Ssn6 (Cyc8) is a component of the yeast general corepressor Ssn6-Tup1 that inhibits the transcription of many diversely regulated genes. The corepressor does not interact directly with DNA but is recruited to different promoters through interactions with distinct pathway-specific, DNA-binding repressor proteins. Using yeast two-hybrid and GST chromatography interaction experiments, we have determined that Sfl1, a novel repressor protein, interacts directly with Ssn6, and in vivo repression data suggest that Sfl1 inhibits transcription by recruiting Ssn6-Tup1 via a specific domain in the Sfl1 protein. Sin4 and Srb10, components of specific RNA polymerase II sub-complexes that are required for Ssn6-Tup1 repression activity, are found to be required for Sfl1 repression function. These results indicate a possible mechanism for Sfl1-mediated repression via Ssn6-Tup1 and specific subunits of the RNA polymerase II holoenzyme. Electrophoretic mobility shift and chromatin immuno-precipitation assays demonstrate that Sfl1 is present at the promoters of three Ssn6-Tup1-repressible genes; namely, FLO11, HSP26, and SUC2. Sfl1 is known to interact with Tpk2, a cAMP-dependent protein kinase that negatively regulates Sfl1 function. Consistently, we show that phosphorylation by protein kinase A inhibits Sfl1 DNA binding in vitro, and that a tpk2Delta mutation increases the levels of Sfl1 protein associated with specific promoter elements in vivo. These data indicate a possible mechanism for regulating Sfl1-mediated repression through modulation of DNA binding by cAMP-dependent protein kinase-dependent phosphorylation. Taken together with previous data, these new observations suggest a link between cAMP signaling and Ssn6-Tup1-mediated transcriptional repression.
|Evidence ID||Analyze ID||Interactor||Interactor Systematic Name||Interactor||Interactor Systematic Name||Type||Assay||Annotation||Action||Modification||Phenotype||Source||Reference||Note|
|Evidence ID||Analyze ID||Gene||Gene Systematic Name||Gene Ontology Term||Gene Ontology Term ID||Qualifier||Aspect||Method||Evidence||Source||Assigned On||Annotation Extension||Reference|
|Evidence ID||Analyze ID||Gene||Gene Systematic Name||Phenotype||Experiment Type||Experiment Type Category||Mutant Information||Strain Background||Chemical||Details||Reference|
|Evidence ID||Analyze ID||Regulator||Regulator Systematic Name||Target||Target Systematic Name||Experiment||Assay||Construct||Conditions||Strain Background||Reference|