooij DG, Morello D: The RNA-binding protein ELAVL1/HuR is essential for mouse spermatogenesis, acting both at meiotic and postmeiotic stages. Molecular Biology of the Cell. 2011, 22 (16): 2875-2885. 10.1091/mbc.E11-03-0212.PubMed CentralPubMedGoogle Scholar
- Parfitt DA, Michael GJ, Vermeulen EGM, Prodromou NV, Webb TR, Gallo J-M, Cheetham ME, Nicoll WS, Blatch GL, Chapple JP: The ataxia protein sacsin is a functional co-chaperone that protects against polyglutamine-expanded ataxin-1. Hum Mol Genet. 2009, 18 (9): 1556-1565. 10.1093/hmg/ddp067.PubMed CentralPubMedGoogle Scholar
- Kruse E, Uehlein N, Kaldenhoff R: The aquaporins. Genome Biol. 2006, 7 (2): 206-10.1186/gb-2006-7-2-206.PubMed CentralPubMedGoogle Scholar
- Britton C, Murray L: Cathepsin L protease (CPL-1) is essential for yolk processing during embryogenesis in Caenorhabditis elegans. Journal of Cell Science. 2004, 117 (21): 5133-5143. 10.1242/jcs.01387.PubMedGoogle Scholar
- Gilbert JJ: Females from resting eggs and parthenogenetic eggs in the rotifer Brachionus calyciflorus: lipid droplets, starvation resistance and reproduction. Freshw Biol. 2004, 49: 1505-1515. 10.1111/j.1365-2427.2004.01282.x.Google Scholar
- 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
- Caprioli M, Katholm AK, Melone G, Ramlov H, Ricci C, Santo N: Trehalose in desiccated rotifers: a comparison between a bdelloid and a monogonont species. Comparative Biochemistry and Physiology. 2004, 139: 527-532. 10.1016/j.cbpb.2004.10.019.PubMedGoogle Scholar
- Erkut C, Penkov S, Khesbak H, Vorkel D, Verbavatz J-M, Fahmy K, Kurzchalia Teymuras V: Trehalose renders the dauer larva of Caenorhabditis elegans resistant to extreme desiccation. Curr Biol. 2011, 21: 1331-1336. 10.1016/j.cub.2011.06.064.PubMedGoogle Scholar
- Denekamp NY, Reinhardt R, Kube M, Lubzens E: Late Embryogenesis Abundant (LEA) Proteins in nondesiccated, encysted, and diapausing embryos of rotifers. Biol Reprod. 2010, 82 (4): 714-724. 10.1095/biolreprod.109.081091.PubMedGoogle Scholar
- Warner AH, Miroshnychenko O, Koxarova A, Vacratsis PO, MacRae TH, Kim J, Clegg JS: Evidence for multiple group 1 late embryogenesis abundant (LEA) proteins in encysted embryos of Artemia and their organelles. J Biochem. 2010, 148: 581-592. 10.1093/jb/mvq091.PubMedGoogle Scholar
- Zamudio N, Bourc’his D: Transposable elements in the mammalian germline: a comfortable niche or a deadly trap?. Heredity. 2010, 105 (1): 92-104. 10.1038/hdy.2010.53.PubMedGoogle Scholar
- Bao J, Yan W: Male germline control of transposable elements. Biol Reprod. 2012, 86 (5): 162-10.1095/biolreprod.111.095463. 162PubMed CentralPubMedGoogle Scholar
- Mostowy S, Cossart P: Septins: The fourth component of the cytoskeleton. Nature Publishing Group. 2012, 13 (3): 183-194.Google Scholar
- Ricci C: Anhydrobiotic capabilities of bdelloid rotifers. Hydrobiologia. 1998, 387/388: 321-326.Google Scholar
- Krisko A, Leroy M, Radman M, Meselson M: Extreme anti-oxidant protection against ionizing radiation in bdelloid rotifers. Proc Natl Acad Sci USA. 2012, 109 (7): 2354-2357. 10.1073/pnas.1119762109.PubMed CentralPubMedGoogle Scholar
- Kunz C, Schar P: Meiotic recombination: sealing the partnership at the junction. Curr Biol. 2004, 14: R962-R964. 10.1016/j.cub.2004.10.043.PubMedGoogle Scholar
- Gilbert JJ: Rotifera. Reproductive Biology of Invertebrates. 1989, 4 part A: 179-199.Google Scholar
- Schurko AM, Logsdon JM, Eads BD: Meiosis genes in Daphnia pulex and the role of parthenogenesis in genome evolution. BMC Evol Biol. 2009, 9 (78): 1-27.Google Scholar
- Hiruta C, Nishida C, Tochinai S: Abortive meiosis in the oogenesis of parthenogenetic Daphnia pulex. Chromosome Res. 2010, 18: 833-840. 10.1007/s10577-010-9159-2.PubMedGoogle Scholar
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