On the role of the Escherichia coli RNA polymerase σ70 region 4.2 and α-subunit C-terminal domains in promoter complex formation on the extended-10 galP1 promoter

Bacterial promoters of the extended -10 class contain a single consensus element, and the DNA sequence upstream of this element is not critical for promoter activity. Open promoter complexes can be formed on an extended -10 Escherichia coli galP1 promoter at temperatures as low as 6degreesC, when complexes on most promoters are closed. Here, we studied the contribution of upstream contacts to promoter complex formation using galP1 and its derivatives lacking the extended -10 motif and/or containing the -35 promoter consensus element. A panel of E. coli RNA polymerase holoenzymes containing two, one, or no alpha-subunit C-terminal domains (alphaCTD) and either wild-type sigma(70) subunit or sigma(70) lacking region 4.2 was assembled and tested for promoter complex formation. At 37degreesC, alphaCTD and sigma(70) region 4.2 were individually dispensable for promoter complex formation on galP1 derivatives with extended -10 motif. However, no promoter complexes formed when both alphaCTD and sigma(70) region 4.2 were absent. Thus, in the context of an extended -10 promoter, alphaCTD and sigma(70) region 4.2 interactions with upstream DNA can functionally substitute for each other. In contrast, at low temperature, alphaCTD and sigma(70) region 4.2 interactions with upstream DNA were found to be functionally distinct, for sigma(70) region 4.2 but not alphaCTD was required for open promoter complex formation on galP1 derivatives with extended -10 motif. We propose a model involving sigma(70) region 4.2 interaction with the beta flap domain that explains these observations.