AUTHOR=Qin Lei , Wang Xingqiang , Gao Yingli , Bi Keran , Wang Weixia 

TITLE=Roles of EvpP in Edwardsiella piscicida-Macrophage Interactions

JOURNAL=Frontiers in Cellular and Infection Microbiology

VOLUME=10

YEAR=2020

URL=https://www.frontiersin.org/journals/cellular-and-infection-microbiology/articles/10.3389/fcimb.2020.00053

DOI=10.3389/fcimb.2020.00053

ISSN=2235-2988

ABSTRACT=<p><italic>Edwardsiella piscicida</italic> is found to be an important facultative intracellular pathogen with a broad host range. These organisms can replicate and survive within host macrophages to escape from the subversion of the immune defense. <italic>E. piscicida</italic>-macrophage interaction is very important in determining the outcome of edwardsiellasis. As an effector protein of <italic>E. piscicida</italic> T6SS, EvpP has been determined to be a very important virulence factor for <italic>E. piscicida</italic>, although its precise role in <italic>E. piscicida</italic>-macrophage interactions is not yet clear. In this study, the roles of EvpP in <italic>E. piscicida</italic>-macrophage interactions were characterized. Here, we constructed the deletion mutants of <italic>evp</italic>P (Δ<italic>evp</italic>P) and complementation (Δ<italic>evp</italic>P-C) by the allelic exchange method. Compared to wild type strain (WT), Δ<italic>evp</italic>P was found to be attenuated for growth within macrophages. In line with this observation, we found its survival capacity was lower than WT under oxidative and acid stress <italic>in vitro</italic>, which simulate conditions encountered in host macrophages. Attenuation of Δ<italic>evp</italic>P also correlated with enhanced activation of macrophages, as reflected by augmented NO production in Δ<italic>evp</italic>P-treated macrophages. Moreover, compared to WT, Δ<italic>evp</italic>P induced markedly increased apoptosis of macrophages, characterized by increased Annexin V binding and the activation of cleaved caspase-3. These findings provided strong evidence that EvpP is involved in the process of <italic>E. piscicida-</italic>macrophage interactions and is required for its survival and replication in macrophages. Thus, we propose that EvpP might be an important factor that controlling the fate of <italic>E. piscicida</italic> inside macrophages. To further exploring the underlying mechanism of EvpP action, the cDNA library was constructed from <italic>E. piscicida</italic>-infected macrophages and a yeast two-hybrid screen was performed to search for cellular proteins interacting with EvpP. Ribosomal protein S5 (RPS5) was identified as a target of EvpP. Furthermore, the interaction was validated with co-immunoprecipitation assay. This result implies that the observed effect of EvpP on macrophages might be related to RPS5-mediated regulation, contributing to a better understanding of the mechanisms of EvpP involved in <italic>E. piscicida</italic>-macrophage interactions.</p>