Two Novel Type III-Secreted Proteins of Xanthomonas campestris pv. vesicatoria Are Encoded within the hrp Pathogenicity Island

Abstract

The Hrp type III protein secretion system (TTSS) is essential for pathogenicity of gram-negative plant pathogen Xanthomonas campestris pv. vesicatoria. cDNA-amplified fragment length polymorphism and reverse transcription-PCR analyses identified new genes, regulated by key hrp regulator HrpG, in the regions flanking the hrp gene cluster. Sequence analysis revealed genes encoding HpaG, a predicted leucine-rich repeatcontaining protein, the lysozyme-like HpaH protein, and XopA and XopD, which are similar in sequence to Hpa1 from Xanthomonas oryzae pv. oryzae and PsvA from Pseudomonas syringae, respectively. XopA and XopD (Xanthomonas outer proteins) are secreted by the Xanthomonas Hrp TTSS and thus represent putative effector proteins. Mutations in xopA, but not in xopD, resulted in reduced bacterial growth in planta and delayed plant reactions in susceptible and resistant host plants. Since the xopD promoter contains a putative hrp box, which is characteristic of hrpL-regulated genes in P. syringae and Erwinia spp., the gene was probably acquired by horizontal gene transfer. Interestingly, the regions flanking the hrp gene cluster also contain insertion sequences and genes for a putative transposase and a tRNA Arg . These features suggest that the hrp gene cluster of X. campestris pv. vesicatoria is part of a pathogenicity island.

A genetic screen to isolate type III effectors translocated into pepper cells during Xanthomonas infection

“…xopN, xopQ, xopP, and the C terminus of xopF1 are conserved across Xanthomonas spp. (Table 2), and their GC content is comparable with that of the Xcv hrp cluster (8). These xop genes may be members of a ''core'' group of Xanthomonas spp.…”

“…xopC and xopO did not hybridize to many distant Xanthomonas spp., but have homologs in two tomato phytopathogens, Pst and R. solanacearum (Tables 1 and 2), suggesting that these genes may have been introduced into Xcv by horizontal gene transfer. In fact, the xopC gene is flanked by 62-bp IRs and multiple ORFs predicted to encode transposases and integrases (8). A peptide fragment with homology to the Xcc IS1479 transposase (29) was identified upstream of xopO.…”

“…Several Xcv effectors have been identified (5), and molecular analysis has revealed new insight into the role of these proteins in Xcv pathogenesis. AvrBs2, AvrBs3, HpaA, and XopA are required for full virulence of Xcv (6)(7)(8)(9). XopD and AvrXv4 encode cysteine proteases that cleave the small ubiquitin-like modifier (SUMO) from plant SUMO-protein conjugates (10,11), potentially disrupting eukaryotic processes regulated by SUMO modification.…”

Identification of a host 14-3-3 protein that interacts with Xanthomonas effector AvrRxv

“…The PIP-box that is 31 bases upstream of the -10 promoter motif has not yet been shown to function in HrpX control of expression of avrRxv [6]. Analysis of the genome of Xcv revealed that there are PIP-boxes upstream of coding sequences for genes that have no obvious association with pathogenicity and also, HrpX controlled genes that do not contain this motif [11,12,85]. More work has to be done to understand the role of the PIP-box.…”

“…In addition to AvrRxv, there are three other AvrRxv family members present in Xcv. These are AvrBsT, XopJ and AvrXv4 [6,10,11], all of which are translocated to the host cell [8,12,13]. Genes with high levels of sequence similarity are also present in other plant pathogens [14][15][16][17][18], and in the plant symbiont Rhizobium [19].…”