The role of upstream sequences in determining the strength of an rRNA promoter of E. coli.

In vitro transcription experiments were carried out with recombinant plasmids containing the promoters of the rrnB gene of Escherichia coli, and with deletion mutants lacking various lengths of the AT-rich sequence upstream from the P1 promoter of that gene. The main conclusions are as follows: The in vitro transcriptional activity of the P1 and P2 promoters of the rrnB gene are an order of magnitude higher on closed-circular (supercoiled) templates than on linear DNA; the strong P1 and P2 promoters are heparin-sensitive on linear templates, and on circular DNA only P2 is heparin-resistant; removal of the upstream AT-rich region did not decrease the apparent in vitro strength of the P1 promoter under standard conditions (50 mM KCl, high RNA polymerase/DNA ratio); at higher salt concentrations, or with a lower RNA polymerase/DNA ratio, the deletion mutants displayed much lower in vitro transcriptional activity than the wild-type, and the apparent weakening of the P1 promoter was roughly proportional to the length of the deleted AT-rich sequence. The implications of these findings for the possible in vivo role of the AT-rich region are discussed.