MODULAR ORGANIZATION OF THE E2F1 ACTIVATION DOMAIN AND ITS INTERACTION WITH GENERAL TRANSCRIPTION FACTORS TBP AND TFIIH

The transcriptional activator E2F1 regulates the expression of genes a t the G1/S boundary. We have characterized interactions of the E2F1 ac tivation domain with two general transcription factors, the TATA-box b inding protein (TBP) and TFIIH. Two distinct binding sites on E2F1 wer e identified for TBP (amino acids 386-417 and 415-437) each of which s upported activation in mammalian cells when expressed as a fusion to a heterologous DNA-binding domain. Neither of these minimal activation domains independently bound TFIIH; rather, the TFIIH binding site of E 2F1 overlaps both domains. Loss of TFIIH-binding by E2F1 resulted in a 60-65% reduction in transactivation, suggesting that the E2F1/TFIIH i nteraction is important, but not essential, for transactivation. The r etinoblastoma protein (Rb) binds directly to E2F1 and represses E2F1-m ediated transactivation. We have demonstrated that recombinant Rb can compete with TBP and the p62 subunit of TFIIH for binding to immobiliz ed E2F1. A tumorigenic form of Rb deficient in repressing E2F1-mediate d transactivation is likewise deficient in displacing TBP from E2F1. W e propose that competition between Rb and both TBP and TFIIH for bindi ng to E2F1 is a mechanism by which Rb inhibits transactivation by E2F1 .