Regulation of Yeast Transcription Factors Sko1 and Cst6 by Sumoylation

Sumoylation is a post-translational modification that plays an essential role in cellular processes, including transcriptional regulation. Transcription factors represent one of the largest groups of proteins that are modified by the SUMO (Small Ubiquitin-like Modifier) protein. In this study, we focused on finding roles for sumoylation in regulating two gene-specific bZIP transcription factors, Sko1 and Cst6, in Saccharomyces cerevisiae. Sko1 plays a unique bifunctional role in regulating transcription: it is a repressor during normal growth, by interacting with co-repressor complexes, and an activator during osmotic stress, via interaction with Hog1 Kinase. We show that Sko1 is poly-sumoylated at Lys 567 but its sumoylation is not regulated by stress. Along with sumoylation, Sko1 also undergoes phosphorylation, by PKA and Hog1, and our experiments show that these two modifications are not interdependent. We find that DNA binding is a requirement for Sko1 sumoylation and genome-wide chromatin immunoprecipitation (ChIP-seq) analysis shows that Sko1 sumoylation controls the occupancy level of Sko1 on target promoters and is involved in preventing Sko1 from binding to non-target promoters. Moreover, blocking sumoylation attenuated the interaction between Sko1 and Hog1 on target promoters.

Cst6, on the other hand, is required for survival during ethanol stress and has roles in the utilization of carbon sources other than glucose. In this study, we show that Cst6 is multi-sumoylated at Lys residues 139, 461 and 547. The level of Cst6 sumoylation increases in ethanol and oxidative stress conditions, but decreases if ethanol is used as the sole carbon source. Unlike Sko1, protein levels of SUMO-deficient Cst6 were moderately reduced compared to the wild-type form, implying that sumoylation promotes Cst6 stability. ChIP experiments suggests that sumoylation is important for the timely recruitment of Cst6 to its target promoters. In addition, we provide evidence that Cst6 sumoylation reduces the expression of some target genes, during non-stress and ethanol stress conditions. Taken together, our studies suggest that the specific effects of sumoylation in regulating transcription factors are target specific. Nevertheless, SUMO plays a general role by controlling the transcription factor-DNA interaction to maintain proper gene expression.