Section of Microbial Pathogenesis, Yale University School of Medicine, Boyer Center for Molecular Medicine, Yale University, New Haven, Connecticut, CT 06536, USA.
Nature. 2011 Aug 7;477(7362):103-6. doi: 10.1038/nature10335.
The intracellular pathogen Legionella pneumophila modulates the activity of host GTPases to direct the transport and assembly of the membrane-bound compartment in which it resides. In vitro studies have indicated that the Legionella protein DrrA post-translationally modifies the GTPase Rab1 by a process called AMPylation. Here we used mass spectrometry to investigate post-translational modifications to Rab1 that occur during infection of host cells by Legionella. Consistent with in vitro studies, DrrA-mediated AMPylation of a conserved tyrosine residue in the switch II region of Rab1 was detected during infection. In addition, a modification to an adjacent serine residue in Rab1 was discovered, which was independent of DrrA. The Legionella effector protein AnkX was required for this modification. Biochemical studies determined that AnkX directly mediates the covalent attachment of a phosphocholine moiety to Rab1. This phosphocholine transferase activity used CDP-choline as a substrate and required a conserved histidine residue located in the FIC domain of the AnkX protein. During infection, AnkX modified both Rab1 and Rab35, which explains how this protein modulates membrane transport through both the endocytic and exocytic pathways of the host cell. Thus, phosphocholination of Rab GTPases represents a mechanism by which bacterial FIC-domain-containing proteins can alter host-cell functions.
胞内病原体军团菌会调节宿主 GTP 酶的活性，以指导其所在的膜结合隔室的运输和组装。体外研究表明，军团菌蛋白 DrrA 通过一种称为 AMPylation 的过程对 GTP 酶 Rab1 进行翻译后修饰。在这里，我们使用质谱法研究了宿主细胞感染期间 Rab1 发生的翻译后修饰。与体外研究一致，在感染过程中检测到 DrrA 介导的 Rab1 开关 II 区域保守酪氨酸残基的 AMPylation。此外，还发现 Rab1 中一个相邻丝氨酸残基的修饰，这一修饰独立于 DrrA。军团菌效应蛋白 AnkX 是这种修饰所必需的。生化研究表明，AnkX 直接介导磷酸胆碱部分与 Rab1 的共价连接。这种磷酸胆碱转移酶活性以 CDP-胆碱为底物，需要 AnkX 蛋白 FIC 结构域中的保守组氨酸残基。在感染过程中，AnkX 修饰了 Rab1 和 Rab35，这解释了该蛋白如何通过宿主细胞的内吞和外排途径来调节膜运输。因此，Rab GTP 酶的磷酸胆碱化代表了细菌 FIC 结构域蛋白能够改变宿主细胞功能的一种机制。
