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PMID: 16194240 Published · ppublish English Journal Article Research Support, N.I.H., Extramural Research Support, U.S. Gov't, Non-P.H.S. Research Support, U.S. Gov't, P.H.S.

Agrobacterium tumefaciens oncogenic suppressors inhibit T-DNA and VirE2 protein substrate binding to the VirD4 coupling protein.

Molecular microbiology ·Vol. 58 ·No. 2 ·2005-10-00 ·Pages 565-79

Cascales E, Atmakuri K, Liu Z, Binns AN, Christie PJ

Abstract

Agrobacterium tumefaciens uses a type IV secretion (T4S) system composed of VirB proteins and VirD4 to deliver oncogenic DNA (T-DNA) and protein substrates to susceptible plant cells during the course of infection. Here, by use of the Transfer DNA ImmunoPrecipitation (TrIP) assay, we present evidence that the mobilizable plasmid RSF1010 (IncQ) follows the same translocation pathway through the VirB/D4 secretion channel as described previously for the T-DNA. The RSF1010 transfer intermediate and the Osa protein of plasmid pSa (IncW), related in sequence to the FiwA fertility inhibition factor of plasmid RP1 (IncPalpha), render A. tumefaciens host cells nearly avirulent. By use of a semi-quantitative TrIP assay, we show that both of these 'oncogenic suppressor factors' inhibit binding of T-DNA to the VirD4 substrate receptor. Both factors also inhibit binding of the VirE2 protein substrate to VirD4, as shown by coimmunoprecipitation and bimolecular fluorescence complementation assays. Osa fused to the green fluorescent protein (GFP) also blocks T-DNA and VirE2 binding to VirD4, and Osa-GFP colocalizes with VirD4 at A. tumefaciens cell poles. RSF1010 and Osa interfere specifically with VirD4 receptor function and not with VirB channel activity, as shown by (i) TrIP and (ii) a genetic screen for effects of the oncogenic suppressors on pCloDF13 translocation through a chimeric secretion channel composed of the pCloDF13-encoded MobB receptor and VirB channel subunits. Our findings establish that a competing plasmid substrate and a plasmid fertility inhibition factor act on a common target, the T4S receptor, to inhibit docking of DNA and protein substrates to the translocation apparatus.

MeSH Terms
Agrobacterium tumefaciens/genetics,metabolism Bacterial Proteins/genetics,metabolism DNA, Bacterial/metabolism DNA-Binding Proteins/genetics,metabolism Gene Expression Regulation, Bacterial Genes, Tumor Suppressor Genetic Complementation Test Ion Channels/genetics,metabolism Plant Leaves/microbiology Plasmids/genetics,metabolism Protein Binding Recombinant Fusion Proteins/genetics,metabolism
Chemicals
Bacterial Proteins DNA, Bacterial DNA-Binding Proteins Ion Channels Recombinant Fusion Proteins T-DNA virE2 protein, Agrobacterium
Authors & Affiliations
5 authors, click to expand affiliations / ORCID
Cascales Eric
Department of Microbiology and Molecular Genetics, University of Texas-Houston, Medical School, Houston, TX 77030, USA.
Atmakuri Krishnamohan
Liu Zhenying
Binns Andrew N
Christie Peter J
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Article Info
Journal
Molecular microbiology
Abbr.
Mol Microbiol
ISSN
0950-382X
DOI
10.1111/j.1365-2958.2005.04852.x
Published
2005-10-00
Pages
565-79
Language
English
Region
England
NLM ID
8712028
PMCID
PMC2749481
Subset
IM
Grants
NIGMS NIH HHS · R01 GM048746 · United States
NIGMS NIH HHS · R01 GM048746-14 · United States
NIGMS NIH HHS · GM48746 · United States
External Links
PubMed source DOI: 10.1111/j.1365-2958.2005.04852.x PMC Full Text View journal details
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