THE ROLE OF TBP IN RDNA TRANSCRIPTION BY RNA-POLYMERASE-I IN SACCHAROMYCES-CEREVISIAE - TBP IS REQUIRED FOR UPSTREAM ACTIVATION FACTOR-DEPENDENT RECRUITMENT OF CORE FACTOR

Transcription of Saccharomyces cerevisiae rDNA by RNA polymerase I inv olves at least two transcription factors characterized previously: ups tream activation factor (UAF) consisting of Rrn5p, Rrn9p, Rrn10p, and two more uncharacterized proteins; and core factor (CF) consisting of Rrn6p, Rrn7p, and Rrn11p. UAF interacts directly with an upstream elem ent of the promoter and mediates its stimulatory function, and CE subs equently joins a stable preinitiation complex. The TATA-binding protei n (TBP) has been known to be involved in transcription by all three nu clear RNA polymerases. We found that TBP interacts specifically with b oth UAF and CF, the interaction with UAF being stronger than that with CF. Using extracts from a TBP (I143N) mutant, it was shown that TBP i s required for stimulation of transcription mediated by the upstream e lement, but not for basal transcription directed by a template without the upstream element. By template competition experiments, it was sho wn that TBP is required for UAF-dependent recruitment of CF to the rDN A promoter, explaining the TBP requirement for stimulatory activity of the upstream element. We also studied protein-protein interactions an d found specific interactions of TBP with Rrn6p and with Rrn9p both in vitro and in the yeast two-hybrid system in vivo. Thus, these two int eractions may be involved in the interactions of TBP with CF and UAF, respectively, contributing to the recruitment of CF to the rDNA promot er. Additionally, we observed an interaction between Rrn9p and Rrn7p b oth in vitro and in the two-hybrid system; thus, this interaction migh t also contribute to the recruitment of CF.