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Journal of Bacteriology, June 2001, p. 3310-3317, Vol. 183, No. 11
0021-9193/01/$04.00+0   DOI: 10.1128/JB.183.11.3310-3317.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

ChvD, a Chromosomally Encoded ATP-Binding Cassette Transporter-Homologous Protein Involved in Regulation of Virulence Gene Expression in Agrobacterium tumefaciens

Zhenying Liu,1 Mark Jacobs,2 Dennis A. Schaff,1,dagger Colleen A. McCullen,1 and Andrew N. Binns1,*

Plant Science Institute, Department of Biology, University of Pennsylvania, Philadelphia, Pennsylvania 19104-6018,1 and Department of Biology, Swarthmore College, Swarthmore, Pennsylvania 190812

Received 30 November 2000/Accepted 6 March 2001

A yeast two-hybrid screen searching for chromosomally encoded proteins that interact with the Agrobacterium tumefaciens VirB8 protein was carried out. This screen identified an interaction candidate homologous to the partial sequence of a gene that had previously been identified in a transposon screen as a potential regulator of virG expression, chvD. In this report, the cloning of the entire chvD gene is described and the gene is sequenced and characterized. Insertion of a promoterless lacZ gene into the chvD locus greatly attenuated virulence and vir gene expression. Compared to that of the wild-type strain, growth of the chvD mutant was reduced in rich, but not minimal, medium. Expression of chvD, as monitored by expression of beta -galactosidase activity from the chvD-lacZ fusion, occurred in both rich and minimal media as well as under conditions that induce virulence gene expression. The ChvD protein is highly homologous to a family of ATP-binding cassette transporters involved in antibiotic export from bacteria and has two complete Walker box motifs. Molecular genetic analysis demonstrated that disruption of either Walker A box, singly, does not inactivate this protein's effect on virulence but that mutations in both Walker A boxes renders it incapable of complementing a chvD mutant strain. Constitutive expression of virG in the chvD mutant strain restored virulence, supporting the hypothesis that ChvD controls virulence through effects on virG expression.

* Corresponding author. Mailing address: Plant Science Institute, Department of Biology, University of Pennsylvania, Philadelphia, PA 19104-6018. Phone: (215) 898-8684. Fax: (215) 898-8780. E-mail: abinns{at}sas.upenn.edu.

dagger Present address: Agricultural Products Group, FMC Corporation, Philadelphia, PA 19103.

Journal of Bacteriology, June 2001, p. 3310-3317, Vol. 183, No. 11
0021-9193/01/$04.00+0   DOI: 10.1128/JB.183.11.3310-3317.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.


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