Kinetics of repression by the Tup1‐Ssn6 co‐repressor

The Tup1‐Ssn6 complex from budding yeast is one of the best studied co‐repressors and has served as a model for the study of similar co‐repressor complexes in higher eukaryotes. It is brought to promoters by DNA binding repressors, where it represses target genes that include those involved in DNA damage response, mating type genes, glucose metabolism and anaerobic stress. Recent studies have shown interaction of the co‐repressor with hypoacetylated histones, the RNA transcriptional machinery and histone deacetylases (HDACS) and have led to a model of how repression may be established. In this model, Tup1‐Ssn6 is recruited to the promoter by a sequence specific repressor, where it interacts with RNA polymerase and histones to establish repression of the gene. Tup1‐Ssn6 then recruits HDACs, which de‐acetylate the histones, reinforcing the repressive state. While the importance of these interactions with Tup1‐Ssn6 has been established, the timing of these events is still unknown. To analyze repression by Tup1‐Ssn6, we have used an inducible myc tagged repressor, myc ‐CRT, to follow the establishment of repression over time at the RNR2 gene. We have used chromatin immunoprecipitation to follow the recruitment of proteins to the promoter after induction. Two hours after induction of the myc ‐Crt, we can detect both Crt and Tup1 at the RNR2 promoter. At the same time histone acetylation and RNA polymerase occupancy decrease. As predicted in our model, repression is established by repressor mediated recruitment of Tup1‐Ssn6 to the promoter, concurrent with histone deacetylation and removal of RNA polymerase. Supported by : NIH 5 T32 HD07325‐19.