Protein interactions involved in nuclear import of the Agrobacterium VirE2 protein in vivo and in vitro.
Agrobacterium, the only known organism capable of trans-kingdom DNA transfer, genetically transforms plants by transferring a segment of its DNA, T-DNA, into the nucleus of the host cell where it integrates into the plant genome. One of the central events in this genetic transformation process is nuclear import of the T-DNA molecule, which to a large degree is mediated by the bacterial virulence protein VirE2. VirE2 is distinguished by its nuclear targeting, which occurs only in plant but not in animal cells and is facilitated by the cellular VIP1 protein. The molecular mechanism of the VIP1 function is still unclear. Here, we used in vitro assays for nuclear import and quantification of protein-protein interactions to directly demonstrate formation of ternary complexes between VirE2, VIP1, and a component of the cellular nuclear import machinery, karyopherin alpha. Our results indicate that VIP1 functions as a molecular bridge between VirE2 and karyopherin alpha, allowing VirE2 to utilize the host cell nuclear import machinery even without being directly recognized by its components.
Citovsky
Vitaly
V
Department of Biochemistry and Cell Biology, State University of New York at Stony Brook, Stony Brook, New York 11794-5215, USA. vitaly.citovsky@stonybrook.edu
Kapelnikov
Anat
A
Oliel
Shachar
S
Zakai
Nehama
N
Rojas
Maria R
MR
Gilbertson
Robert L
RL
Tzfira
Tzvi
T
Loyter
Abraham
A
D021581:Active Transport, Cell Nucleus / Q000502:physiology*; D000818:Animals; D029681:Arabidopsis Proteins / Q000235:genetics / Q000378:metabolism*; D001426:Bacterial Proteins; D019556:COS Cells; D002522:Chlorocebus aethiops; D004269:DNA, Bacterial / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D005456:Fluorescent Dyes / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D007473:Ion Channels / Q000378:metabolism*; D046911:Macromolecular Substances; D019913:Nuclear Localization Signals; D010940:Plant Proteins / Q000378:metabolism; D011485:Protein Binding; D012231:Rhizobium / Q000235:genetics / Q000378:metabolism*; D014026:Nicotiana / Q000166:cytology / Q000378:metabolism / Q000382:microbiology; D028901:alpha Karyopherins / Q000378:metabolism
D029681:Arabidopsis Proteins; D001426:Bacterial Proteins; D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; D005456:Fluorescent Dyes; D007473:Ion Channels; D046911:Macromolecular Substances; D019913:Nuclear Localization Signals; D010940:Plant Proteins; C036483:T-DNA; C432104:VIP1 protein, Arabidopsis; D028901:alpha Karyopherins; C051583:virE2 protein, Agrobacterium