FUNCTIONAL INTERACTION OF THE BOVINE PAPILLOMAVIRUS E2 TRANSACTIVATION DOMAIN WITH TFIIB

Induction of gene expression by the papillomavirus E2 protein requires its similar to 220-amino-acid amino-terminal transactivation domain ( TAD) to interact with cellular factors that lead to formation of an ac tivated RNA polymerase complex These interaction partners have yet to be identified and characterized. The E2 protein localizes the transcri ption complex to the target promoter through its carboxy-terminal sequ ence-specific DNA binding domain, This domain has been reported to bin d the basal transcription factors TATA-binding protein and TFIIB. We p resent evidence establishing a direct interaction between amino acids 74 to 134 of the E2 TAD and TFIIB. Within this region, the E2 point mu tant N127Y was partially defective and W99C was completely defective f or TFIIB binding in vitro, and these mutants displayed reduced or no t ranscriptional activity. respectively, upon transfection into C33A cel ls. Overexpression of TFIIB specifically restored transactivation by N 127Y to close to wild-type levels, while W99C remained inactive. To fu rther demonstrate the functional interaction of TFIIB with the wild-ty pe E2 TAD, this region was fused to a bacterial DNA binding domain (Le xA:E2:1-216). Upon transfection with increasing amounts of LexA:E2:1-2 16, there was reduction of its transcriptional activity, a phenomenon thought to result from titration of limiting factors, or squelching. S quelching of LexA:E2:1-216, or the wild-type E2 activator, was partial ly relieved by overexpression of TFIIB. We conclude that a specific re gion of the E2 TAD functionally interacts with TFIIB.