A fork junction DNA-protein switch that controls promoter melting by the bacterial enhancer-dependent sigma factor.

Results of binding assays using DNA fork junction probes indicate that sigma 54 contains multiple determinants that regulate melting to allow RNA polymerase to remain in closed promoter complexes in order to respond to enhancers. Gel mobility shift studies indicate that the -12 promoter element and parts of sigma 54 act together to form a molecular switch that controls melting. The DNA sequences and the sigma 54 N-terminus help direct polymerase to the location within the -12 promoter element where melting will initiate. However, the fork junction that would lead to melting does not form, due to the action of an inhibitory DNA element. Such unregulated melting is inhibited further by the lack of availability of the single-strand binding elements, which are needed to spread opening from the junction to the transcription start site. Thus, in the absence of looping enhancer protein, proper regulation is maintained as the sigma 54 polymerase remains bound in an inactive state. These complex protein-DNA interactions allow the controls over protein recruitment and DNA melting to be separated, enhancing the diversity of accessible mechanisms of transcription regulation.