Promoter Interference in a Bacteriophage Lambda Control Region: Effects of a Range of Interpromoter Distances

Abstract

The p R and p RM promoters of bacteriophage lambda direct transcription in divergent directions from start sites separated by 83 phosphodiester bonds. We had previously shown that the presence of an RNA polymerase at p R interfered with open complex formation at p RM and that this effect was alleviated by the deletion of 10 bp between the two promoters. Here we present a detailed characterization of the dependence of the interference on the interpromoter distance. It was found that the reduced interference between the two promoters is unique to the 10-bp deletion. The relief of interference was demonstrated to be due to the facilitation of a step subsequent to RNA polymerase binding to the p RM promoter. A model to explain these observations is proposed. A search of known Escherichia coli promoters identified three pairs of divergent promoters with similar separations to those investigated here.In the rightward control region of bacteriophage lambda, transcription is initiated in divergent directions from two promoters, p R and p RM , that have start sites separated by 83 phosphodiester bonds (pdb; we are using this designation to avoid ambiguity in the representation of the distance between start sites). These two promoters are among those responsible for implementing the decision as to whether viral development will proceed along the lytic or lysogenic pathways (27). The p R promoter has greater similarity to the promoter consensus sequence than the p RM promoter (27). As a consequence, open complex formation at p R is accomplished in seconds but under the same conditions requires tens of minutes at p RM (15,27,34). Therefore, for the wild-type control region, in vitro RNA polymerase (RNAP)-p RM interactions occur almost exclusively in the context of another RNAP already bound to p R . It has been previously shown that this p R -bound RNAP interferes with open complex formation at p RM (16,17,21,34,37). The effect is not exerted at the initial binding of RNAP to the promoter but rather at a subsequent step (16, 34) that is likely a conformational change in the RNAP (9). Eventually, open complexes do form at p RM and coexist with those at p R (16,25). The converse of the situation described above has also been shown: when p R has been weakened due to base substitutions, its ability to form open complexes is affected by the presence of p RM on the same DNA fragment (11).Only 13 pdb separate the start site-distal edges of the Ϫ35 regions of the p R and p RM promoters. Given such a short interpromoter distance, it was suggested that the p R -bound RNAP was slowing open complex formation at p RM because of steric hindrance. Consistent with this notion, deletion of 1 bp between the Ϫ35 regions was found to further reduce the rate of open complex formation at p RM (40). However, it has also been shown that when the distance between the Ϫ35 regions of the promoters is shortened by the deletion of 10 bp (one turn of the DNA helix), unexpectedly the inhibition of open complex formation at p RM is greatly diminished (21)...

A Mathematical Model for Transcriptional Interference by RNA Polymerase Traffic in Escherichia coli

“…We also plan to adapt the model to account for promoter competition, where the promoters are so close that their SDCs overlap. 27,28 We expect our model to be directly transferable to other eubacterial systems. Some aspects, particularly our treatment of interference by collisions, might also be applied to eukaryotes.…”

Regulatory Cross-talk in the Double par Locus of Plasmid pB171

“…In most cases, transcriptional interference between divergent promoters is positive (19,20), and only few examples of negative interference are known (21). The deletion analysis presented in Fig.…”

Sigma70 Promoters in Escherichia coli: Specific Transcription in Dense Regions of Overlapping Promoter-like Signals

“…Interference among closely located promoters has been documented for some divergent promoters. 37 It has been shown experimentally that additional RNAP binding sites occur in the regions near functional promoters. 33,34,38 These promoter-like signals are embedded within A/T-rich sequences, perhaps to facilitate the localized melting required for transcription initiation process.…”