Binding of Drosophila heat-shock gene transcription factor to the hsp 70 promoter. Evidence for symmetric and dynamic interactions.

A Drosophila heat-shock gene transcription factor (HSTF) has been shown to bind to three domains upstream from the TATA homology on a hsp 70 gene. The domain closest to the TATA homology consists of two contiguous binding sites with different binding affinities. Occupancy of the TATA homology proximal site (site 1) coordinates HSTF binding to the neighboring site (site 2) in a cooperative manner (Topol, J., Ruden, D. M., and Parker, C. S. (1985) Cell 42, 527-537). We have used alkylation interference and protection experiments to determine which residues within the binding sites are closely contacted by the HSTF. The contacts inferred from these studies included the residues present in the consensus sequence found in all HSTF binding sites and exhibit rotational symmetry, suggestive of a multimeric HSTF. By employing a gel electrophoresis separation technique we were able to resolve two protein-DNA complexes consisting of site 1 occupancy (complex A) and sites 1 and 2 occupancy (complex B). Analysis of these discrete species reveals that a subset of contacts within site 1 change upon HSTF binding to site 2, suggesting that a conformational change in the protein-DNA complex occurs. Implications for the activation of heat-shock gene transcription are discussed.