Direct visualization of the expression and localization of chlamydial effector proteins within infected host cells.
Direct visualization of the expression and localization of chlamydial effector proteins within infected host cells.
复制标题
直接可视化受感染宿主细胞内衣原体效应蛋白的表达和定位。
DOI：
10.1093/femspd/fty011
复制
发表时间：
2018
影响因子：
3.3
通讯作者：
Stephens,RichardS
中科院分区：
文献类型：
--
作者：
Wang,Xiaogang;Hybiske,Kevin;Stephens,RichardS
衣原体分泌到宿主细胞的效应蛋白的多样化阵列，但在表征这些蛋白质的时空定位的进展已被阻碍了缺乏遗传方法inChlamydia和活细胞环境中研究这些蛋白质的挑战。我们采用了一种分裂的绿色荧光蛋白（GFP）系统用于衣原体标记衣原体效应蛋白，并跟踪它们在天然环境下在宿主细胞中的定位。通过对几种已知的衣原体蛋白（包括印加、CT 005和CT 694）的检测，证实了该系统的有效性。我们进一步使用这种方法来检测两个衣原体deubiquitinases（CT 867和CT 868）在感染过程中的活细胞内。CT 868只定位于早期和晚期发育阶段的包涵体膜。CT 867定位于衣原体包涵体膜在早期发育阶段，并伴随着定位于宿主质膜在感染的后期阶段。这些数据表明，衣原体去泛素化酶通过靶向细胞膜和衣原体包涵体膜上的蛋白质在衣原体发病机制中发挥重要作用。split-GFP技术被证明是一个强大的和有效的方法来确定重要的衣原体毒力因子的分泌和细胞定位。
Chlamydiasecrete into host cells a diverse array of effector proteins, but progress in characterizing the spatiotemporal localization of these proteins has been hindered by a paucity of genetic approaches inChlamydiaand also by the challenge of studying these proteins within the live cellular environment. We adapted a split-green fluorescent protein (GFP) system for use inChlamydiato label chlamydial effector proteins and track their localization in host cells under native environment. The efficacy of this system was demonstrated by detecting several knownChlamydiaproteins including IncA, CT005 and CT694. We further used this approach to detect two chlamydial deubiquitinases (CT867 and CT868) within live cells during the infection. CT868 localized only to the inclusion membrane at early and late developmental stages. CT867 localized to the chlamydial inclusion membrane at an early developmental stage and was concomitantly localized to the host plasma membrane at a late stage during the infection. These data suggest that chlamydial deubiquitinase play important roles for chlamydial pathogenesis by targeting proteins at both the plasma membrane and the chlamydial inclusion membrane. The split-GFP technology was demonstrated to be a robust and efficient approach to identify the secretion and cellular localization of important chlamydial virulence factors.