The yeast heat shock response is induced by conversion of cells to spheroplasts and by potent transcriptional inhibitors.
We report here that procedures commonly used to measure transcription and mRNA decay rates in Saccharomyces cerevisiae induce the heat shock response. First, conversion of cells to spheroplasts with lyticase, a prerequisite for nuclear runoff transcription, induces the expression of HSP70 and HSP90 heat shock genes. The transcript levels of the non-heat-shock gene ACT1 are slightly depressed, consistent with the general yeast stress response. Second, the DNA intercalator, 1,10-phenanthroline, widely employed as a general transcriptional inhibitor in S. cerevisiae, enhances the mRNA abundance of certain heat shock genes (HSP82, SSA1-SSA2) although not of others (HSC82, SSA4, HSP26). Third, the antibiotic thiolutin, previously demonstrated to inhibit all three yeast RNA polymerases both in vivo and in vitro, increases the RNA levels of HSP82 5- to 10-fold, those of SSA4 greater than 25-fold, and those of HSP26 greater than 50-fold under conditions in which transcription of non-heat-shock genes is blocked. By using an episomal HSP82-lacZ fusion gene, we present evidence that lyticase and thiolutin induce heat shock gene expression at the level of transcription, whereas phenanthroline acts at a subsequent step, likely through message stabilization. We conclude that, because of the exquisite sensitivity of the yeast heat shock response, procedures designed to measure the rate of gene transcription or mRNA turnover can themselves impact upon each process.
Department of Biochemistry and Molecular Biology, Louisiana State University Medical Center, Shreveport 71130.
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D000935:Antifungal Agents / Q000494:pharmacology*; D015152:Blotting, Northern; D002467:Cell Nucleus / Q000502:physiology; D005800:Genes, Fungal; D006360:Heat-Shock Proteins / Q000235:genetics*; D006358:Hot Temperature; D007700:Kinetics; D010618:Phenanthrolines / Q000494:pharmacology*; D011760:Pyrrolidinones / Q000494:pharmacology; D012331:RNA, Fungal / Q000235:genetics / Q000302:isolation & purification; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000235:genetics / Q000502:physiology*; D013104:Spheroplasts / Q000502:physiology; D014158:Transcription, Genetic / Q000187:drug effects*
D000935:Antifungal Agents; D006360:Heat-Shock Proteins; D010618:Phenanthrolines; D011760:Pyrrolidinones; D012331:RNA, Fungal; D012333:RNA, Messenger; C006361:acetopyrrothine; C025205:1,10-phenanthroline