Agrobacterium tumefaciens and A. rhizogenes transfer genes and proteins into plant cells via a type IV secretion system. Transferred DNA (T-DNA) is stably inherited and expressed in plant cells, causing crown gall or hairy root disease. VirE2 is a secreted ssDNA-binding protein crucial for efficient T-DNA transfer and integration. VirE2 targets T-DNA into the nucleus. Despite the importance of VirE2 for gene transfer to plants, some strains of A. rhizogenes lack VirE2, but they still transfer T-DNA efficiently. A gene (GALLS) from A. rhizogenes restores pathogenicity to virE2 - A. tumefaciens. GALLS, like VirE2, may be secreted into plant cells. GALLS contains a putative nuclear localization signal and helicase motifs similar to those in TraA, which mediates conjugal DNA transfer. GALLS contains three repeats that do not resemble any known protein. GALLS substitutes for VirE2 but does not resemble it. Does GALLS perform similar biochemical functions, or does it compensate for loss of VirE2 by providing a new activity? To address this, the following will be done: Create mutations in GALLS to identify domains important for DNA transfer. Express GALLS in plants to ask whether it functions in plant cells and accumulates in the nucleus. Ask whether GALLS protects T- DNA in plant cells or bacteria. Test GALLS for helicase activity and binding to ATP or DNA. Crown gall is economically important, and A. tumefaciens type IV secretion is an excellent model for related systems in human pathogens. Plant transformation by A. tumefaciens is widely used to create transgenic crops. GALLS mediates essential steps in gene transfer to plants and may provide an alternate route into the nucleus. This may allow better control over transgene structure, copy number, and location. Thus, it is important to understand how GALLS works.
