ADR1-Mediated Transcriptional Activation Requires the Presence of an Intact TFIID Complex

ABSTRACT

The yeast transcriptional activator ADR1, which is required forADH2 and other genes’ expression, contains four transactivation domains (TADs). While previous studies have shown that these TADs act through GCN5 and ADA2, and presumably TFIIB, other factors are likely to be involved in ADR1 function. In this study, we addressed the question of whether TFIID is also required for ADR1 action. In vitro binding studies indicated that TADI of ADR1 was able to retain TAF_II90 from yeast extracts and TADII could retain TBP and TAF_II130/145. TADIV, however, was capable of retaining multiple TAF_IIs, suggesting that TADIV was binding TFIID from yeast whole-cell extracts. The ability of TADIV truncation derivatives to interact with TFIID correlated with their transcription activation potential in vivo. In addition, the ability of LexA-ADR1-TADIV to activate transcription in vivo was compromised by a mutation in TAF_II130/145. ADR1 was found to associate in vivo with TFIID in that immunoprecipitation of either TAF_II90 or TBP from yeast whole-cell extracts specifically coimmunoprecipitated ADR1. Most importantly, depletion of TAF_II90 from yeast cells dramatically reducedADH2 derepression. These results indicate that ADR1 physically associates with TFIID and that its ability to activate transcription requires an intact TFIID complex.

ACKNOWLEDGMENTS

We thank K. Struhl for providing strains and M. Green for antibodies and strains used in this project. The technical assistance of J. Farrell is also appreciated.

This research was supported by NIH grant GM41215, NSF grant MCB95-13412, Hatch project 291 to C.L.D., and NIH grant GM52461 to P.A.W. P.B.K. was partially supported by a Dissertation Fellowship Award from the University of New Hampshire.