PSa causes oncogenic suppression of Agrobacterium by inhibiting VirE2 protein export

When coresident with the Ti (tumor-inducing) plasmid, the 21-kDa product of the osa gene of the plasmid pSa can suppress crown gap tnmorigenesis incit ed by Agrobacterium tumefaciens. Neither T-DNA processing nor vir (virulenc e) gene induction is affected by the presence of asa in the bacterium. We u sed Arabidopsis thaliana root segments and tobacco leaf discs to demonstrat e that Osa inhibits A. tumefaciens from transforming these plants to the st able phenotypes of tumorigenesis, kanamycin resistance, and stable beta-glu curonidase (GUS) expression. When A. tumefaciens contained osa, the lack of expression of transient GUS activity in infected plant tissues, as well as the lack of systemic viral symptoms following agroinfection of Nicotiana B enthamiana by tomato mottle virus, suggested that oncogenic suppression by Osa occurs before T-DNA enters the plant nucleus. The extracellular complem entation of an A. tumefaciens virE2 mutant (the T-DNA donor strain) by an A . tumefaciens strain lacking T-DNA but containing a wild-type virE2 gene (t he VirE2 donor strain) was blocked when osa was present in the VirE2 donor strain, but not If hen osa was present in the T-DNA donor strain. These dat a indicate that osa inhibits VirE2 protein, but not T-DNA export from A. tu mefaciens, These data further suggest that VirE2, protein and T-DNA are sep arately exported from the bacterium. The successful infection of Datura str amonium plants and leaf discs of transgenic tobacco plants expressing VirE2 protein by an A. tumefaciens virE2 mutant carrying osa confirmed that onco genic suppression by osa does not occur by blocking T-DNA transfer. Overexp ression of virB9, virB10, and virB11 in A. tumefaciens did not overcome onc ogenic suppression by osa. The finding that the expression of the osa gene by itself rather than the formation of a conjugal intermediate with pSa, bl ocks transformation suggests that the mechanism of oncogenic suppression by osa may differ from that of the IncQ plasmid RSF1010.