TAU-91, AN ESSENTIAL SUBUNIT OF YEAST TRANSCRIPTION FACTOR-IIIC, COOPERATES WITH TAU-138 IN DNA-BINDING

Transcription factor IIIC (TFIIIC) (or tau) is a large multisubunit an d multifunctional factor required for transcription of all class III g enes in Saccharomyces cerevisiae. It is responsible for promoter recog nition and TFIIIB assembly. We report here the cloning and characteriz ation of TFC6, an essential gene encoding the 91-kDa polypeptide, tau 91, present in affinity-purified TFIIIC. tau 91 has a predicted molecu lar mass of 74 kDa. It harbors a central cluster of His and Cys residu es and has basic and acidic amino acid regions, but it shows no specif ic similarity to known proteins or predicted open reading frames. The TFIIIC subunit status of tau 91 was established by the following bioch emical and genetic evidence. Antibodies to tau 91 bound TFIIIC-DNA com plexes in gel shift assays; in vivo, a B block deficient U6 RNA gene ( SNR6) harboring GALA binding sites was reactivated by fusing the GAL4 DNA binding domain to tau 91; and a point mutation in TFC6 (tau 91-E33 0K) was found to suppress the thermosensitive phenotype of a tfc3-G349 E mutant affected in the B block binding subunit (tau 138). The suppre ssor mutation alleviated the DNA binding and transcription defects of mutant TFIIIC in vitro. These results indicated that tau 91 cooperates with tau 138 for DNA binding. Recombinant tau 91 by itself did not in teract with a tRNA gene, although it showed a strong affinity for sing le-stranded DNA.