A. Miranda et al., AGROBACTERIUM-TUMEFACIENS TRANSFERS EXTREMELY LONG T-DNAS BY A UNIDIRECTIONAL MECHANISM, Journal of bacteriology, 174(7), 1992, pp. 2288-2297
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.