Bidirectional transcription in the mom promoter region of bacteriophage Mu

Abstract

Transcription of the Mu mom operon requires activation by the phage gene product, C, a site-specific DNA binding protein. Previous in vivo and in vitro footprinting studies showed that Escherichia coli RNA polymerase (Eσ 70=RNAP) bound the wild-type (wt) mom promoter (P mom ) region in the absence of C; this site, now designated momP2 (−11 to −64), is slightly upstream of, but overlapping with, momP1 (+16 to −49), the functional binding site for mom operon (rightward) transcription. The location/distribution of KMnO 4-sensitive sites on the two DNA strands suggested that RNAP bound at momP2 was in an open-complex, but that transcription was in the opposite direction. Here, we used both runoff transcription and reverse transcriptase-primer extension sequencing to provide direct evidence that in the absence of C protein, RNAP carries out leftward transcription from momP2 both in vitro and in vivo. In addition, the 5′ ends of these transcripts were mapped to the same upstream initiation site, −58G, relative to the initiation site of C-activated rightward transcription. We also present evidence that leftward transcription from momP2 requires RNAP recognition of an UP-element by the carboxyl-terminal domain of the α subunit.