The Agrobacterium tumefaciens VirB7 lipoprotein is required for stabilization of VirB proteins during assembly of the T-complex transport apparatus

• Published in: Journal of Bacteriology 1996-06, Vol.178 (11), p.3168-3176
• Main Authors: Fernandez, D. (The University of Texas Health Science Center at Houston, Houston, TX.), Spudich, G.M, Zhou, X.R, Christie, P.J
• Format: Article
• Language: English
• Subjects: ADN; AGALLAS; AGROBACTERIUM TUMEFACIENS; Agrobacterium tumefaciens - chemistry; BACTERIA; Bacterial Proteins - chemistry; Bacterial Proteins - genetics; Bacterial Proteins - physiology; Bacteriology; Biological Transport; Flowers & plants; GALLE; Genetics; KALANCHOE; LIPOPROTEINAS; LIPOPROTEINE; Lipoproteins - physiology; MEMBRANAS CELULARES; MEMBRANE CELLULAIRE; MUTANT; MUTANTES; Mutation; Nucleoproteins - metabolism; PATOGENICIDAD; POUVOIR PATHOGENE; PROTEINAS; PROTEINAS AGLUTINANTES; PROTEINE; PROTEINE DE LIAISON; Proteins; SINTESIS DE PROTEINAS; SYNTHESE PROTEIQUE; Virulence Factors
• ISSN: 0021-9193; 1098-5530; 1067-8832
• DOI: 10.1128/jb.178.11.3168-3176.1996

The Agrobacterium tumefaciens virB7 gene product is a lipoprotein whose function is required for the transmission of oncogenic T-DNA to susceptible plant cells. Three lines of study provided evidence that VirB7 interacts with and stabilizes other VirB proteins during the assembly of the putative T-complex transport apparatus. First, a precise deletion of virB7 from the pTiA6NC plasmid of wild-type strain A348 was correlated with significant reductions in the steady-state levels of several VirB proteins, including VirB4, VirB9, VirB10, and VirB11; trans expression of virB7 in the (delta)virB7 mutant partially restored the levels of these proteins, and trans coexpression of virB7 and virB8 fully restored the levels of these proteins to wild-type levels. Second, modulation of VirB7 levels resulted in corresponding changes in the levels of other VirB proteins in the following cell types: (i) a (delta)virB7 mutant expressing virB7 and virB8 from isopropyl-beta-D-thiogalactopyranoside (IPTG)-inducible P lac and other virB genes from acetosyringone (AS)-inducible PvirB; (ii) a (delta)virB operon mutant expressing virB7 and virB8 from Plac and virB9, virB10, and virB11 from PvirB; and (iii) a (delta)virB operon mutant expressing virB7 from IPTG-inducible Plac, and virB9 from an AS-inducible PvirB. Third, the synthesis of a VirB7::PhoA fusion protein in strain A348 was correlated with a significant reduction in the steady-state levels of VirB4, VirB5, and VirB7 through VirB11; these cells also exhibited a severely attenuated virulence phenotype, indicating that synthesis of the fusion protein perturbs the assembly of VirB proteins into a stabilized protein complex required for T-complex transport. Extracts of AS-induced cells electrophoresed under nonreducing conditions possessed undetectable levels of the 32-kDa VirB9 and 4.5-kDa VirB7 monomers and instead possessed a 36-kDa complex that cross-reacted with both VirB7 and VirB9 antisera and accumulated as a function of virB7