Mutants of Agrobacterium tumefaciens with elevated vir gene expression.
Expression of Agrobacterium tumefaciens virulence (vir) genes requires virA, virG, and a plant-derived inducing compound such as acetosyringone. To identify the critical functional domains of virA and virG, a mutational approach was used. Agrobacterium A136 harboring plasmid pGP159, which contains virA, virG, and a reporter virB:lacZ gene fusion, was mutagenized with UV light or nitrosoguanidine. Survivors that formed blue colonies on a plate containing 5-bromo-4-chloro-3-indolyl beta-D-galactoside were isolated and analyzed. Quantification of beta-galactosidase activity in liquid assays identified nine mutant strains. By plasmid reconstruction and other procedures, all mutations mapped to the virA locus. These mutations caused an 11- to 560-fold increase in the vegetative level of virB:lacZ reporter gene expression. DNA sequence analysis showed that the mutations are located in four regions of VirA: transmembrane domain one, the active site, a glycine-rich region with homology to ATP-binding sites, and a region at the C terminus that has homology to the N terminus of VirG.
Pazour
G J
GJ
Department of Biochemistry, University of Minnesota, St. Paul 55108.
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C N
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Das
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D000595:Amino Acid Sequence; D003062:Codon / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D005798:Genes, Bacterial; D005838:Genotype; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009604:Nitrosoguanidines / Q000494:pharmacology; D010641:Phenotype; D010957:Plasmids; D011994:Recombinant Proteins / Q000378:metabolism; D012231:Rhizobium / Q000187:drug effects / Q000235:genetics* / Q000472:pathogenicity / Q000528:radiation effects; D014466:Ultraviolet Rays; D014774:Virulence / Q000235:genetics; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism
D003062:Codon; D009604:Nitrosoguanidines; D011994:Recombinant Proteins; D001616:beta-Galactosidase