Important Role In Thermotolerance Research Articles

Evolutionary Relationship Between Arabian Camel Breeds Based on Sequencing of Heat-Stress Genes under Egyptian Conditions

The Arabian camel (Camelus dromedarius), which is domesticated in semi-desert environments in Egypt, is one of the neglected animals in the scientific community. Although, it is the best model for studying acclimatization as it is well adapted to withstand severe drought and high temperatures and survive for long periods. Many physiological and behavioral aspects were found to play a role in such adaptations. At the molecular level, it is well known that heat shock genes play an important role in thermotolerance of camels. Therefore, this study was designed to investigate relationship among the most dominant breeds in Egypt (Fellahi, Maghrebi and Sudani) in order to determine the phylogenetic relationship based on sequences of two heat stress genes (HSP family B (small) member 9: HSPB9 and HSP 70 kDa 1B: HSPA1B). Both genes were isolated and partially sequenced from Fellahi, Maghrebi and Sudani Egyptian breeds. It has been deposited in the NCBI database with accession numbers MZ243318, MZ243319, MZ243320, MZ851990 and MZ851991. By studying the phylogenetic tree based on HSPB9 gene sequence, our results showed that Maghrebi and Sudani breeds are closely related whereas, Fellahi breed was found to be more related to the reference breed on database. Relying on HSPA1B gene sequence, it was found that Maghrebi breed is closely related to wild strain (Camelus ferus). Additionally, Fellahi breed was separated into distinct group with reference breed, which indicates strength of its relationship to reference origin.

Thermal Stress Has Minimal Effects on Bacterial Communities of Thermotolerant Symbiodinium Cultures

Algae in the dinoflagellate family Symbiodiniaceae are endocellular photosymbionts of corals and other cnidarians. This close relationship is disrupted when seawater temperature increases, causing coral bleaching eventually affecting entire coral reefs. Although the relationship between animal host and photosymbiont has been well-studied, little is known about the bacterial community associated with Symbiodiniaceae in culture. We compared the microbial communities of three isolates from different species of the genus Symbiodinium (formerly known as Symbiodinium clade A) with different ecophysiology, levels of interaction with the animal host, and thermal adaptations. Two species, Symbiodinium microadriaticum and Symbiodinium necroappettens, exhibit intermediate thermotolerance, with a decrease of both growth rate and photochemical efficiency with increased temperature. The third species, Symbiodinium pilosum, has high thermotolerance with no difference in growth rate or photochemical efficiency at 32°C. Microbial communities were characterized after 27 days of growth under control (26°C) and high temperature (32°C). Data shows stronger grouping of bacterial assemblages based on Symbiodinium species than temperature. Microbial communities did not group phylogenetically. We found a shared set of fifteen ASVs belonging to four genera and three families that remained in all three Symbiodiniaceae species. These included Labrenzia, Phycisphaeraceae (SM1A02), Roseovarius, and Muricauda, which are all commonly associated with corals and Symbiodiniaceae cultures. Few ASVs differed significantly by temperature within species. S. pilosum displayed significantly lower levels of microbial diversity and greater individual variability in community composition at 32°C compared to 26°C. These results suggest that bacteria associated or co-cultured with thermotolerant Symbiodinium might play an important role in thermotolerance. Further research on the functional metabolic pathways of these bacteria might hold the key to understanding Symbiodinium’s ability to tolerate thermal stress.

Heat shock protein 101 (HSP101) promotes flowering under nonstress conditions.

Heat shock proteins (HSPs) are stress-responsive proteins that are conserved across all organisms. Heat shock protein 101 (HSP101) has an important role in thermotolerance owing to its chaperone activity. However, if and how it functions in development under nonstress conditions is not yet known. By using physiological, molecular, and genetic methods, we investigated the role of HSP101 in the control of flowering in Arabidopsis (Arabidopsis thaliana (L.) Heynh.) under nonstress conditions. Knockout and overexpression of HSP101 cause late and early flowering, respectively. Late flowering can be restored by rescue of HSP101. HSP101 regulates the expression of genes involved in the six known flowering pathways; the most negatively regulated genes are FLOWERING LOCUS C (FLC) and SHORT VEGETATIVE PHASE (SVP); downstream integrators of the flowering pathways are positively regulated. The late-flowering phenotype of loss-of-HSP101 mutants is suppressed by both the mutations of FLC and SVP. The responses of flowering time to exogenous signals do not change in HSP101 mutants. HSP101 is also found in nonspecific regions according to subcellular localization. We found that HSP101 promotes flowering under nonstress conditions and that this promotion depends on FLC and SVP. Our data suggest that this promotion could occur through a multiple gene regulation mechanism.

Transcriptomic Analysis Reveals Important Roles of Lignin and Flavonoid Biosynthetic Pathways in Rice Thermotolerance During Reproductive Stage.

Rice is one of the major staple cereals in the world, but heat stress is increasingly threatening its yield. Analyzing the thermotolerance mechanism from new thermotolerant germplasms is very important for rice improvement. Here, physiological and transcriptome analyses were used to characterize the difference between two germplasms, heat-sensitive MH101 and heat-tolerant SDWG005. Two genotypes exhibited diverse heat responses in pollen viability, pollination characteristics, and antioxidant enzymatic activity in leaves and spikelets. Through cluster analysis, the global transcriptomic changes indicated that the ability of SDWG005 to maintain a steady-state balance of metabolic processes played an important role in thermotolerance. After analyses of gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment, we found that the thermotolerance mechanism in SDWG00 was associated with reprogramming the cellular activities, such as response to abiotic stress and metabolic reorganization. In contrast, the down-regulated genes in MH101 that appeared to be involved in DNA replication and DNA repair proofreading, could cause serious injury to reproductive development when exposed to high temperature during meiosis. Furthermore, we identified 77 and 11 differentially expressed genes (DEGs) involved in lignin and flavonoids biosynthetic pathways, respectively. Moreover, we found that more lignin deposition and flavonoids accumulation happened in SDWG005 than in MH101 under heat stress. The results indicated that lignin and flavonoid biosynthetic pathways might play important roles in rice heat resistance during meiosis.

Effect of high temperature stress on heat shock protein expression and antioxidant enzyme activity of two morphs of the mud crab Scylla paramamosain

The present study aimed to investigate the effect rapid temperature change from moderate temperature to high temperatures on heat shock protein (HSP) expression and antioxidant enzyme activities in mud crabs. Two mud crabs, one with one spine on the outer margin of the carpus of cheliped (Sp1) and another with two spines (Sp2), were acclimated at 25 °C and then transferred to a 33 °C environment, and HSP expression and antioxidant enzyme activity were assessed. HSP70 and HSP60 were markedly up-regulated in the gills and hepatopancreas of Sp1 and Sp2 after exposure to 35 °C. Exposure to 35 °C also significantly increased superoxide dismutase and catalase activity in the gills of Sp1 and Sp2, with transient changes in hepatopancreas. Apart from changes in antioxidant enzyme activities, HSPs were highly up-regulated after exposure to 37 °C, especially for HSP70. Gill HSP70 expression in Sp2 was 6.1 folds that of the control after 24 h of exposure to 37 °C, and 9.2 folds that of Sp1. Moreover, exposure to 37 °C further up-regulated HSP70 in the hepatopancreas of Sp1, compared to that in Sp2. Hence, HSPs play important roles in thermotolerance in S. paramamosain and Sp1 might have a stronger tolerance to hyperthermal stress than Sp2.

Polymorphisms in Hsp90ab1 gene and their association with heattolerance in Sahiwal and Karan Fries cows

Heat shock protein functions as a molecular chaperone and plays an important role in thermotolerance. The study was undertaken to identify single nucleotide polymorphisms (SNPs) of Hsp90ab1 gene by comparative sequencing and to analyze their association with thermo-physiological parameters, viz. respiration rate (RR), rectal temperature (RT) and heat tolerance coefficient (HTC) in different seasons, viz. winter, spring and summer, at probable extreme hours of the day in Sahiwal and Karan Fries cows. The RR, RT and HTC in summers were significantly (P£0.01) higher than the other two seasons. Polymorphism of Hsp90ab1 gene, evaluated by comparative sequencing revealed six SNPs, viz. T17871421C, C17871485del, C17872061T, T17872112C, T17872148G and A17872199C, two each located in exon 8, intron 10 and exon 11. SNP loci T17871421C and C17871485del were found to be monomorphic for allele C and Deletion (-) respectively in experimental population. Individuals with AA genotype showed significantly (P £0.01) lower RR than AC genotype. While, individuals with CT genotype recorded significantly (P £ 0.01) lower RT than CC genotype. Therefore, it is inferred that propagation of AA and CT genotypes may be an aid to selection and breeding programmes to enhance thermo-tolerance in dairy cattle.

Constitutive Expression of a Tomato Small Heat Shock Protein Gene LeHSP21 Improves Tolerance to High-Temperature Stress by Enhancing Antioxidation Capacity in Tobacco

It is well established that small heat shock proteins (sHSPs) play an important role in thermotolerance in various organisms due to their abundance and diversity. In the present study, a chloroplast small heat shock protein gene (LeHSP21) from tomato (Lycopersicon esculentum cv PKM-1) was constitutively expressed in tobacco (Nicotiana tabacum L. cv Wisconsin 38) plants via Agrobacterium-mediated transformation. When compared to wild-type control plants, transgenic tobacco plants constitutively expressing LeHSP21, driven by the cauliflower mosaic virus 35S promoter, exhibited improved tolerance to both high temperature and oxidative stress. Furthermore, when the seedlings were subjected to high temperature treatment, the activities of anti-oxidative enzymes and the content of proline were significantly higher in transgenic plants than those in the wild-type plants. Our results presented here demonstrate the feasibility of improving high temperature and oxidative stress tolerance in plants through the expression of LeHSP21 gene.

A SNP in the 3'-UTR of HSF1 in dairy cattle affects binding of target bta-miR-484.

The heat shock transcription factor 1 gene (HSF1) plays a key role in the heat stress response. We previously found a single nucleotide polymorphism (SNP) in the 3'-untranslated region (g.4693G>T) of HSF1 that was related to thermo tolerance in Chinese Holstein cattle through association analysis. However, it is not known whether other SNPs also affect thermo tolerance.In this study a novel SNP, g.1451G>T, was identified by DNA sequencing and genotyped using creating restriction site-polymerase chain reaction methodology. The g.1451G>T polymorphic site met Hardy-Weinberg equilibrium (P > 0.05). Association analysis demonstrated that this SNP had no effect on thermo tolerance traits in Holstein cattle. Findings of the study compared to the analysis of g.4693 G>T further indicated that g.4693 G>T may play an important role in thermo tolerance, although the mechanism is not clear. RNA hybrid and Targetscan prediction showed that the minimum free energy hybridization of bta-miR-484 with HSF1 3'-UTR was -31.9 kcal/mol and g.4693 G>T was in the seed sequence of bovine HSF1 that binds to bta-miR-484. Analysis by Luciferase assay indicated that HSF1 expression was directly targeted by bta-miR-484 in HEK 293T cells, and the Rluc/luc ratio of wildtype (GG) was lower than that of the mutant (TT) (P < 0.05). These results suggest that g.4693 G>T affects binding of HSF1 to bta-miR-484.

Transcriptome-Wide Characterization of Novel and Heat-Stress-Responsive microRNAs in Radish (Raphanus Sativus L.) Using Next-Generation Sequencing

microRNAs (miRNAs) are a class of single-stranded endogenous non-coding RNAs that play critical roles in plant growth, development, and environmental stress responses. Temperature is one of the major physical parameters disturbing cellular homeostasis and causing leaf etiolation in plants. Previous studies have reported that several conserved and novel miRNAs were responsive to heat stress in plants. However, the characterization of miRNAs responsive to heat stress in radish remains poorly understood. To better understand miRNAs and their target genes under heat stress, two small RNA libraries were constructed from heat-treated (Heat24) and heat-untreated (CK) radish roots. Using Solexa system, totally, 26 known and 19 novel miRNAs were identified as differentially expressed under heat stress. Expression patterns of a set of heat-responsive miRNAs were validated by quantitative real-time PCR (qRT-PCR). Furthermore, 422 sliced targets for 25 known miRNAs were identified by degradome sequencing technology, and most of the identified targets are involved in multiple biological processes including transcriptional regulation and response to biotic and abiotic stresses. Moreover, some miRNAs and their corresponding targets, which are related to the accumulation of heat stress transcription factors and heat shock proteins, played important roles in thermo-tolerance in radish. These findings could enhance the understanding of molecular mechanisms underlying miRNAs and their targets in regulating plant responses to heat stress.

Arabidopsis thaliana Phosphoinositide-Specific Phospholipase C Isoform 3 (AtPLC3) and AtPLC9 have an Additive Effect on Thermotolerance

The heat stress response is an important adaptation, enabling plants to survive challenging environmental conditions. Our previous work demonstrated that Arabidopsis thaliana Phosphoinositide-Specific Phospholipase C Isoform 9 (AtPLC9) plays an important role in thermotolerance. During prolonged heat treatment, mutants of AtPLC3 showed decreased heat resistance. We observed no obvious phenotypic differences between plc3 mutants and wild type (WT) seedlings under normal growth conditions, but after heat shock, the plc3 seedlings displayed a decline in thermotolerance compared with WT, and also showed a 40-50% decrease in survival rate and chlorophyll contents. Expression of AtPLC3 in plc3 mutants rescued the heat-sensitive phenotype; the AtPLC3-overexpressing lines also exhibited much higher heat resistance than WT and vector-only controls. The double mutants of plc3 and plc9 displayed increased sensitivity to heat stress, compared with either single mutant. In transgenic lines containing a AtPLC3:GUS promoter fusion, GUS staining showed that AtPLC3 expresses in all tissues, except anthers and young root tips. Using the Ca(2+)-sensitive fluorescent probe Fluo-3/AM and aequorin reconstitution, we showed that plc3 mutants show a reduction in the heat-induced Ca(2+) increase. The expression of HSP genes (HSP18.2, HSP25.3, HSP70-1 and HSP83) was down-regulated in plc3 mutants and up-regulated in AtPLC3-overexpressing lines after heat shock. These results indicated that AtPLC3 also plays a role in thermotolerance in Arabidopsis, and that AtPLC3 and AtPLC9 function additionally to each other.

Short-Term Increased Expression of the Heat Shock Protein 70 Family Members During Heat Shock is Positively Correlated with Basal Thermotolerance in the Midgut of Three Strains of the Silkworm,Bombyx mori

Heat shock proteins have been shown to play an essential role in protecting organisms from environmental stresses and can be divided into multiple families, including the heat shock protein 70 family. The family is composed of heat shock protein 70 (Hsp70) and heat shock cognate 70 (Hsc70). In the present study, to investigate whether the Hsp70 family plays an important role in the thermotolerance of Bombyx mori (silkworm), we analysed the expression of the Hsp70 family before and during heat shock in the midgut of three strains, including a thermotolerant strain cs-10, a thermosensitive strain cs-11, and cs-7, a thermotolerant near isogenic line of cs-11. qRT-PCR and Northern blot analyses revealed that, before heat shock, there was no significant difference in the expression of the selected four Hsp70 family members (Hsp70B, Hsp68, Hsc70-4 and Hsc70-3) among the three strains. Interestingly, after a heat shock of 2 h at 40 °C, increased expression of the selected four members was observed in all three strains, suggesting they are all inducible and involved in thermotolerance. Remarkably their expression, especially that of Hsp70B and Hsp68, was positively correlated with basal thermotolerance. After a heat shock of 8 h at 40 °C, there was no significant difference in the transcript level of Hsc70-3 between cs-10 and c-11. However, its transcript levels in both strains were still significantly higher than its pre-shock levels, indicating that Hsc70-3 may play an important role in long-term basal thermotolerance by working with other proteins yet to be identified. Unexpectedly, transcript levels of Hsp68, Hsc70-3 and Hsc-4 were found to be higher in cs-7 than in cs-10. A hypothesis is proposed to explain these results.

Involvement of Arabidopsis CPR5 in thermotolerance

The constitutive expressor of pathogenesis-related genes 5 (CPR5) plays a role in pathogen defence responses, programmed cell death, cell wall biogenesis, seed generation and senescence regulation in plants. Here, we investigated the functional characteristics of CPR5 to long-term heat stress in Arabidopsis with different genotypes: wild type (WT), cpr5 mutant and cpr5/CPR5 complementary transgenic plant. The cpr5 mutant showed increased susceptibility to long-term heat stress, displaying significant decreases in hypocotyl elongation, seedling and inflorescence survival, membrane integrity and photosystem II activity (Fv/Fm) during heat stress. However, the thermotolerance was recovered when cpr5 mutant was transformed with a CPR5 gene. H2O2 accumulation and lipid peroxidation were lower in cpr5/CPR5 plants and WT than in cpr5 mutants after exposure to 36 °C for 5 days. The alleviated oxidative damage was associated with increased activities of superoxide dismutase, catalase, and ascorbate peroxidase. Furthermore, the induced expression of HSP17.6A-CI, HSP101 and HSP70B under long-term heat stress was more substantial in cpr5/CPR5 plants and WT than in cpr5 mutants. These findings suggest that CPR5 plays an important role in thermotolerance of Arabidopsis by regulating the activities of antioxidant enzymes and the expressions of heat shock protein genes.

A mutation in THERMOSENSITIVE MALE STERILE 1, encoding a heat shock protein with DnaJ and PDI domains, leads to thermosensitive gametophytic male sterility in Arabidopsis

In most flowering plant species, pollination and fertilization occur during the hot summer, so plants must have evolved a mechanism that ensures normal growth of their pollen tubes at high temperatures. Despite its importance to plant reproduction, little is known about the molecular basis of thermotolerance in pollen tubes. Here we report the identification and characterization of a novel Arabidopsis gene, Thermosensitive Male Sterile 1 (TMS1), which plays an important role in thermotolerance of pollen tubes. TMS1 encodes a Hsp40-homologous protein with a DnaJ domain and an a_ERdj5_C domain found in protein disulfide isomerases (PDI). Purified TMS1 expressed in Escherichia coli (BL21 DE3) had the reductive activity of PDI. TMS1 was expressed in pollen grains, pollen tubes and other vegetative tissues, including leaves, stems and roots. Heat shock treatment at 37 degrees C increased its expression levels in growing pollen tubes as well as in vegetative tissues. A knockout mutation in TMS1 grown at 30 degrees C had greatly retarded pollen tube growth in the transmitting tract, resulting in a significant reduction in male fertility. Our study suggests that TMS1 is required for thermotolerance of pollen tubes in Arabidopsis, possibly by functioning as a co-molecular chaperone.

The Transcriptional Co-activator MBF1c Is a Key Regulator of Thermotolerance in Arabidopsis thaliana

The ability of an organism to acclimate to its environment is a key determinant in its global distribution and capacity to compete with other organisms. The heat stress response, a highly conserved environmental and developmental program in eukaryotic and prokaryotic organisms, is an important component of the acclimation response of plants. Previous studies have shown that heat shock transcription factors play an important role in thermotolerance in plants and other organisms, controlling the expression of different heat shock proteins and detoxifying enzymes. In contrast, although several other pathways, involving ethylene, salicylic acid (SA), and trehalose, were recently shown to play a central role in thermotolerance in plants, a key regulator of these responses was not identified. Here we report that the highly conserved transcriptional co-activator, MBF1c (multiprotein bridging factor 1c), is a key regulator of thermotolerance in Arabidopsis thaliana. MBF1c protein accumulates rapidly and is localized to nuclei during heat stress. MBF1c is required for thermotolerance and functions upstream to SA, trehalose, ethylene, and pathogenesis-related protein 1 during heat stress. In contrast, MBF1c is not required for the expression of transcripts encoding HSFA2 and different heat shock proteins. Interestingly, MBF1c interacts with TPS5 (trehalose phosphate synthase 5), which is also heat-inducible, and mutants deficient in TPS5 are thermosensitive. Our results provide evidence for the existence of a tightly coordinated heat stress-response network, involving trehalose-, SA-, and ethylene-signaling pathways, that is under the control of MBF1c.

The C-terminal Extension of Saccharomyces cerevisiae Hsp104 Plays a Role in Oligomer Assembly

The Saccharomyces cerevisiae protein Hsp104, a member of the Hsp100/Clp AAA+ family of ATPases, and its orthologues in plants (Hsp101) and bacteria (ClpB) function to disaggregate and refold thermally denatured proteins following heat shock and play important roles in thermotolerance. The primary sequences of fungal Hsp104's contain a largely acidic C-terminal extension not present in bacterial ClpB's. In this work, deletion mutants were used to determine the role this extension plays in Hsp104 structure and function. Elimination of the C-terminal tetrapeptide DDLD diminishes binding of the tetratricopeptide repeat domain cochaperone Cpr7 but is dispensable for Hsp104-mediated thermotolerance. The acidic region of the extension is also dispensable for thermotolerance and for the stimulation of Hsp104 ATPase activity by poly-l-lysine, but its truncation results in an oligomerization defect and reduced ATPase activity in vitro. Finally, sequence alignments reveal that the C-terminal extension contains a sequence (VLPNH) that is conserved in fungal Hsp104's but not in other orthologues. Hsp104 lacking the entire C-terminal extension including the VLPNH region does not assemble and has very low ATPase activity. In the presence of a molecular crowding agent the ATPase activities of mutants with longer truncations are partially restored possibly through enhanced oligomer formation. However, elimination of the whole C-terminal extension results in an Hsp104 molecule which is unable to assemble and becomes aggregation prone at high temperature, highlighting a novel structural role for this region.

Pleurotus sajor-caju HSP100 complements a thermotolerance defect inhsp104 mutantSaccharomyces cerevisiae

A putative Hsp100 gene was cloned from the fungus Pleurotus sajor-caju. mRNA expression studies demonstrated that this gene (designated PsHsp100) is highly induced by high temperature,induced less strongly by exposure to ethanol, and not induced by drought or salinity. Heat shock induction is detectable at 37 degrees C and reaches a maximum level at 42 degrees C. PsHsp100 mRNA levels sharply increased within 15 min of exposure to high temperature, and reached a maximum expression level at 2 h that was maintained for several hours. These results indicate that PsHsp100 could work at an early step in thermotolerance. To examine its function, PsHsp100 was transformed into a temperature-sensitive hsp104 deletion mutant Saccharomycetes cerivisiae strain to test the hypothesis that PsHSP100 is an protein that functions in thermotolerance. Overexpression of PsHSP100 complemented the thermotolerance defect of the hsp104 mutant yeast, allowing them being survive even at 50 degree C for 4 h. These results indicate that PsHSP100 protein is functional as an HSP100 in yeast and could play and important role in thermotolerance in P. sajor-caju.

An Arabidopsis heat shock protein complements a thermotolerance defect in yeast.

The heat shock protein Hsp104 of the yeast Saccharomyces cerevisiae plays a key role in promoting survival at extreme temperatures. We found that when diverse higher plant species are exposed to high temperatures they accumulate proteins that are antigenically related to Hsp104. We isolated a cDNA corresponding to one of these proteins from Arabidopsis. The protein, AtHSP101, is 43% identical to yeast Hsp104. DNA gel blot analysis indicated that AtHSP101 is encoded by a single- or low-copy number gene. AtHsp101 mRNA was undetectable in the absence of stress but accumulated to high levels during exposure to high temperatures. When AtHSP101 was expressed in yeast, it complemented the thermotolerance defect caused by a deletion of the HSP104 gene. The ability of AtHSP101 to protect yeast from severe heat stress strongly suggests that this HSP plays an important role in thermotolerance in higher plants.

Genetic evidence for a functional relationship between Hsp104 and Hsp70

The phenotypes of single Hsp104 and Hsp70 mutants of the budding yeast Saccharomyces cerevisiae provide no clue that these proteins are functionally related. Mutation of the HSP104 gene severely reduces the ability of cells to survive short exposures to extreme temperatures (thermotolerance) but has no effect on growth rates. On the other hand, mutations in the genes that encode Hsp70 proteins have significant effects on growth rates but do not reduce thermotolerance. The absence of a thermotolerance defect in S. cerevisiae Hsp70 mutants is puzzling, since the protein clearly plays an important role in thermotolerance in a variety of other organisms. In this report, examination of the phenotypes of combined Hsp104 and Hsp70 mutants uncovers similarities in the functions of Hsp104 and Hsp70 not previously apparent. In the absence of the Hsp104 protein, Hsp70 is very important for thermotolerance in S. cerevisiae, particularly at very early times after a temperature upshift. Similarly, Hsp104 plays a substantial role in vegetative growth under conditions of decreased Hsp70 protein levels. These results suggest a close functional relationship between Hsp104 and Hsp70.

Methionine-mediated lethality in yeast cells at elevated temperature.

Saccharomyces cerevisiae cells grown at 30 degrees C in minimal medium containing methionine lose viability upon transfer to 45 degrees C, whereas cells grown in the absence of methionine survive. Cellular levels of two intermediates in the sulfate assimilation pathway, adenosine 5'-phosphosulfate (APS) and adenosine 5'-phosphosulfate 3'-phosphate, are increased by a posttranslational mechanism after sudden elevation of temperature in yeast cultures grown in the absence of methionine. Yeast cells unable to synthesize APS because of repression by methionine or mutation of the MET3 gene do not survive the temperature shift. Thus, methionine-mediated lethality at elevated temperature is linked to the inability to synthesize APS. The results demonstrate that APS plays an important role in thermotolerance.