The Type VI Secretion TssEFGK-VgrG Phage-Like Baseplate Is Recruited to the TssJLM Membrane Complex via Multiple Contacts and Serves As Assembly Platform for Tail Tube/Sheath Polymerization.
The Type VI Secretion TssEFGK-VgrG Phage-Like Baseplate Is Recruited to the TssJLM Membrane Complex via Multiple Contacts and Serves As Assembly Platform for Tail Tube/Sheath Polymerization.
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DOI：
10.1371/journal.pgen.1005545
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发表时间：
2015-10
期刊：
影响因子：
4.5
通讯作者：
Cascales E
中科院分区：
文献类型：
--
作者：
Brunet YR;Zoued A;Boyer F;Douzi B;Cascales E
VI型分泌系统（T6 SS）是一种广泛使用的武器，专门用于将毒素蛋白递送到真核和原核细胞中。13个T6 SS亚基通过跨膜复合物组装细胞质收缩结构锚定到细胞包膜。这种结构在进化上、结构上和功能上与收缩性噬菌体的尾部相关。在噬菌体中，尾部组装到蛋白质复合物上，称为基板，其不仅在管和鞘的组装期间充当平台，而且还触发鞘的收缩。尽管在了解T6 SS组装和功能方面取得了进展，但T6 SS胫骨托的组成仍然是未知的。在这里，我们报告，六T6 SS蛋白TssA，TssE，TssF，TssG，TssK和VgrG所需的正确组装的T6 SS尾管，和VgrG，TssE，-F和-G之间的复合物可以分离。此外，我们表明，TssF和TssG共享有限的序列同源性与已知的噬菌体组件，我们报告这些亚基和其他基板和尾部组件之间的相互作用网络。在协议的基板作为装配平台的尾巴，荧光显微镜分析功能GFP-TssF和TssK-GFP融合蛋白显示，这些蛋白质组装稳定和静态的集群上的鞘聚合。最后，我们表明，招聘的基板的装置需要初始定位的膜复合物和接触之间的TssG和内膜TssM蛋白。在环境中，细菌竞争获得营养或特定生态位的特权。因此，细菌进化出了消灭竞争对手的机制。其中，VI型分泌系统（T6 SS）是一个收缩机器，其功能与尿道相当：一个内管被一个收缩结构包裹。在该外鞘收缩时，内管被推向靶细胞并递送抗菌效应物。管状结构组装在一种称为基板的蛋白质复合物上。在这里，我们定义了基板的组成，表明它是由五个亚基：TssE，TssF，TssG，TssK和VgrG。我们进一步详细的TSSF和TSSG蛋白的作用，通过定义它们的本地化和识别它们的合作伙伴。我们发现，除了TssE和VgrG已被证明与噬菌体gp 25和gp 27-gp 5蛋白共享同源性，TssF和TssG蛋白也与噬菌体组分具有同源性。最后，我们表明，该基板被招募到TssJLM膜复合物之前，组装的收缩性尾部结构。这项研究可以更好地了解这种分子武器的组装途径的早期事件。
The Type VI secretion system (T6SS) is a widespread weapon dedicated to the delivery of toxin proteins into eukaryotic and prokaryotic cells. The 13 T6SS subunits assemble a cytoplasmic contractile structure anchored to the cell envelope by a membrane-spanning complex. This structure is evolutionarily, structurally and functionally related to the tail of contractile bacteriophages. In bacteriophages, the tail assembles onto a protein complex, referred to as the baseplate, that not only serves as a platform during assembly of the tube and sheath, but also triggers the contraction of the sheath. Although progress has been made in understanding T6SS assembly and function, the composition of the T6SS baseplate remains mostly unknown. Here, we report that six T6SS proteins–TssA, TssE, TssF, TssG, TssK and VgrG–are required for proper assembly of the T6SS tail tube, and a complex between VgrG, TssE,-F and-G could be isolated. In addition, we demonstrate that TssF and TssG share limited sequence homologies with known phage components, and we report the interaction network between these subunits and other baseplate and tail components. In agreement with the baseplate being the assembly platform for the tail, fluorescence microscopy analyses of functional GFP-TssF and TssK-GFP fusion proteins show that these proteins assemble stable and static clusters on which the sheath polymerizes. Finally, we show that recruitment of the baseplate to the apparatus requires initial positioning of the membrane complex and contacts between TssG and the inner membrane TssM protein. In the environment, bacteria compete for privileged access to nutrients or to a particular niche. Bacteria have therefore evolved mechanisms to eliminate competitors. Among them, the Type VI secretion system (T6SS) is a contractile machine functionally comparable to a crossbow: an inner tube is wrapped by a contractile structure. Upon contraction of this outer sheath, the inner tube is propelled towards the target cell and delivers anti-bacterial effectors. The tubular structure assembles on a protein complex called the baseplate. Here we define the composition of the baseplate, demonstrating that it is composed of five subunits: TssE, TssF, TssG, TssK and VgrG. We further detail the role of the TssF and TssG proteins by defining their localizations and identifying their partners. We show that, in addition to TssE and VgrG that have been shown to share homologies with the bacteriophage gp25 and gp27-gp5 proteins, the TssF and TssG proteins also have homologies with bacteriophage components. Finally, we show that this baseplate is recruited to the TssJLM membrane complex prior to the assembly of the contractile tail structure. This study allows a better understanding of the early events of the assembly pathway of this molecular weapon.