The Phenolic Recognition Profiles of the Agrobacterium tumefaciens VirA Protein Are Broadened by a High Level of the Sugar Binding Protein ChvE

Agrobacterium tumefaciens infects a wide range of plants by transferring and integrating a piece (the T-DNA) of its tumor-inducing (Ti) plasmid into the plant genome, resulting in the formation of crown gall tumors. To initiate this process, a set of genes, the virulence (vir) genes, on the Ti plasmid must be activated by three kinds of environmental signals at the wound site of a plant. These signals include specific classes of plant phenolic compounds, monosaccharides, and an acidic pH. The signals are perceived by three proteins, the VirA-VirG two-component transduction system and ChvE, a periplasmic sugar binding protein. In response to these signals, the VirA sensor protein is autophosphorylated at His-474, and this phosphate is then transferred to Asp-52 of the response regulator, VirG. The activated VirG then induces the expression of all of the vir genes (for reviews, see references 15, 17, and 32). The key molecule which allows Agrobacterium to sense environmental conditions favorable for T-DNA transfer is the VirA protein, which is anchored to the cytoplasmic membrane by two transmembrane domains. This protein contains four other domains: an amino-terminal periplasmic domain and three cytoplasmic domains. The latter domains include a linker, a kinase, and a carboxyl-terminal region termed the receiver because it contains a region that is homologous to the phosphorylatable receiver domain of VirG (5). The periplasmic domain is responsible for sensing a variety of monosaccharides involved in vir gene induction. The linker domain is necessary for perceiving phenolic compounds and acidity (5). The function of the receiver domain is less clear. Chang and Winans (5) reported that the A6 VirA receiver domain might play an inhibitory role in signal transduction, because when this domain was deleted, monosaccharides alone induced vir gene expression in the absence of phenolic compounds. Similar results were obtained by Gubba et al. (14). However, deleting the C-terminal receiver domain from the VirA protein of Agrobacterium rhizogenes resulted in a strain which showed poor vir gene induction and was nontumorigenic on Kalanchoe diagremontiana leaves (11). The A6 VirA receiver domain apparently restricts the recognition of phenolic inducers by VirA, since certain mutations in the domain or its removal widens the spectrum of inducers which VirA recognizes (6). The chvE gene plays several roles, one in the uptake of specific sugars, another in chemotaxis to these sugars (4, 18), and a third in the VirA-VirG two-component signal transduction system. ChvE is homologous to two periplasmic proteins involved in sugar recognition and uptake in Escherichia coli (4, 18). The inducing monosaccharides, all of which are components of the plant cell wall, are sensed by the VirA protein via the ChvE protein (1, 4). The ChvE protein interacts with each of the various inducing monosaccharides, and the ChvE::monosaccharide complex in turn interacts with the periplasmic region of VirA (4, 26). The periplasmic domain of VirA may repress its function, and this repression is overcome by the binding of the ChvE::monosaccharide complex to the periplasmic domain of VirA (4, 15, 26). All Agrobacterium strains containing a defective chvE are defective in vir gene induction and have limited host ranges compared with those of wild-type strains (13, 18). In strain A348, mutants lacking the periplasmic domain of VirA have the same host range defects and inducing properties as chvE mutants. They are avirulent on some, but not all, plants (4). However, in strain C58, the loss of the ChvE protein results in the almost-complete loss of vir gene induction and an extremely limited host range (10). The chvE gene is regulated by the inducing monosaccharides through the gbpR gene product, GbpR, a member of the LysR family of transcriptional regulators (9). A subset of the monosaccharides which are involved in the induction of the vir genes induces the expression of ChvE a maximum of eightfold; in their absence, GbpR represses its expression (9). In this paper, we present evidence that specific monosaccharides can broaden the spectrum of phenolic compounds which can serve as vir gene inducers. This effect is due to an increased level of the ChvE protein resulting from the induction of chvE gene expression by GbpR and the inducing sugar.