STRUCTURAL ANALYSIS OF THE TBP/TFIIA/TATA COMPLEX. J. H. Geiger, S. Hahn, P. B. Sigler, Yale University, New Haven, CT 06510

The goal of this project is to investigate structure-function relationships of eukaryotic transcription initiation. For RNA polymerase II, there are a host of general transcription factors that must associate at the promoter with the polymerase before transcription initiation begins. Some are required for the initiation of all genes (TFIID, TFIIB) in vivo and in vitro, while some are required on a subset of promoters, or serve to activate transcription, but are not necessary for basal levels of in vitro transcription. TFIIA falls in the latter category. An essential gene in yeast, TFIIA counteracts the effects of transcriptional repressors and may be involved in the activity of transcriptional activators. It increases the affinity of TBP to the TATA box by three orders of magnitude, and extends the TBP footprint on DNA by seven base pairs in the upstream direction.

We have crystallized and solved the structure of a complex of TBP/TFIIA/TATA to 3.0 angstroms. We are using this structure to elucidate the stereochemical determinants of this assembly and to give insight into the interaction of this complex with the rest of the basal machinery and its possible interaction with activators and repressors of transcription initiation.