THE XANTHOMONAS TYPE III EFFECTOR XOPAE ENCODES A E3 UBIQUITIN LIGASE THAT INHIBITS PAMP-TRIGGERED IMMUNITY

The Gram-negative bacterium Xanthomonas euvesicatoria (Xcv) is the causal agent of spot disease in pepper and tomato plants. Xcv pathogenicity depends on a type III secretion system (T3S) that translocates effector proteins into the plant cell. Collectively, T3S effectors suppress plant PAMP-triggered immunity (PTI), alter metabolism and gene expression of the host for the benefit of the pathogen. In the genome of the Xcv strain 85-10 there are about 35 genes that encode T3S effectors. The XCV0408Ψ gene, which in other Xanthomonas strains encodes the XopAE effector, was annotated as a pseudogene in the Xcv 85-10 genome because of a frameshift mutation in its open reading frame. By expression and translocation analysis of XCV0408Ψ, we found that this gene encodes a functional T3S effector protein. Xcv 85-10 XopAE displayed the ability to interfere with PTI as it inhibited the activation of a flg22-responsive promoter in a pathogen-free protoplast assay and prevented callose deposition at the plant cell wall. PTI signaling involves activation of mitogen-activated protein (MAP) kinase cascades. However, expression of XopAE did not affect flg22-mediated phosphorylation of the MPK3 and MPK6 MAP kinases indicating that this effector interferes with PTI signaling downstream to immunity-associated MAP kinase cascades. Homology modeling of the XopAE C-terminus revealed a three-dimensional fold similar to that of the E3 ubiquitin ligase domain of the Xcv 85-10 effector XopL. Follow up biochemical analysis demonstrated that XopAE is an active E3 ubiquitin ligase in vitro. Together our findings provide new insights into the function and biochemical properties of a previously uncharacterized Xcv effector protein.