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Regulation of the transcription factor STE12 by the RNA polymerase II associated CDK SRB10

University of British Columbia

The transcription factor Ste12 is a heavily phosphorylated protein that activates genes required for mating, as well as filamentous growth of the yeast Saccharomyces cerevisiae. Both processes require activation by an upstream MAPK cascade, and this model system is representative of signal responsive transcription in higher organisms. I have identified a signaling pathway that utilizes the RNA polymerase holoenzyme machinery as a means to simultaneously transmit information to potentially all gene specific transcription factors. In my thesis research I have demonstrated that Srb10, a CDK in the mediator subcomplex of the RNA Pol II holoenzyme, directly phosphorylates Ste12, and targets the protein for ubiquitin dependent proteolysis. Using tryptic phosphopeptide mapping I identified two serine residues on Ste12 that are phosphorylated by Srb10 in vitro and required for phosphorylation by Srb10 in vivo. These phosphorylations, at S261 and S451, have a negative effect on Ste12 activity, and filamentous growth as mutation of these sites to alanine results in elevated Ste12 dependent transcription, and hyperfilamentous growth. The ability of Srb10 to phosphorylate Ste12 is dependent on fermentable carbon, an observation that is consistent with the model that Srb10 activity is sensitive to environmental and nutritional stress. Together these observations support a model whereby Srb10 limits Ste12 activity and filamentous response in rich media. Starvation antagonizes Srb10, causing stabilization of Ste12, facilitating induction of filamentous response genes. Because other transcription factors are also regulated by Srb10 phosphorylation, this pathway appears to coordinate expression of diverse gene sets with the growth potential of the cell.

Thesis/Dissertation

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2009-11-30

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http://hdl.handle.net/2429/16005

Biochemistry and Molecular Biology

University of British Columbia

UBCV

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