The Gram‐negative bacterium Xanthomonas campestris pv. vesicatoria is the causal agent of bacterial spot disease on pepper and tomato plants. Pathogenicity of X. campestris pv. vesicatoria depends on a type III secretion (T3S) system which translocates bacterial effector proteins into plant cells. At least nine membrane‐associated and cytoplasmic components of the secretion apparatus are homologous to corresponding Sct (secretion and cellular translocation) proteins from animal pathogens, suggesting a similar structural organisation of T3S systems in different bacterial species. T3S in X. campestris pv. vesicatoria also depends on non‐conserved proteins with yet unknown function including the essential pathogenicity factor HrpB4. Here, we show that HrpB4 localises to the cytoplasm and the bacterial membranes and interacts with the cytoplasmic domain of the inner membrane (IM) ring component HrcD and the cytoplasmic HrcQ protein. The analysis of HrpB4 deletion derivatives revealed that deletion of the N‐ or C‐terminal protein region affects the interaction of HrpB4 with HrcQ and HrcD as well as its contribution to pathogenicity. HrcQ is a component of the predicted sorting platform, which was identified in animal pathogens as a dynamic heterooligomeric protein complex and associates with the IM ring via SctK proteins. HrcQ complex formation was previously shown by fluorescent microscopy analysis and depends on the presence of the T3S system. In the present study, we provide experimental evidence that the absence of HrpB4 severely affects the docking of HrcQ complexes to the T3S system but does not significantly interfere with HrcQ complex formation in the bacterial cytoplasm. Taken together, our data suggest that HrpB4 links the predicted cytoplasmic sorting platform to the IM rings of the T3S system.

中文翻译：

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来自野油菜黄单胞菌的 HrpB4。vesicatoria 的作用类似于 SctK 蛋白，并促进预测的分选平台与 III 型分泌系统的对接

革兰氏阴性菌Xanthomonas campestris pv。vesicatoria是辣椒和番茄植物细菌性斑点病的病原体。X. campestris pv. 的致病性。vesicatoria依赖于 III 型分泌 (T3S) 系统，该系统将细菌效应蛋白转移到植物细胞中。分泌器的至少九个膜相关和细胞质成分与来自动物病原体的相应 Sct（分泌和细胞易位）蛋白同源，表明不同细菌物种中 T3S 系统的结构组织相似。X. campestris pv. 中的T3S 。风湿病还取决于具有未知功能的非保守蛋白质，包括必需的致病因子 HrpB4。在这里，我们显示 HrpB4 定位于细胞质和细菌膜，并与内膜 (IM) 环组件 HrcD 和细胞质 HrcQ 蛋白的细胞质域相互作用。HrpB4 缺失衍生物的分析表明，N 端或 C 端蛋白区域的缺失影响 HrpB4 与 HrcQ 和 HrcD 的相互作用及其对致病性的贡献。HrcQ 是预测分选平台的一个组成部分，在动物病原体中被鉴定为动态异源寡聚蛋白复合物，并通过 SctK 蛋白与 IM 环相关联。HrcQ 复合物的形成以前是通过荧光显微镜分析显示的，并且取决于 T3S 系统的存在。在本研究中，我们提供的实验证据表明，HrpB4 的缺失严重影响 HrcQ 复合物与 T3S 系统的对接，但不会显着干扰细菌细胞质中 HrcQ 复合物的形成。总之，我们的数据表明 HrpB4 将预测的细胞质分选平台与 T3S 系统的 IM 环联系起来。