The short-lived Matalpha2 transcriptional repressor is protected from degradation in vivo by interactions with its corepressors Tup1 and Ssn6.
The Matalpha2 (alpha2) protein is a transcriptional repressor necessary for the proper expression of cell type-specific genes in Saccharomyces cerevisiae. Like many transcription factors, alpha2 is rapidly degraded in vivo by the ubiquitin-proteasome pathway. At least two different ubiquitin-dependent pathways target alpha2 for destruction, one of which recognizes the well-characterized Deg1 degradation determinant near the N terminus of the protein. Here we report that the alpha2 corepressors Tup1 and Ssn6 modify the in vivo degradation rate of alpha2. Tup1 modulates the metabolic stability of alpha2 by directly binding to the Deg1-containing region of the protein. TUP1 overexpression specifically stabilizes Deg1-containing proteins but not other substrates of the same ubiquitination enzymes that recognize Deg1. Point mutations in both alpha2 and Tup1 that compromise the alpha2-Tup1 binding interaction disrupt the ability of Tup1 to stabilize Deg1 proteins. The physical association between Tup1 and alpha2 competes with the ubiquitination machinery for access to the Deg1 signal. Finally, we observe that overproduction of both Tup1 and Ssn6, but not either alone, strongly stabilizes the endogenous alpha2 protein. From these results, we propose that the fraction of alpha2 found in active regulatory complexes with Tup1 and Ssn6 is spared from rapid proteolytic destruction and is stabilized relative to the uncomplexed pool of the protein.
Laney
Jeffrey D
JD
Department of Molecular Biophysics and Biochemistry, Brown University, Providence, RI 02912, USA.
Mobley
Erika F
EF
Hochstrasser
Mark
M
D004268:DNA-Binding Proteins / Q000378:metabolism*; D019751:Intramolecular Transferases; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D017354:Point Mutation; D011506:Proteins / Q000235:genetics / Q000378:metabolism*; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic; D025801:Ubiquitin / Q000378:metabolism
C065455:CYC8 protein, S cerevisiae; D004268:DNA-Binding Proteins; C062928:MATA2 protein, S cerevisiae; D009687:Nuclear Proteins; D011506:Proteins; D012097:Repressor Proteins; D029701:Saccharomyces cerevisiae Proteins; C066502:TUP1 protein, S cerevisiae; D025801:Ubiquitin; D019751:Intramolecular Transferases; C068903:DEG1 protein, S cerevisiae