Transcriptional Activation of Agrobacterium tumefaciens Virulence Gene Promoters in Escherichia coli Requires the A. tumefaciens rpoA Gene, Encoding the Alpha Subunit of RNA Polymerase

摘要

The two-component regulatory system, composed of virAand virG, is indispensable for transcription of virulence genes within Agrobacterium tumefaciens. However,virA and virG are insufficient to activate transcription from virulence gene promoters within Escherichia coli cells, indicating a requirement for additional A. tumefaciens genes. In a search for these additional genes, we have identified the rpoA gene, encoding the α subunit of RNA polymerase (RNAP), which confers significant expression of avirB promoter (virBp)::lacZ fusion in E. coli in the presence of an active transcriptional regulatorvirG gene. We conducted in vitro transcription assays using either reconstituted E. coli RNAP or hybrid RNAP in which the α subunit was derived from A. tumefaciens. The two forms of RNAP were equally efficient in transcription from a ?70-dependent E. coli galP1 promoter; however, only the hybrid RNAP was able to transcribe virBpin a virG-dependent manner. In addition, we provide evidence that the α subunit from A. tumefaciens, but not from E. coli, is able to interact with the VirG protein. These data suggest that transcription of virulence genes requires specific interaction between VirG and the α subunit of A. tumefaciens and that the α subunit from E. coli is unable to effectively interact with the VirG protein. This work provides the basis for future studies designed to examinevir gene expression as well as the T-DNA transfer process in E. coli.