AGROBACTERIUM-TUMEFACIENS TRANSFERS EXTREMELY LONG T-DNAS BY A UNIDIRECTIONAL MECHANISM

During crown gall tumorigenesis, part of the Agrobacterium tumefaciens tumor-inducing (Ti) plasmid, the T-DNA, integrates into plant DNA. Di rect repeats define the left and right ends of the T-DNA, but tumorige nesis requires only the right-hand repeat. Virulence (vir) genes act i n trans to mobilize the T-DNA into plant cells. Transfer of T-DNA begi ns when the VirD endonuclease cleaves within the right-hand border rep eat. Although the T-DNA right-border repeat promotes T-DNA transmissio n best in its normal orientation, an inverted right border exhibits re duced but significant activity. Two models may account for this dimini shed tumorigenesis. The right border may function bidirectionally, wit h strong activity only in its wild-type orientation, or it may promote T-DNA transfer in a unidirectional manner such that, with an inverted right border, transfer proceeds around the entire Ti plasmid before r eaching the T-DNA. To determine whether a substantial portion of the T i plasmid is transferred to plant cells, as predicted by the unidirect ional-transfer hypothesis, we examined T-DNAs in tumors induced by str ains containing a Ti plasmid with a right border inverted with respect to the T-DNA oncogenes. These tumors contained extremely long T-DNAs corresponding to most or all of the Ti plasmid. To test whether the ri ght border can function bidirectionally, we inserted T-DNAs with eithe r a properly oriented or an inverted right border into a specific site in the A. tumefaciens chromosome. A border situated to transfer the o ncogenes first directed T-DNA transfer even from the bacterial chromos ome, whereas a border in the opposite (inverted) orientation did not t ransfer the oncogenes to plant cells. Our results indicate that the ri ght-border repeat functions in a unidirectional manner.