The marine bacterium Vibrio vulnificus causes food-borne diseases, which may lead to life-threatening septicemia in some individuals. Therefore, identifying virulence factors in V. vulnificus is of high priority. We performed a transcriptome analysis on V. vulnificus after infection of human intestinal HT29-methotrexate cells and found induction of plpA, encoding a putative phospholipase, VvPlpA. Bioinformatics, biochemical, and genetic analyses demonstrated that VvPlpA is a phospholipase A2 secreted in a type II secretion system-dependent manner. Compared with the wild type, the plpA mutant exhibited reduced mortality, systemic infection, and inflammation in mice as well as low cytotoxicity toward the human epithelial INT-407 cells. Moreover, plpA mutation attenuated the release of actin and cytosolic cyclophilin A from INT-407 cells, indicating that VvPlpA is a virulence factor essential for causing lysis and necrotic death of the epithelial cells. plpA transcription was growth phase– dependent, reaching maximum levels during the early stationary phase. Also, transcription factor HlyU and cAMP receptor protein (CRP) mediate additive activation and host-dependent induction of plpA. Molecular biological analyses revealed that plpA expression is controlled via the promoter, PplpA, and that HlyU and CRP directly bind to PplpA upstream sequences. Taken together, this study demonstrated that VvPlpA is a type II secretion system-dependent secretory phospholipase A2 regulated by HlyU and CRP and is essential for the pathogenicity of V. vulnificus.
Bibliographical noteFunding Information:
This work was supported by National Research Foundation Mid-career Researcher Program Grant 2015R1A2A1A13001654 from the Ministry of Science, ICT, and Future Planning and the R&D Convergence Center Sup-port Program of the Ministry of Agriculture, Food, and Rural Affairs, Repub-lic of Korea (to S. H. C.). The authors declare that they have no conflicts of interest with the contents of this article. This article contains supplemental Tables S1 and S2. † Deceased November 2, 2015. 1To whom correspondence may be addressed. Tel.: 82-42-879-8212; E-mail: firstname.lastname@example.org. 2To whom correspondence may be addressed. Tel.: 82-2-880-4857; E-mail: email@example.com.
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