Abstract
Organisms respond to environmental stress by adopting changes in gene expression at the transcriptional level. Rpb4, a nonessential subunit of the core RNA polymerase II has been proposed to play a role in non-stress-specific transcription and in the regulation of stress response in yeast. We find that in addition to the temperature sensitivity of the null mutant of Rpb4, diploid null mutants are also compromised in sporulation and show morphological changes associated with nitrogen starvation. Using whole genome expression analysis, we report here the effects of Rpb4 on expression of genes during normal growth and following heat shock and nutritional starvation. Our analysis shows that Rpb4 affects expression of a small yet significant fraction of the genome in both stress and normal conditions. We found that genes involved in galactose metabolism were dependent on the presence of Rpb4 irrespective of the environmental condition. Rpb4 was also found to affect the expression of several other genes specifically in conditions of nutritional starvation. The general defect in the absence of Rpb4 is in the expression of metabolic genes, especially those involved in carbon metabolism and energy generation. We report that various stresses are affected by RPB4 and that on overexpression the stress-specific activators can partially rescue the corresponding defects.
Highlights
The survival of a cell depends on its ability to respond rapidly to environmental changes
We find that in addition to the temperature sensitivity of the null mutant of Rpb4, diploid null mutants are compromised in sporulation and show morphological changes associated with nitrogen starvation
Using whole genome expression analysis, we report here the effects of Rpb4 on expression of genes during normal growth and following heat shock and nutritional starvation
Summary
Strains and Plasmids—The rpb4⌬ and wild type strains (SY10-1: Mat a, his3⌬-200, ura, leu 112, lys, rpb4⌬::HIS3/pPS2; SY10-2: Mat a, his3⌬-200, ura, leu 112, lys, rpb4⌬::HIS3/pNS114) have been described before [8]. PHD1 as well as IME4 open reading frames were PCR-amplified and expressed in yeast from multicopy plas-. Sporulation—The yeast cultures were grown until mid-log phase at 25 °C in YEPD [13]. The cells were harvested after 12 h of incubation in sporulation medium for RNA isolation. The cells were harvested, washed with sterile water, and transferred to sterile SLAD medium (0.67% yeast nitrogen base without ammonium sulfate and amino acids, 0.05 mM ammonium sulfate, 2% glucose). The cells were harvested after 12 h of incubation in SLAD medium for RNA isolation. Data from published microarray experiments used to compare with data generated in our experiments was retrieved using yMGV (Yeast Microarray Global Viewer) [19]
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