Update on Genomics of Agrobacterium-Mediated Plant Transformation Agrobacterium in the Genomics Age

Members of the genus Agrobacterium cause the neoplastic diseases crown gall, hairy root, and cane gall on numerous plant species. Extensive genetic analyses conducted in the 1980s identified key bacterial genes involved in virulence. During the past decade, however, genomic technologies have revealed numerous additional bacterial genes that more subtly influence transformation. The results of these genomic analyses allowed scientists to develop a more integrated view of how Agrobacterium interacts with host plants. In a similar manner, genomic technologies have identified numerous plant genes important for Agrobacteriummediated genetic transformation. Knowledge of these genes and their roles in transformation has revealed how Agrobacterium manipulates its hosts to increase the probability of a successful transformation outcome. In this article, I review our current knowledge of Agrobacterium-plant interactions and how genomic and proteomic technologies have increased our understanding of this unique plant-microbe interaction. Agrobacterium species are phytopathogens that cause a variety of neoplastic diseases, including crown gall (Agrobacterium tumefaciens and Agrobacterium vitis), hairy root (Agrobacterium rhizogenes), and cane gall (Agrobacterium rubi). Virulent strains of Agrobacterium contain tumor-inducing (Ti) or root-inducing (Ri) plasmids. During infection, enzymes encoded by plasmidlocalized virulence (vir) genes process the T-DNA region of these plasmids. The resulting single-strand DNA (T-strand) linked to VirD2 protein exits the bacterium via a type IV protein secretion system and enters the plant cell. Within the plant, T-strands likely form complexes with other secreted virulence effector proteins, including VirE2, VirE3, VirD5, and VirF, and supercomplexes with plant proteins as they traverse the cytoplasm and target the nucleus. Once inside the nucleus, T-strands integrate randomly into the plant genome and express T-DNA-encoded transgenes. Two classes of T-DNA genes mediate the pathology of Agrobacterium infection. The first group, the oncogenes, either effect phytohormone production (iaa and ipt; Akiyoshi et al., 1984; Schroder et al., 1984), sensitize the plant to endogenous hormone levels (rol and other genes of pRi, gene5 and gene6 of pTi; Shen et al., 1988; Spanier et al., 1989; Tinland et al., 1990; Korber et al., 1991), or may be involved in chromatin remodeling (gene6b; Terakura et al., 2007). Expression of these genes results in tumorigenic or rhizogenic growth. A second set of genes directs the synthesis of various low Mr compounds, opines, that can serve as energy sources for the inciting bacterial strain and can perhaps affect virulence (Veluthambi et al., 1989). For reviews, the reader should see Gelvin (2000, 2003), Tzfira and Citovsky (2001, 2003), McCullen and Binns (2006), and Citovsky et al. (2007). In addition, the reader is directed to an excellent new book on Agrobacterium biology (Tzfira and Citovsky, 2008). Most plant biologists, however, best know Agrobacterium as an agent of horizontal gene transfer that plays an essential role in basic scientific research and in agricultural biotechnology. In the 1980s, scientists learned to disarm (delete the oncogenes and, usually, the opine synthase genes) virulent Agrobacterium strains such that tissues infected by the bacteria could regenerate into normal plants (Bevan et al., 1983; Fraley et al., 1983; Herrera-Estrella et al., 1983). Substituting genes of interest for oncogenes and opine synthase genes resulted in plants expressing these novel transgenes and, thus, novel phenotypes. Although transgene substitution for oncogenes within T-DNAwas initially conducted in cis (i.e. novel transgenes were placed within T-DNA of Ti-plasmids; Caplan et al., 1983; Fraley et al., 1985), the development of binary systems, in which T-DNA and virulence helper plasmids were separated into two different replicons (de Framond et al., 1983; Hoekema et al., 1983), greatly increased the utility of Agrobacterium as a vehicle for gene transfer in plant biology laboratories. Throughout its development as a gene jockeying tool, genomic studies on Agrobacterium and its plant hosts guided scientists in basic science and agricultural biotechnology developments. In this article, I review some of the key genomic methodologies and findings that have contributed to our knowledge of how Agrobacterium works and will contribute in the future better to utilize Agrobacterium’s amazing gene transfer abilities in the laboratory and in the agricultural biotechnology industry.