Role of type VI secretion system protein TssJ-3 in virulence and intracellular survival of Burkholderia pseudomallei.

TssJ-3 is an outer-membrane lipoprotein and is one of the key components of the type VI secretion system in Burkholderia pseudomallei. TssJ translocates effector proteins to target cells to induce innate immune response in the host. However, the tssJ gene has not been identified in B. pseudomallei and its function in this bacterium has not yet been characterized. tssJ-3 knockout and tssJ-3-complemented B. pseudomallei strains were constructed to determine the effects of tssJ-3 on bacterial growth, biofilm formation, flagellum synthesis, motility, host cell infection, and gene expression in B. pseudomallei. We found that the ΔtssJ-3 mutant strain of B. pseudomallei showed significantly suppressed biofilm formation, flagellum synthesis, bacterial growth, motility, and bacterial invasion into host cells (A549 cells). Furthermore, the ΔtssJ-3 mutation downregulated multiple key genes, including biofilm and flagellum-related genes in B. pseudomallei and induced interleukin-8 gene expression in host cells. These results suggest that tssJ-3, an important gene controlling TssJ-3 protein expression, has regulatory effects on biofilm formation and flagellum synthesis in B. pseudomallei. In addition, B. pseudomallei-derived tssJ-3 contributes to cell infiltration and intracellular replication. This study provides a molecular basis of tssJ-3 for developing therapeutic strategies against B. pseudomallei infections.

Construction and analysis of the immune effect of Vibrio harveyi subunit vaccine and DNA vaccine encoding TssJ antigen.

Vibrio harveyi, a severe pathogen infects different kinds of sea animals, causes huge economic loss in aquaculture industry. In order to control the Vibriosis disease caused mainly by V. harveyi and other Vibrio spp., the best solution lies in developing corresponding efficient vaccines. In this study, we have cloned and analysed a putative antigen TssJ from the T6SS of V. harveyi, which has the potential as a vaccine against infection. The sequence analysis and western blotting experiments indicated that TssJ anchored in outer membrane and there were several antigenic determinants existed on its extracellular region. Two forms of universal vaccines, subunit vaccine and DNA vaccine, were developed based on TssJ and applied in Trachinotus ovatus. The results showed that both of the two vaccines could generate a moderate protection in fish against V. harveyi. The relative percentage survival (RPS) of subunit vaccine and DNA vaccine were 52.39% and 69.11%, respectively. Immunological analysis showed both subunit vaccine and DNA vaccine enhanced acid phosphatase, alkaline phosphatase, superoxide dismutase, and lysozyme activities. Specific serum antibodies against TssJ in the fish vaccinated with subunit vaccine was much higher than that in the DNA vaccine group. Several immune-related genes, i.e., IL10, C3, MHC Iα, MHC IIα, and IgM, were induced both by the two forms of vaccines. TNFα and Mx were only upregulated in the DNA vaccine group. However, the induction levels of these genes induced by DNA vaccine were higher than subunit vaccine. All these findings suggested that TssJ from V. harveyi had a potential application value in vaccine industry.

Structure of the T6SS lipoprotein TssJ1 from Pseudomonas aeruginosa.

The type VI secretion system of Pseudomonas aeruginosa has been shown to be responsible for the translocation of bacteriolytic effectors into competing bacteria. A mechanistic understanding of this widely distributed secretion system is developing and structural studies of its components are ongoing. Two representative structures of one highly conserved component, TssJ, from Escherichia coli and Serratia marcescens have been published. Here, the X-ray crystal structure of TssJ1 from P. aeruginosa is presented at 1.4 Å resolution. The overall structure is conserved among the three proteins. This finding suggests that the homologues function in a similar manner and bolsters the understanding of the structure of this family of proteins.