Antagonistic control of the Escherichia coli bgl promoter by FIS and CAP in vitro

Abstract

The wild-type Escherichia coli bgl promoter is silent in vivo but active in vitro. Silencing in vivo is directed by silencer sequences that flank the promoter, and requires nucleoid-associated protein H-NS and other unidentified cellular factors. Here we show that the DNA bending protein FIS is a repressor of the bgl promoter. Two FIS binding sites, centred at positions -52 and -27, overlap the CAP binding site and the -35 box respectively. FIS efficiently competes with CAP for binding to the wild-type promoter. However, FIS does not prevent binding of RNA polymerase. It interferes with the formation of a heparin-resistant complex and represses transcription initiation up to 40-fold. The presence of CAP has very little effect on the FIS-mediated repression of the wild-type bgl promoter in vitro. However, when a bgl promoter allele was tested that carries an improved CAP binding site (which leads to activation in vivo) CAP effectively counteracted repression by FIS in vitro. These results suggest that FIS contributes to silencing of the wild-type bgl promoter in vivo, presumably in the early exponential phase when FIS is predominantly expressed.