GalR-mediated repression and activation of hybrid lacUV5 promoter: differential contacts with RNA polymerase.

Abstract

The GalR repressor regulates expression of genes of the gal regulon in Escherichia coli. We studied the regulatory effect of GalR in vitro on a heterologous promoter, lacUV5, by placing the GalR-binding site, OE, at different locations upstream of this promoter. Despite the fact that the lacUV5 promoter is transcribed efficiently by RNA polymerase (RNP) alone, GalR modulated transcription from many of the PlacUV5 variants. Depending on the location of OE and the neighboring DNA sequence, GalR repressed, activated or had no effect on the promoter. Both repression and activation involved formation of GalR-RNP-DNA ternary complexes and required an intact c-domain of the alpha subunit of the holoenzyme. These results support the differential contact model of a regulator action, in which a regulator differentially binds to, and lowers the energy of, intermediates of transcription initiation either to hinder or to facilitate a step of initiation. The nature of the contacts depends upon the context, i.e. the geometry of the ternary complex. The observed repression and activation effect of GalR on a heterologous promoter also underscores the point that a regulator is not a dedicated protein for repression or for activation.