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PMID: 12560481 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

Analysis of Vir protein translocation from Agrobacterium tumefaciens using Saccharomyces cerevisiae as a model: evidence for transport of a novel effector protein VirE3.

Nucleic acids research ·Vol. 31 ·No. 3 ·2003-02-01 ·Pages 860-8

Schrammeijer B, den Dulk-Ras A, Vergunst AC, Jurado Jácome E, Hooykaas PJ

Abstract

Agrobacterium tumefaciens causes crown gall disease on a variety of plants. During the infection process Agrobacterium transfers a nucleoprotein complex, the VirD2 T-complex, and at least two Vir proteins, VirE2 and VirF, into the plant cell via the VirB/VirD4 type IV secretion system. Recently, we found that T-DNA could also be transferred from Agrobacterium to Saccharomyces cerevisiae. Here, we describe a novel method to also detect trans-kingdom Vir protein transfer from Agrobacterium to yeast, using the Cre/lox system. Protein fusions between Cre and VirE2 or VirF were expressed in AGROBACTERIUM: Transfer of the Cre-Vir fusion proteins from Agrobacterium to yeast was monitored by a selectable excision event resulting from site-specific recombination mediated by Cre on a lox-flanked transgene in yeast. The VirE2 and VirF proteins were transported to yeast via the virB-encoded transfer system in the presence of coupling factor VirD4, analogous to translocation into plant cells. The yeast system therefore provides a suitable and fast model system to study basic aspects of trans-kingdom protein transport from Agrobacterium into host cells. Using this method we showed that VirE2 and VirF protein transfer was inhibited by the presence of the Osa protein. Besides, we found evidence for a novel third effector protein, VirE3, which has a similar C-terminal signature to VirE2 and VirF.

MeSH Terms
Agrobacterium tumefaciens/genetics,metabolism Bacterial Proteins/genetics,metabolism,physiology DNA-Binding Proteins/genetics,metabolism Integrases/genetics Ion Channels/genetics,metabolism Models, Biological Protein Transport Recombinant Fusion Proteins/metabolism Saccharomyces cerevisiae/metabolism Viral Proteins/genetics Virulence Factors
Chemicals
Bacterial Proteins DNA-Binding Proteins Ion Channels OSA protein, Shigella flexneri Recombinant Fusion Proteins Viral Proteins Virulence Factors virE2 protein, Agrobacterium Cre recombinase Integrases
Authors & Affiliations
5 authors, click to expand affiliations / ORCID
Schrammeijer Barbara
Institute of Molecular Plant Sciences, Clusius Laboratory, Leiden University, Wassenaarseweg 64, 2333 AL Leiden, The Netherlands.
den Dulk-Ras Amke
Vergunst Annette C
Jurado Jácome Esmeralda
Hooykaas Paul J J
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Article Info
Journal
Nucleic acids research
Abbr.
Nucleic Acids Res
ISSN
1362-4962
DOI
10.1093/nar/gkg179
Published
2003-02-01
Pages
860-8
Language
English
Region
England
NLM ID
0411011
PMCID
PMC149200
Subset
IM
External Links
PubMed source DOI: 10.1093/nar/gkg179 PMC Full Text View journal details
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