Roles of EvpP in Edwardsiella piscicida-Macrophage Interactions
Roles of EvpP in Edwardsiella piscicida-Macrophage Interactions
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EvpP 在杀鱼爱德华氏菌-巨噬细胞相互作用中的作用
DOI：
10.3389/fcimb.2020.00053
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发表时间：
2020-02
影响因子：
5.7
通讯作者：
Wang Weixia
中科院分区：
文献类型：
--
作者：
Qin Lei;Wang Xingqiang;Gao Yingli;Bi Keran;Wang Weixia
杀鱼爱德华氏菌是一种重要的兼性胞内病原菌，具有广泛的寄主范围。这些生物体可以在宿主巨噬细胞内复制和存活，以逃避免疫防御的破坏。E.杀鱼-巨噬细胞相互作用在决定爱德华氏菌病的结果中非常重要。作为E. EvpP是E.picidaT6SS的一个重要毒力因子。piscicida，虽然它的确切作用，在E.杀鱼-巨噬细胞相互作用尚不清楚。本研究探讨了EvpP在E.表征杀鱼-巨噬细胞相互作用。本文采用等位基因交换法构建了evpP缺失突变体（ΔevpP）和互补突变体（ΔevpP-C）。与野生型菌株（WT）相比，发现ΔevpP在巨噬细胞内的生长减弱。与此观察结果一致，我们发现其在体外氧化和酸应激下的存活能力低于WT，这模拟了宿主巨噬细胞中遇到的条件。ΔevpP的减弱也与巨噬细胞活化增强相关，如Δ evpP处理的巨噬细胞中NO产生增加所反映的。此外，与WT相比，ΔevpP诱导的巨噬细胞凋亡显著增加，其特征在于增加的Annexin V结合和裂解的caspase-3的活化。这些结果为EvpP参与E.杀鱼-巨噬细胞相互作用，并且是其在巨噬细胞中存活和复制所必需的。因此，我们推测EvpP可能是控制E.巨噬细胞内的杀鱼素为了进一步研究EvpP的作用机制，我们从大肠杆菌中构建了EvpP的cDNA文库。进行杀鱼素感染的巨噬细胞和酵母双杂交筛选以寻找与EvpP相互作用的细胞蛋白。核糖体蛋白S5（RPS 5）被鉴定为EvpP的靶标。此外，通过免疫共沉淀试验验证了相互作用。这一结果表明，所观察到的EvpP对巨噬细胞的作用可能与RPS 5介导的调节有关，有助于更好地理解EvpP参与E.杀鱼-巨噬细胞相互作用。
Edwardsiella piscicida 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. E. piscicida-macrophage interaction is very important in determining the outcome of edwardsiellasis. As an effector protein of E. piscicida T6SS, EvpP has been determined to be a very important virulence factor for E. piscicida, although its precise role in E. piscicida-macrophage interactions is not yet clear. In this study, the roles of EvpP in E. piscicida-macrophage interactions were characterized. Here, we constructed the deletion mutants of evpP (ΔevpP) and complementation (ΔevpP-C) by the allelic exchange method. Compared to wild type strain (WT), ΔevpP 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 in vitro, which simulate conditions encountered in host macrophages. Attenuation of ΔevpP also correlated with enhanced activation of macrophages, as reflected by augmented NO production in ΔevpP-treated macrophages. Moreover, compared to WT, ΔevpP 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 E. piscicida-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 E. piscicida inside macrophages. To further exploring the underlying mechanism of EvpP action, the cDNA library was constructed from E. piscicida-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 E. piscicida-macrophage interactions.
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杀鱼爱德华氏菌细胞内易位候选效应子的系统鉴定
影响因子：
5.7
作者：
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通讯作者：
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影响因子：
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作者：
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通讯作者：
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影响因子：
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作者：
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通讯作者：
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