title: Interaction of the Agrobacterium tumefaciens virulence protein VirD2 with histones.
abstract: Agrobacterium tumefaciens is a Gram-negative soil bacterium that genetically transforms plants and, under laboratory conditions, also transforms non-plant organisms, such as fungi and yeasts. During the transformation process a piece of ssDNA (T-strand) is transferred into the host cells via a type IV secretion system. The VirD2 relaxase protein, which is covalently attached at the 5' end of the T-strand through Tyr29, mediates nuclear entry as it contains a nuclear localization sequence. How the T-strand reaches the chromatin and becomes integrated in the chromosomal DNA is still far from clear. Here, we investigated whether VirD2 binds to histone proteins in the yeast Saccharomyces cerevisiae. Using immobilized GFP-VirD2 and in vitro synthesized His6-tagged S. cerevisiae proteins, interactions between VirD2 and the histones H2A, H2B, H3 and H4 were revealed. In vivo, these interactions were confirmed by bimolecular fluorescence complementation experiments. After co-cultivation of Agrobacterium strains expressing VirD2 tagged with a fragment of the yellow fluorescent protein analogue Venus with yeast strains expressing histone H2A or H2B tagged with the complementary part of Venus, fluorescence was detected in dot-shaped structures in the recipient yeast cells. The results indicated that VirD2 was transferred from Agrobacterium to yeast cells and that it interacted with histones in the host cell, and thus may help direct the T-DNA (transferred DNA) to the chromatin as a prelude to integration into the host chromosomal DNA.
authors.affiliation: Section Molecular and Developmental Genetics, Institute of Biology, Leiden University, Leiden, The Netherlands.
authors.forename: Suzanne
authors.initials: S
authors.lastname: Wolterink-van Loo
authors.affiliation: Section Molecular and Developmental Genetics, Institute of Biology, Leiden University, Leiden, The Netherlands.
authors.forename: Abril A Escamilla
authors.initials: AAE
authors.lastname: Ayala
authors.affiliation: Section Molecular and Developmental Genetics, Institute of Biology, Leiden University, Leiden, The Netherlands.
authors.forename: Paul J J
authors.initials: PJJ
authors.lastname: Hooykaas
authors.affiliation: Section Molecular and Developmental Genetics, Institute of Biology, Leiden University, Leiden, The Netherlands.
authors.forename: G Paul H
authors.initials: GPH
authors.lastname: van Heusden
mesh_terms: D016960:Agrobacterium tumefaciens / Q000235:genetics / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D006657:Histones / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D020798:Two-Hybrid System Techniques; D037521:Virulence Factors / Q000235:genetics / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D006657:Histones; D029701:Saccharomyces cerevisiae Proteins; D037521:Virulence Factors