Genetic analysis of the virD operon of Agrobacterium tumefaciens: a search for functions involved in transport of T-DNA into the plant cell nucleus and in T-DNA integration.

The transferred DNA(T-DNA) is transported from Agrobacterium tumefaciens to the nucleus and is stably integrated into the genome of many plant species. It has been proposed that the VirD2 protein, tightly attached to the T-DNA, pilots the T-DNA into the plant cellnucleus and that it is involved in integration. Using agroinfection and beta-glucuronidaseexpression as two different very sensitive transient assays for T-DNA transfer, together with assays for stable integration, we have shown that the C-terminal half of the VirD2 protein and the VirD3 protein are not involved in T-DNA integration. However, the bipartite nuclear localization signal, which is located within the C terminusof the VirD2 protein and which has previously been shown to be able to target a foreign protein into the plant cellnucleus, was shown to be required for efficient T-DNA transfer. virD4 mutants were shown by agroinfection to be completely inactive in T-DNA transfer.