MULTIPLE REGIONS OF TBP PARTICIPATE IN THE RESPONSE TO TRANSCRIPTIONAL ACTIVATORS IN-VIVO

We used mutant yeast and human TBP molecules with an altered DNA-binding specificity to examine the role of TBP in transcriptional activation in vivo. Fire show that yeast TBP is functionally equivalent to human TBP for response to numerous transcriptional activators in human cells, including those that do not function in yeast. Despite the extensive conservation of TBP, its ability to respond to transcriptional activators in vivo is curiously resistant to clustered sets of alanine substitution mutations in different regions of the protein, including those that disrupt DNA binding and basal transcription in vitro. Combined sets of these mutations, however, can attenuate the in vivo activity of TBP and can differentially affect response to different activation domains. Although the activity of TBP mutants in vivo did not correlate with DNA binding or basal transcription in vitro, it did correlate with binding in vitro to the largest subunit of TFIID, hTAF(II)250. Together, these data suggest that TBP utilizes multiple interactions across its surface to respond to RNA polymerase II transcriptional activators in vivo; some of these interactions appear to involve recruitment of TBP into TFIID, whereas others are involved in response to specific types of transcriptional activators.