IDENTIFICATION OF TFIID COMPONENTS REQUIRED FOR TRANSCRIPTIONAL ACTIVATION BY UPSTREAM STIMULATORY FACTOR

A TATA box-binding initiation factor, TFIID, plays a central role in t he transcriptional regulation by activators. Using anti-TFIIDtau (a TA TA box-binding component of native TFIID) immunoaffinity chromatograph y, nine polypeptides (230, 110, 85, 62, 58, 42, 28, 22, and 21 kDa) we re identified as native Drosophila TFIID components that are tightly a ssociated with TFIIDtau. To verify the functional activity of the puri fied TFIID complex, template DNA and other transcription factors were reconstituted with purified TFIID bound to the antibody-Sepharose matr ix. Immobilized TFIID mediated not only basal transcription but transc riptional activation by upstream stimulatory factor (USF). On the othe r hand, recombinant TFIIDtau immobilized on the same antibody-Sepharos e matrix could not mediate activation by USF. These results suggest th at one or more of these additional polypeptides are required as functi onal TFIID subunits for activator-dependent transcription in conjuncti on with TFIIDtau. As further evidence of the relevance of the Drosophi la TFIID components identified in this analysis, including the previou sly unrecognized p230 (Dynlacht, B. D., Hoey, T., and Tjian, R. (1991) Cell 66, 563-576), protein blot analysis showed that TFIIDtau interac ts specifically and exclusively with p230. This suggests that p230 is an integral subunit of TFIID and that it may play a major role in teth ering other subunits to TFIIDtau.