THE CYC8 (SSN6)-TUP1 COREPRESSOR COMPLEX IS COMPOSED OF ONE CYC8 AND 4 TUP1 SUBUNITS

The Cyc8 (Ssn6)-Tup1 corepressor complex is required for repression in several important regulatory systems in yeast cells, including glucos e repression and mating type, Cyc8-Tup1 is recruited to target genes b y interaction with diverse repressor proteins that bind directly to DN A, Since the complex has a large apparent molecular mass of 1,200 kDa on nondenaturing gels (F. E. Williams, U. Varanasi, and R. J. Trumbly, Mel. Cell. Biol, 11:3307-3316, 1991), we used a variety of approaches to determine its actual subunit composition, Immunoprecipitation of e pitope-tagged complex and reconstitution of the complex from in vitro- translated proteins demonstrated that only the Cyc8 and Tup1 proteins were present in the complex, Hydrodynamic properties showed that these proteins have unusually large Stokes radii, low sedimentation coeffic ients, and high frictional ratios, all characteristic of asymmetry whi ch partly accounts for the apparent high molecular weight, Calculation of native molecular weights from these properties indicated that the Cyc8-Tup1 complex is composed of one Cyc8 subunit and four Tup subunit s, This composition was confirmed by reconstitution of the complex fro m Cyc8 and Tup1 expressed in vitro and analysis by one- and two-dimens ional gel electrophoresis.