Polysome profiling reveals broad translatome remodeling during endoplasmic reticulum (ER) stress in the pathogenic fungus Aspergillus fumigatus
© Krishnan et al.; licensee BioMed Central Ltd. 2014
Received: 17 October 2013
Accepted: 17 February 2014
Published: 25 February 2014
The unfolded protein response (UPR) is a network of intracellular signaling pathways that supports the ability of the secretory pathway to maintain a balance between the load of proteins entering the endoplasmic reticulum (ER) and the protein folding capacity of the ER lumen. Current evidence indicates that several pathogenic fungi rely heavily on this pathway for virulence, but there is limited understanding of the mechanisms involved. The best known functional output of the UPR is transcriptional upregulation of mRNAs involved in ER homeostasis. However, this does not take into account mechanisms of translational regulation that involve differential loading of ribosomes onto mRNAs. In this study, a global analysis of transcript-specific translational regulation was performed in the pathogenic mold Aspergillus fumigatus to determine the nature and scope of the translational response to ER stress.
ER stress was induced by treating the fungus with dithiothreitol, tunicamycin, or a thermal up-shift. The mRNAs were then fractionated on the basis of ribosome occupancy into an under-translated pool (U) and a well-translated pool (W). The mRNAs were used to interrogate microarrays and the ratio of the hybridization signal (W/U) was used as an indicator of the relative translational efficiency of a mRNA under each condition. The largest category of translationally upregulated mRNAs during ER stress encoded proteins involved in translation. Components of the ergosterol and GPI anchor biosynthetic pathways also showed increased polysome association, suggesting an important role for translational regulation in membrane and cell wall homeostasis. ER stress induced limited remodeling of the secretory pathway translatome. However, a select group of transcription factors was translationally upregulated, providing a link to subsequent modification of the transcriptome. Finally, we provide evidence that one component of the ER stress translatome is a novel mRNA isoform from the yvc1 gene that is induced by ER stress in a UPR-dependent manner.
Together, these findings define a core set of mRNAs subject to translational control during the adaptive response to acute ER stress in A. fumigatus and reveal a remarkable breadth of functions that are needed to resolve ER stress in this organism.
KeywordsAspergillus fumigatus UPR Unfolded protein response ER stress Translational regulation Polysome profiling Yvc1
The opportunistic mold pathogen Aspergillus fumigatus causes life-threatening pulmonary infections that have the potential to progress into invasive aspergillosis, a disseminated disease with a very high rate of mortality [1, 2]. Infections with this fungus continue to impede the successful management of patients with hematologic malignancies or solid-organ and bone marrow transplants worldwide, accounting for the highest per person hospitalization costs of all the systemic mycoses [3–5]. The ongoing expansion of the immunosuppressed population is expected to increase the incidence of the disease, which is galvanizing studies to understand more about fungal stress response pathways that could yield novel vulnerabilities for future therapeutic targeting.
Current evidence indicates that pathogenic fungi are under endoplasmic reticulum (ER) stress in the host environment and therefore depend upon adaptive stress responses pathways to support their survival during infection [6–10]. The unfolded protein response (UPR) is the major ER stress response pathway, responsible for maintaining an ER lumenal environment that is conducive to optimal protein folding . A. fumigatus depends upon the UPR to support the expression of clinically relevant traits such as thermotolerance, cell wall/membrane homeostasis, hypoxia adaptation, iron homeostasis, nutrient assimilation from complex substrates and antifungal drug resistance [6, 7]. Similar findings have also been reported in Cryptococcus neoformans, Candida albicans, Candida glabrata, and Alternaria brassicicola, suggesting that the UPR is used by diverse fungal pathogens as a regulatory hub for the expression of multiple attributes that promote virulence in the host. The UPR is triggered in response to the accumulation of unfolded proteins, a condition that arises during infection when there is an imbalance between the level of nascent proteins entering the ER and the ability of the organelle to process that load. ER protein folding may also be perturbed by adverse conditions encountered in the host such as mammalian body temperature, oxidative stress, hypoxia and nutrient limitation . The UPR counters the resulting ER stress by expanding the quantity of ER-resident chaperones and folding enzymes that are needed to help membrane and secreted proteins achieve their native conformation. The current understanding of the fungal UPR is based upon the paradigm established in the model yeast Saccharomyces cerevisiae. The pathway is controlled by Ire1 (IreA in A. fumigatus), an ER-transmembrane protein that detects disturbances in the ER that lead to the accumulation of unfolded proteins. Ire1 contains a lumenal sensing domain and a cytosolic effector region that contains dual enzymes: a kinase linked to an endoribonuclease (RNase). In the absence of ER stress, Ire1p exists as an inactive monomer in association with the ER-resident chaperone, Bip (also known as Kar2p). When the folding capacity of the ER is exceeded, BiP dissociates from the lumenal domain to assist with protein folding. This triggers the activation of Ire1 by a ligand-dependent two-step mechanism in which BiP dissociation is followed by direct interaction of Ire1 with unfolded proteins [15–18]. These events elicit Ire1 oligomerization in the ER membrane, resulting in a conformational change that activates the C-terminal RNase [19, 20]. The substrate of this RNAse is a cytoplasmic mRNA known as HAC1 (hacA in A. fumigatus). The excision of an unconventional intron from the HAC1 mRNA allows in-frame translation of the bZIP transcription factor, Hac1 (HacA in A. fumigatus). Hac1 re-establishes ER homeostasis by remodeling the transcriptome to enhance the protein folding capacity of the ER.
Genome-wide expression profiling has demonstrated that A. fumigatus responds to acute ER stress by upregulating the levels of a core group of mRNAs that encode proteins with functions that support the secretory pathway . However, mRNA abundance measurements do not take translational efficiency into consideration, which is a mechanism of gene regulation that can have potent effects on protein production [21–23]. Translational regulation provides the cell with a rapid-response mechanism to fine-tune protein levels in proportion to need, and is particularly important in situations where an immediate response to an environmental stress is key for survival [24, 25]. Translational regulation can be studied on a global scale by interrogating microarrays with mRNAs that have been fractionated based upon ribosome occupancy . This approach is based on the fact that translationally quiescent mRNAs are sequestered within messenger ribonucleoprotein (mRNP) particles or associated with single ribosomes (monosomes), whereas actively translated mRNAs are associated with multiple ribosomes (polysomes). The hybridization of a microarray with these polysome-fractionated mRNAs can thus provide insight into how the translational efficiency of individual mRNAs is modified by environmental cues. Analogous approaches have been used to study the ER stress translatome in S. cerevisiae and Aspergillus niger[27, 28]. However, a global analysis of transcript-specific translational regulation has not been performed in A. fumigatus. In this study, polysome fractionation of mRNA was coupled with microarray detection in order to identify changes in the translational status of the A. fumigatus transcriptome under conditions that perturb ER homeostasis. The findings establish a core ER stress translatome and uncover evidence for extensive translational regulation during the response of A. fumigatus to ER stress.
Results & discussion
Translatome remodeling is a major component of the ER stress response in A. fumigatus
A total of 90 mRNAs showed a decrease in translational efficiency in response to DTT and TM treatment (Figure 2), 46 of which were unannotated. Analysis of the remaining 44 mRNAs using the Kyoto Encyclopedia of Genes and Genomes (KEGG) database revealed enrichment in metabolism and RNA processing categories, consistent with a shift towards more limited metabolic functions under ER stress conditions (Additional file 2). Of the 1253 mRNAs that showed altered translational efficiency in response to DTT alone, 536 were down-regulated. This contrasts a study of translational efficiency during DTT-induced stress in A. niger, which revealed down-regulation of 253 mRNAs and upregulation of 26 mRNAs. A direct comparison between these two datasets is difficult however, because the A. niger paper treated the fungus with 20 mM DTT for 2 h and fractionated mRNAs based upon occupancy with < 2 and ≥2 ribosomes , whereas our study used 1 mM DTT for 1 h and fractionation into pools associated with <5 and ≥5 ribosomes. The more severe ER stress conditions used in the A. niger study may account for the predominance of translationally repressed mRNAs in that organism, relative to the largely inductive response in A. fumigatus.
ER stress induces limited remodeling of the secretory pathway translatome
List of mRNAs with increased polysome association during ER stress (treatment with DTT or TM)
60S ribosomal protein L27a (AFUA_3G05600)
40S ribosomal protein S29 (AFUA_6G12720)
40S ribosomal protein S8 (AFUA_6G07360)#
50S ribosomal protein L36 (AFUA_4G12810)
37S ribosomal protein S16 (AFUA_5G08350)
Mitochondrial ribosomal protein L11 (AFUA_5G11830)
Ribosome biogenesis protein (AFUA_8G04790)
37S ribosomal protein S5 (AFUA_5G11540)
40S ribosomal protein S9 (AFUA_3G06970)#
40S ribosomal protein Rps16 (AFUA_2G10500)#
60S ribosomal protein L6 (AFUA_6G09060)
Aconitate hydratase/Mitochondrial ribosomal protein subunit L49 (AFUA_3G08080)#
Eukaryotic translation initiation factor 3 subunit eIF-Ca (AFUA_1G05200)
Translation elongation factor eEF-1B gamma subunit, putative (AFUA_1G17120)
GTP binding protein Guf1 (AFUA_3G14350)
Eukaryotic translation initiation factor 3 subunit eIF-Ck (AFUA_3G09280)
Cell membrane/cell wall
Squalene monooxygenase Erg1 (AFUA_5G07780)
Ergosterol biosynthesis protein Erg28 (AFUA_2G11550)
1,3-beta-glucanosyltransferase Gel2 (AFUA_6G11390)
Cell wall proline rich protein (AFUA_1G13450)
Actin cortical patch protein Sur7 (AFUA_2G02310)
CFEM domain protein (AFUA_6G14090)
GPI anchored protein (AFUA_2G07800)
GPI anchored dioxygenase (AFUA_3G01800)
Dolichol phosphate-mannose biosynthesis regulatory protein Dpm2 (AFUA_1G03020)
N-acetylglucosaminyl-phosphatidylinositol deacetylase, putative (AFUA_5G12550)
Integral membrane protein (Pth11) (AFUA_6G03600)#
Protein folding & modification
Alpha-1,2-mannosyltransferase (Alg2) (AFUA_5G13210)
Disulfide isomerase (TigA) (AFUA_5G12260)#
Protein disulfide isomerase Pdi1 (AFUA_2G06150)
N-acetyltransferase family protein (AFUA_4G10930)
N-acetyltransferase complex ARD1 subunit (AFUA_1G09600)
Prefoldin subunit 5 (AFUA_1G10740)#
Endosome/protein transport and sorting
Rho GTPase activator (Bem3) (AFUA_6G06400)
Fasciclin domain family protein (AFUA_1G14300)
Ras-like GTP-binding protein (AFUA_4G03100)
Endosomal cargo receptor (Erv14) (AFUA_6G07290)
Synaptobrevin-like protein Sybl1 (AFUA_6G11270)
RAB GTPase Vps21/Ypt51 (AFUA_3G10740)#
Vacuolar protein sorting 55 superfamily (AFUA_6G04780)#
AP-1 adaptor complex subunit sigma (AFUA_2G01570)
Mitochondrial import inner membrane translocase subunit (TIM22) (AFUA_5G02200)#
bZIP transcription factor JlbA/IDI-4 (AFUA_5G01650)#
CBF/NF-Y family transcription factor (AFUA_2G14250)
C6 transcription factor (AFUA_3G09130)
Transcription factor RfeF (AFUA_4G10200)
CP2 transcription factor (AFUA_1G17350)
bZIP transcription factor (LziP) (AFUA_1G16460)#
Transcription initiation factor TFIID, 31kd subunit, putative (AFUA_1G14600)
C6 transcription factor (AFUA_6G11230)
RNA polymerase II mediator complex component Srb8, putative (AFUA_3G06250)
CHCH domain protein (AFUA_3G06370)#
Nitrogen metabolite repression regulator NmrA (AFUA_5G02920)
General stress response phosphoprotein phosphatase Psr1/2 (AFUA_1G04790)#
Glutathione peroxidase Hyr1 (AFUA_3G12270)
ER stress increases the translational state of the translation machinery
We found that components of the translational machinery were subject to increased polysome association in the presence of either DTT or TM (Table 1). This was somewhat surprising, since previous studies have shown downregulation of ribosome biogenesis genes in A. niger and S. cerevisiae exposed to DTT [27, 28]. This discrepancy is likely to reflect the higher concentrations of DTT used in those studies, and/or species-specific differences in sensitivity to DTT. We speculate that a limited expansion of the translational apparatus is beneficial to A. fumigatus during ER stress because it provides a mechanism to rapidly increase the level of proteins that are needed to protect the ER from damage until the appropriate transcriptional modifications can be implemented. Since only a subset of the translational machinery was upregulated in A. fumigatus, a second possibility is that some of these proteins may have unrecognized ‘moonlighting’ functions that are relevant to ER stress responses, a possibility that is supported by an emerging literature on extra-ribosome functions for ribosomal proteins .
ER stress induces remodeling of the cell wall and membrane translatome
A. fumigatus β(1-3)glucanoxyltransferases (Gel1 and Gel2) catalyze the elongation of β(1-3) glucan side chains and influence morphogenesis and virulence [36, 37]. A previous report indicates that both Gel1 and Gel2 are constitutively transcribed in A. fumigatus. However, here we demonstrate that the translational efficiency of the gel2 mRNA increases 2.5 fold during ER stress, suggesting that an increase in Gel2 protein is needed to protect the wall under these conditions. Gel2 contains a glycosylphosphatidylinositol (GPI) anchor that tethers it to the plasma membrane , which facilitates its role in maintaining cell wall integrity. Interestingly, at least three other mRNAs encoding GPI-anchored proteins of unknown function also showed increased ribosome occupancy during ER stress. In addition, ER stress caused increased polysome association of the mRNA encoding the major regulatory component for the rate-limiting step in GPI anchor biosynthesis, Dpm2, as well as the subsequent enzyme in the pathway, AfPIG-L. Together, these findings argue that rapid translation of GPI-anchored proteins is necessary to protect the fungus under conditions that disrupt ER homeostasis, mostly likely due to their role in maintaining the cell wall [37–39]. It is worth noting that GPI anchor biosynthesis is an emerging target for the development of new antifungal therapy [40–42]. Further understanding of the mechanism(s) by which translational regulation impacts GPI anchor production could suggest novel strategies to enhance pharmacologic inhibition of this pathway.
Host-temperature adaptation involves distinct translatome remodeling
The ER stress translatome contains a UPR-dependent mRNA isoform
In summary, this study reports the first global analysis of transcript-specific translational regulation during ER stress in the pathogenic mold A. fumigatus. The results define a core ER stress translatome and demonstrate that translational regulation is a major component of the response of A. fumigatus to environmental conditions that perturb ER homeostasis. We also provide evidence that the ER stress translatome contains a previously unidentified mRNA that is induced by ER stress in a UPR-dependent manner. Together with our previous analysis of the UPR transcriptome, these findings begin to develop a comprehensive understanding of how A. fumigatus responds to ER stress. Since ER stress is experienced by several fungal pathogens in the host environment [6–10], further understanding of these pathways, and the mechanisms used to deploy them, may provide a new perspective on fungal pathogenesis and offer novel strategies for therapeutic intervention.
Strains and culture conditions
The wt strain used in this study is AfS28 (ΔakuA::ptrA) . Conidia were harvested from colonies grown on OSM plates (Aspergillus minimal medium containing 10 mM ammonium tartrate and osmotically stabilized with 1.2 M sorbitol). For analysis of the ER stress response, a 250 ml flask containing 50 ml of YG medium (0.5% yeast extract, 2% glucose) was inoculated with 5 X 107 conidia and incubated for 16 h with shaking (200 rpm) at 37°C. ER stress was then induced by treating the cultures with 1 mM DTT or 10 μg/ml TM for 1 h. Hyperosmotic stress was induced by adding NaCl to a final concentration of 0.8 M and continuing the incubation for an additional hour. For analysis of the heat-shock response, 250 ml flasks containing 50 ml of YG medium were inoculated with 5 × 107 conidia and incubated for 16 h with shaking (200 rpm) at room temperature. Thermal stress was then applied by transferring the flasks to a shaking 37°C incubator (200 rpm) and harvesting the mycelium after 30 min and 60 min of incubation.
Polysome fractionation and RNA extraction
Sample preparation and polysome analysis was performed as previously described, with modifications . Following the induction of ER or thermal stress in the cultures, translating polyribosomes were halted on the mRNAs by the addition of cycloheximide to 0.1 mg/ml and incubating at 37°C for an additional 5 min. The culture was then chilled in an ice bath for 5 min prior to harvesting the mycelium. The hyphae were washed twice with 5 ml of lysis buffer (10 mM Tris-HCl [pH 7.5], 100 mM NaCl, 30 mM MgCl2, 0.1 mg/ml cycloheximide, and 0.2 mg/ml heparin), flash frozen in liquid nitrogen and mechanically crushed. After resuspending in 0.5 ml of lysis buffer, the lysate was cleared by two subsequent microcentrifugation steps (15,000 x g, for 5 min at 4°C) and the RNA content in the supernatant was quantified by absorbance at 260 nm. Equal amounts of RNA (20-30 A260 units) were loaded onto a 12-ml linear sucrose gradient (7 - 47%) prepared in gradient buffer (50 mM Tris-acetate, 50 mM NH4Cl, 12 mM MgCl2, 1 mM DTT, and 0.2 mg/ml heparin). The gradients were centrifuged at 150,000 × g for 2.5 h at 4°C, using a Sorvall SW 41Ti rotor. Gradient analysis was performed using an ISCO gradient collector with continuous monitoring at 254 nm. Individual fractions were collected with a Foxy Jr. fraction collector and RNA was precipitated from 0.5 ml fractions by mixing with an equal volume of 6 M guanidine thiocyanate and 2 volumes of 100% ethanol and incubating overnight at -20°C. The RNA was pelleted, washed and resuspended using standard procedures. For microarray analysis, RNA from fractions containing less than 5 ribosomes/ mRNA (‘U’) or 5 or more ribosomes/mRNA (‘W’) were pooled and precipitated with 1.5 M LiCl, followed by washing to remove residual heparin. For northern blot analysis of erg1 expression, the sucrose gradient was divided into seven sequential fractions representing the entire gradient, and the RNA was precipitated as indicated above. For experiments that required unfractionated RNA (unfractionated controls for the thermal shift microarray experiment, northern blot analysis of erg1 mRNA, and RNA-seq analysis of DTT-treated cultures), the mycelium was crushed in liquid nitrogen and total RNA was extracted using the TRIZOL method .
The RNA labeling reactions and hybridizations were performed as described in the J. Craig Venter Institute (JCVI) standard operating procedure http://pfgrc.jcvi.org/index.php/microarray/protocols.html) and transcriptional profiles were generated by interrogating the Af293 spotted oligonucleotide microarray containing 10, 503 spots. Each gene was present in triplicate on the array, and all hybridizations were repeated in dye swap experiments. The data for each gene were averaged from the triplicate genes on each array and the duplicate dye swap experiment (a total of six readings for each gene) and the gene expression ratios were log2-transformed. Plotting open reading frame length against fold increase in the W fraction showed no bias towards longer transcripts, indicating that an increase in ribosome loading on a particular transcript is not an artifact of mRNA length (data not shown).
Functional annotation of genes present within the dataset was analyzed using FungiFun  and enrichment of functional groups was performed using FunCat method. Hierarchical clustering was performed using Cluster 3.0  and the cluster tree was visualized using JAVA Treeview . All RNA samples were hybridized with a reference sample obtained from Af293 in order to allow for cross-comparison. The translational efficiency of individual mRNAs during DTT/TM treatment was defined as the ratio of the hybridization signal in fraction-W over that of fraction-U, using a 2-fold difference between conditions as the cut-off value for a change in translational efficiency. Normalization to total mRNA abundance was not performed because the mRNAs that fit these criteria showed no increase in abundance under the same conditions .
The translational efficiency of individual mRNAs at 25°C and following a temperature shift to 37°C (after 30 min or 60 min) was defined as the ratio of the hybridization signal in fraction-W over that of fraction-U, using a 2-fold change between conditions as the cut-off value for a change in translational efficiency. In order to enrich for mRNAs that are predominantly regulated by changes in translational efficiency (as opposed to transcript abundance), the dataset was normalized to transcript levels in unfractionated RNA. RNA abundance was determined by interrogating the microarrays with unfractionated RNA and the change in the translational efficiency of each mRNA upon thermal shift was calculated as (fraction-W/fraction-U)/total transcript abundance.
RNA-seq was performed by the Genomics Sequencing Core (GSC) at the University of Cincinnati. Using TruSeq RNA sample preparation kit (Illumina), total RNA (RIN ≥ 7.0, Agilent 2100 Bioanalyzer) was converted into a library of template molecules suitable for subsequent cluster generation and sequencing by Illumina HiSeq. Poly(A)n mRNA was extracted and fragmented into smaller pieces (~140 nt). The cleaved RNA fragments were converted into first strand cDNA using reverse transcriptase and random primers, followed by second strand synthesis using DNA polymerase I and RNAse H. The cDNA fragments were then subject to end-repair followed by addition of a single ‘A’ base and ligation of adapters. The products were indexed individually, purified and enriched by PCR to create the final cDNA library. The generated library was validated and quantified using Kapa Library Quantification kit (Kapabiosystem). Six individually indexed cDNA libraries of equal amounts were pooled for clustering in cBot system (Illumina). Libraries were clustered onto a flow cell using Illumina’s TruSeq SR Cluster Kit v3, and sequenced for 50 cycles using TruSeq SBS kit on Illumina HiSeq system.
FASTQ files containing 50 bp single-end RNA-Seq reads were mapped to the Aspergillus fumigatus genome sequence (taxid:330879) by TopHat . Transcript assembly and abundance estimation were performed by Cufflinks .
Reads corresponding to 233 genes of interest were filtered and the coverage of each nucleotide position was counted using a semi-automated method in order to ensure accuracy of analysis. Coverage plots for each of the 233 genes under two conditions were plotted using Matlab®.
Analysis of mRNA expression by northern blot analysis and qPCR
RNA samples were fractionated by formaldehyde gel electrophoresis, and visualized by SYBR green staining. The RNA was then transferred to BioBond nylon membranes (Sigma) and hybridized to a 32P-labeled DNA probe as previously described . Probes specific for the A. fumigatus erg1, yvc1 and bipA genes were PCR amplified from genomic DNA using the following primers: erg1: 5′- CGTCAGTGTTGTTGAGAC-3′ and 5′- GAAGGTCGAGAGCTGCTTC-3′; yvc1: 5′- CAATGCTGTGGACGAGTACATG-3′ and 5′ - GTGCTCCTCTGTATCCTTCTTC-3′; bipA: 5′- GTCTGATTGGACGCAAGTTC-3′ and 5′- ATCTGGGAAGACAGAGTACG-3′. Hybridization intensities were quantified by phosphorimager analysis using Image Lab software.
For qPCR analysis one μg of RNA from pooled fractions corresponding to fraction-U or fraction-W was reverse-transcribed with M-MuLV reverse transcriptase (NEB) using oligo (dT)18 and 18S rRNA primers (primer 713-TGAGCCGATAGTCCCCCTAA and primer 714-GACTCAACACGGGGAAACTC). The qPCR was performed using the iTaq™ universal SYBR® green supermix (Bio-Rad) according to the manufacturer’s protocol. The melting curve was monitored to verify specificity of the amplification reaction. Controls reactions in the absence of reverse transcriptase were used verify the absence of DNA contamination. The 18S rRNA present within fraction-U or fraction-W was used as an endogenous control to derive a ∆Ct value for each fraction. A translational efficiency ratio (W/U) was derived by subtracting ∆Ct of fraction-W from that of fraction-U, representing ∆∆Ct. Change in W/U ratios upon treatment with DTT or TM was then plotted using 2-∆∆Ct of untreated samples as the reference. Primers used for qRT-PCR are as follows: β-tubulin (AfuA_1g10910), primer 554-CACGGATCTTGGAGATC and primer 562-ACAACTTCGTCTTCGGCCAG; squalene monooxygenase erg1 (AfuA_5g07780), primer 810-AGCTGCGATCTATGCCGAATTCCT and primer 799-TCCCAGTTGGAAGTAACGGAAGCA; vacuolar protein sorting 55 superfamily vps55 (AFUA_6G04780), primer 804-GCGCTCTCCTTTGTTCTTGCCATT and primer 805-AAGACCTCCGAGGATGGACATGAT; bZIP transcription factor jlbA/IDI-4 (AFUA_5G01650), primer 813-TTGATGTGAACGACTCTCTGCCGT and primer 814-TAGCTTCGACACCCGCATCTTCAA. The data were compared by Student’s t-test and a p < 0.05 was considered significant (indicated by the asterisk, Additional file 1).
The microarray and RNA-seq data sets reported in this article are available in the ArrayExpress database (microarray accession E-MTAB-2027, RNA-seq accession ERP004296).
Unfolded protein response
Kyoto Encyclopedia of Genes and Genomes
Transient receptor potential
Yeast extract/glucose medium.
Supported by National Institutes of Health grant R21AI075237 and R01AI072297 to DSA. The authors thank Jay Card for photography and illustration assistance.
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