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PMID: 10419950 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, Non-P.H.S. Research Support, U.S. Gov't, P.H.S.

Transcriptional activation of Agrobacterium tumefaciens virulence gene promoters in Escherichia coli requires the A. tumefaciens RpoA gene, encoding the alpha subunit of RNA polymerase.

Journal of bacteriology ·Vol. 181 ·No. 15 ·1999-08-00 ·Pages 4533-9

Lohrke SM, Nechaev S, Yang H, Severinov K, Jin SJ

Abstract

The two-component regulatory system, composed of virA and 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 alpha subunit of RNA polymerase (RNAP), which confers significant expression of a virB promoter (virBp)::lacZ fusion in E. coli in the presence of an active transcriptional regulator virG gene. We conducted in vitro transcription assays using either reconstituted E. coli RNAP or hybrid RNAP in which the alpha subunit was derived from A. tumefaciens. The two forms of RNAP were equally efficient in transcription from a sigma(70)-dependent E. coli galP1 promoter; however, only the hybrid RNAP was able to transcribe virBp in a virG-dependent manner. In addition, we provide evidence that the alpha 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 alpha subunit of A. tumefaciens and that the alpha subunit from E. coli is unable to effectively interact with the VirG protein. This work provides the basis for future studies designed to examine vir gene expression as well as the T-DNA transfer process in E. coli.

MeSH Terms
Agrobacterium tumefaciens/genetics,pathogenicity Amino Acid Sequence Bacterial Proteins/genetics,metabolism Cloning, Molecular DNA-Binding Proteins/genetics,metabolism DNA-Directed RNA Polymerases/chemistry,genetics,metabolism Escherichia coli/genetics Gene Expression Regulation, Bacterial Molecular Sequence Data Plasmids Promoter Regions, Genetic Recombinant Proteins/chemistry,metabolism Restriction Mapping Sequence Alignment Sequence Homology, Amino Acid Transcription Factors/genetics,metabolism Transcription, Genetic Transcriptional Activation Virulence/genetics Virulence Factors
Chemicals
Bacterial Proteins DNA-Binding Proteins Recombinant Proteins Transcription Factors Virulence Factors virG protein, Shigella flexneri DNA-Directed RNA Polymerases RNA polymerase alpha subunit
Authors & Affiliations
5 authors, click to expand affiliations / ORCID
Lohrke S M
Department of Microbiology and Immunology, University of Arkansas for Medical Sciences, Little Rock, Arkansas 72205, USA.
Nechaev S
Yang H
Severinov K
Jin S J
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Article Info
Journal
Journal of bacteriology
Abbr.
J Bacteriol
ISSN
0021-9193
DOI
10.1128/JB.181.15.4533-4539.1999
Published
1999-08-00
Pages
4533-9
Language
English
Region
United States
NLM ID
2985120R
PMCID
PMC103583
Subset
IM
Grants
NIGMS NIH HHS · R01 GM059295 · United States
NIGMS NIH HHS · R01 GM 59295 · United States
Databases
GENBANK
AF111855
External Links
PubMed source DOI: 10.1128/JB.181.15.4533-4539.1999 PMC Full Text View journal details
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