title: Involvement of targeted proteolysis in plant genetic transformation by Agrobacterium.
abstract: Genetic transformation of plant cells by Agrobacterium represents a unique case of trans-kingdom DNA transfer. During this process, Agrobacterium exports its transferred (T) DNA and several virulence (Vir) proteins into the host cell, within which T-DNA nuclear import is mediated by VirD2 (ref. 3) and VirE2 (ref. 4) and their host cell interactors AtKAP-alpha and VIP1 (ref. 6), whereas its integration is mediated mainly by host cell proteins. The factors involved in the uncoating of T-DNA from its cognate proteins, which occurs before integration into the host genome, are still unknown. Here, we report that VirF-one of the few known exported Vir proteins whose function in the host cell remains unknown-is involved in targeted proteolysis of VIP1 and VirE2. We show that VirF localizes to the plant cell nucleus and interacts with VIP1, a nuclear protein. VirF, which contains an F-box motif, significantly destabilizes both VIP1 and VirE2 in yeast cells. Destabilization of VIP1 in the presence of VirF was then confirmed in planta. These results suggest that VIP1 and its cognate VirE2 are specifically targeted by the VirF-containing Skp1-Cdc53-cullin-F-box complex for proteolysis. The critical role of proteasomal degradation in Agrobacterium-mediated genetic transformation was also evident from inhibition of T-DNA expression by a proteasomal inhibitor.
authors.affiliation: Department of Biochemistry and Cell Biology, State University of New York, Stony Brook, New York 11794-5215, USA. ttzfira@ms.cc.sunysb.edu
authors.forename: Tzvi
authors.initials: T
authors.lastname: Tzfira
authors.forename: Manjusha
authors.initials: M
authors.lastname: Vaidya
authors.forename: Vitaly
authors.initials: V
authors.lastname: Citovsky
mesh_terms: D001426:Bacterial Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D004269:DNA, Bacterial / Q000235:genetics; D044783:F-Box Proteins / Q000378:metabolism; D015870:Gene Expression; D010447:Peptide Hydrolases / Q000378:metabolism*; D010940:Plant Proteins / Q000378:metabolism; D046988:Proteasome Endopeptidase Complex; D011485:Protein Binding; D011499:Protein Processing, Post-Translational; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D012231:Rhizobium / Q000235:genetics* / Q000378:metabolism* / Q000472:pathogenicity; D044843:SKP Cullin F-Box Protein Ligases / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism; D013379:Substrate Specificity; D014026:Nicotiana / Q000235:genetics* / Q000378:metabolism* / Q000382:microbiology; D014170:Transformation, Genetic; D014774:Virulence; D037521:Virulence Factors / Q000037:antagonists & inhibitors / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D004269:DNA, Bacterial; D044783:F-Box Proteins; D010940:Plant Proteins; D012333:RNA, Messenger; D029701:Saccharomyces cerevisiae Proteins; C036483:T-DNA; D037521:Virulence Factors; D044843:SKP Cullin F-Box Protein Ligases; C120507:SKP1 protein, S cerevisiae; D010447:Peptide Hydrolases; D046988:Proteasome Endopeptidase Complex; C061553:ATP dependent 26S protease