Project description:Similar to other plant-parasitic nematodes, root lesion nematodes possess an array of enzymes that are involved in degradation of the plant cell wall. Here we report the identification of a gene encoding a cell wall degrading enzyme, pectin methylesterase PME (EC 3.1.1.11), in the root lesion nematode Pratylenchus penetrans. Both genomic and coding sequences of the gene were cloned for this species, showing the presence of four introns that excluded a potential bacterial contamination. Expression of the Pp-pme gene was localized in the esophageal glands of P. penetrans as determined by in situ hybridization. Temporal expression of Pp-pme in planta was validated for early time points of infection. The possible function and activity of the gene were assessed by transient expression of Pp-pme in N. benthamiana plants via a Potato virus X-based vector. To our knowledge, this is the first report on identification and characterization of a PME gene within the phylum Nematoda.
Project description:Microbes have evolved elaborate mechanisms to cope with competitors, including the type VIsecretion system (T6SS) of Gram-negative bacteria. The T6SS inhibits target cells through contact-dependent translocation of toxic effector proteins across two cellular membranes via an inverted phage-like apparatus. Proteobacteria accomplish this feat by passage of theT6SS needle through a megadalton-size membrane complex (MC), which is essential for T6SS function. Remarkably, although the phylum Bacteroidota encodes a T6SS, it lacks homologs of the MC. We identified five novel genes, essential for T6SS function, that encode a candidate unique Bacteroidota T6SSMC. We purified the T6SSiii MC and revealed its dimensions using electron microscopy. We identified an intricate protein protein interaction network underlying the assembly of the MC, the stoichiometry of the five TssNQOPR components., tTheir predicted structures were validated using crosslinking mass-spectrometry and we assessed the structural homology with known proteins. Importantly, we determine the connection between the T6SSiii MC and the otherwise conserved baseplate involving the hub protein Tss.Finally, phylogenomic analysis of the distribution of T6SSiii MC genes across the phylum Bacteroidota highlights patterns of conservation including the invariant