Studies of essential pathogenicity determinants in Gram-negative bacteria have revealed the conservation of type III protein secretion systems that allow delivery of virulence factors into host cells from plant and animal pathogens. Ten of 21 Hrp proteins of the plant pathogen Xanthomonas campestris pv. vesicatoria have been suggested to be part of a type III machinery. Here, we report the hrp-dependent secretion of two avirulence proteins, AvrBs3 and AvrRxv, by X. campestris pv. vesicatoria strains that constitutively express hrp genes. Secretion occurred without leakage of a cytoplasmic marker in minimal medium containing BSA, at pH 5.4. Secretion was strictly hrp-dependent because a mutant carrying a deletion in hrcV, a conserved hrp gene, did not secrete AvrBs3 and AvrRxv. Moreover, the Hrp system of X. campestris pv. vesicatoria was able to secrete proteins from two other plant pathogens: PopA, a protein secreted via the Hrp system in Ralstonia solanacearum, and AvrB, an avirulence protein from Pseudomonas syringae pv. glycinea. Interestingly, X. campestris pv. vesicatoria also secreted YopE, a type IIIsecreted cytotoxin of the mammalian pathogen Yersinia pseudotuberculosis in a hrp-dependent manner. YerA, a YopEspecific chaperone, was required for YopE stability but not for secretion in X. campestris pv. vesicatoria. Our results demonstrate the functional conservation of the type III system of X. campestris for secretion of proteins from both plant and mammalian pathogens and imply recognition of their respective secretion signals.Studies of bacterial pathogens of plants and mammals have revealed common strategies to interact with their hosts. A striking example is the conservation of a protein-secretion system essential for pathogenicity in distantly related Gramnegative bacteria. Type III secretion systems serve to deliver virulence factors into host cells and have been identified in 10 pathogenic bacteria, including the mammalian pathogens Yersinia spp., Shigella flexneri, Salmonella spp., enteropathogenic Escherichia coli, Pseudomonas aeruginosa, and Chlamydia spp., and the plant pathogens Erwinia spp., Pseudomonas syringae, Ralstonia solanacearum, and Xanthomonas campestris (for a review on type III systems, see ref.

-associated), which is specifically required for disease development. hpaA mutants are affected in pathogenicity, but retain in part the ability to induce avirulence gene-mediated, host-specific hypersensitive reaction (HR). In addition, HpaA was found to contain two functional nuclear localization signals, which are important for the interaction with the plant. We propose that HpaA is an effector protein that may be translocated into the host cell via the Hrp secretion pathway.

Genetic studies have previously implicated the prp1 gene family in the defence of potato against infection with the late blight fungus Phytophthora infestans. Here, we show that the concentrations of PRP1 mRNA as well as protein rapidly increase in potato leaves after fungal infection and stay at high levels during an extended period of the infection cycle. After separation of subcellular components by differential centrifugation, PRP1 protein was identified in the cytosolic fraction. Expression studies with chimeric promoter/β‐glucuronidase gene constructs in transgenic potato plants provided evidence that transcription of the prp1‐1 gene, representing one member of the prp1 gene family, is at least partly responsible for the accumulation of PRP1 mRNA and protein upon fungal infection. After expression of the prp1‐1 coding sequence in Escherichia coli, the corresponding 26–kDa protein exhibited glutathione S‐transferase activity with Km values of 9.8 mM and 0.11 mM for the artificial standard substrate 1‐chloro‐2,4–dinitrobenzene and glutathione, respectively. Photoaffinity labeling of the protein with tritiated 5–azido‐indole‐3–acetic acid suggested that the phytohormone indole‐3–acetic acid or a structurally related compound serve as a regulator or substrate of the prp1‐1 encoded glutathione S–transferase. This assumption was further supported by the inhibitory effect of the phytohormone on the enzyme activity in vitro. The implications of these findings for a potential involvement of indole‐3–acetic acid in the control of defence reactions are discussed.

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Xanthomonas campestris pv. vesicatoria (Xcv) is the causal agent of bacterial spot disease on pepper and tomato. Pathogenicity on susceptible plants and the induction of the hypersensitive reaction (HR) on resistant plants requires a number of genes, designated hrp, most of which are clustered in a 23-kb chromosomal region. Nine hrp genes encode components of a type III protein secretion apparatus that is conserved in Gram-negative plant and animal pathogenic bacteria. We have recently demonstrated that Xcv secretes proteins into the culture medium in a hrp-dependent manner. Substrates of the Hrp secretion machinery are pathogenicity factors and avirulence proteins, e.g. AvrBs3. The AvrBs3 protein governs recognition, i.e. HR induction, when bacteria infect pepper plants carrying the corresponding resistance gene Bs3. Intriguingly, the AvrBs3 protein contains eukaryotic signatures such as nuclear localization signals (NLS), and has been shown to act inside the plant cell. We postulate that AvrBs3 is transferred into the plant cell via the Hrp type III pathway and that recognition of AvrBs3 takes place in the plant cell nucleus.