Abstract
Opportunistic pathogen Vibrio vulnificus causes severe systemic infection in humans with high mortality. Although multiple exotoxins have been characterized in V. vulnificus, their interactions and potential synergistic roles in pathogen-induced host cell death have not been investigated previously. By employing a series of multiple exotoxin deletion mutants, we investigated whether specific exotoxins of the pathogen functioned together to achieve severe and rapid necrotic cell death. Human epithelial cells treated with V. vulnificus with a plpA deletion background exhibited an unusually prolonged cell blebbing, suggesting the importance of PlpA, a phospholipase A2, in rapid necrotic cell death by this pathogen. Additional deletion of the rtxA gene encoding the multifunctional autoprocessing repeats-in-toxin (MARTX) toxin did not result in necrotic cell blebs. However, if the rtxA gene was engineered to produce an effector-free MARTX toxin, the cell blebbing was observed, indicating that the pore forming activity of the MARTX toxin is sufficient, but the MARTX toxin effector domains are not necessary, for the blebbing. When a recombinant PlpA was treated on the blebbed cells, the blebs were completely disrupted. Consistent with this, MARTX toxin-pendent rapid release of cytosolic lactate dehydrogenase was significantly delayed in the plpA deletion background. Mutations in other exotoxins such as elastase, cytolysin/hemolysin, and/or extracellular metalloprotease did not affect the bleb formation or disruption. Together, these findings indicate that the pore forming MARTX toxin and the phospholipase A2, PlpA, cooperate sequentially to achieve rapid necrotic cell death by inducing cell blebbing and disrupting the blebs, respectively.
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Acknowledgements
This work was supported by the National Research Foundation of Korea funded by the Ministry of Science and ICT (NRF-2020R1F1A1070168 to B.S.K.) and the KRIBB Initiative Program (KGM2112234 to M. H. K.). We thank Dr. S. H. Choi at Seoul National University and Dr. K.-H. Lee at Sogang University for sharing the plasmids pMW0613, pKK1301, pKK1407, and pSKvvpM02. We also thank Dr. Y. Kwon at Ewha Womans University for sharing the 3T3-L1 cells.
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Cho, C., Choi, S., Kim, M.H. et al. Vibrio vulnificus PlpA facilitates necrotic host cell death induced by the pore forming MARTX toxin. J Microbiol. 60, 224–233 (2022). https://doi.org/10.1007/s12275-022-1448-x
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DOI: https://doi.org/10.1007/s12275-022-1448-x