Engineering of Crown Gall Resistant Plants
Presented by: Sarah Andrews
Mentor: Walt Ream
Dept. of Microbiology, Oregon State University
Agrobacterium tumefaciens is responsible for crown pll, a disease that affects woody dicotyledonous plants such as grape vines, nut and fruit trees and lilac bushes. This disease is characterized by the formation of tumors on the roots or graft sites causing the productivity of the plant to decrease.
The tumor-inducing (TI) plasmid in A. tumefaciens is responsible for tumorgenesis. A single- stranded segment of plasmid, called the T-DNA, is transferred from the bacterium to plant cells where it becomes incorporated into the plant genome. This DNA segrnent contains genes that lead to the overproduction of two plant growth horinones, auxin and cytokinin, causing tumorous growth.
Two proteins involved in the transfer of T-DNA are VirE I and VirE2. VirE2 has no role in the bacterial cell, but once transfeffed to the plant cell it binds to T-DNA, targets it to the nucleus and protects the ssDNA against nucleases. VirE I binds to the single-stranded DNA binding domain of VirE2 in the bacterial cell. VirE I, therefore, acts as a secretory chaperone preventing VirE2 from binding T-DNA prematurely before they are both in the plant cell. If VirE I were present in the plant cell, it may compete with the T-DNA for the single-stranded DNA binding domain. If VirE2 were prevented from binding to the T-DNA by VirE I, then the ssDNA would be unstable and not incorporate into the plant genome.
To test this hypothesis I am making three transgenic plants. One will have the virEl gene, another the WrE2 gene and the third both virEl and virE2. The plant containing the virEl trangene will be infected with wild VyW A. tumefaciens to determine whether the presence of VirE I impedes VirE2-T-DNA interaction leading to loss of pathogenicity. The transgenic plants containing WrE2 and both virE genes will be infected with a virE mutant strain of A. tumefaciens and the same question will be asked. Do tumors form?