The capping domain in RalF regulates effector functions.
The capping domain in RalF regulates effector functions.
复制标题
RALF中的上限域调节效应子功能。
DOI：
10.1371/journal.ppat.1003012
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发表时间：
2012
期刊：
影响因子：
6.7
通讯作者：
Roy CR
中科院分区：
文献类型：
--
作者：
Alix E;Chesnel L;Bowzard BJ;Tucker AM;Delprato A;Cherfils J;Wood DO;Kahn RA;Roy CR
嗜肺军团菌效应蛋白RalF作为鸟嘌呤核苷酸交换因子（GEF）激活宿主小GTPase蛋白adp -核糖基化因子（Arf），并将该宿主蛋白招募到病原体所在的空泡中。GEF活性是由位于RalF n端区域的Sec7结构域赋予的。结构研究表明RalF的c端区域与Sec7结构域的残基接触对Arf的相互作用很重要。理论上，RalF的c端区域可以通过阻断Arf与Sec7结构域相互作用的能力来阻止核苷酸交换活性。因此，RalF的c端区域被称为capping结构域。本研究通过比较RalF capping结构域在军团菌RalF蛋白（LpRalF）和从不相关的胞内病原体prowazekii立克次体（RpRalF）分离的RalF同源物中介导的生化和效应活性，研究了RalF capping结构域的作用。这些数据表明，两个RalF蛋白都含有一个功能性的Sec7结构域，并且封盖结构域调节RalF GEF活性。capping结构域具有内在的决定因素，介导宿主细胞内RalF蛋白的定位，并赋予不同的效应活性。由LpRalF的capping结构域介导的定位使GEF能够调节分泌途径中的膜运输，而RpRalF的capping结构域使该细菌GEF能够调节发生在质膜附近的肌动蛋白动力学。因此，这些数据表明，c端capping结构域功能的差异改变了RalF蛋白的体内功能。嗜肺军团菌和普鲁泽氏立克次体是两种致病性细胞内细菌，在系统发育上相距遥远，并表现出不同的细胞内生活方式。有趣的是，这两种生物都编码一种叫做RalF的蛋白质，在军团菌中，它已被证明是一种效应蛋白，可作为真核生物gtp酶Arf家族的鸟嘌呤核苷酸交换因子（GEF）。我们发现这两种蛋白中的Sec7结构域具有相似的激活Arf1的酶促能力，然而，这两种蛋白在哺乳动物细胞内的效应功能不同。军团菌RalF蛋白靶向分泌转运功能，而立克次体RalF蛋白调节质膜上的肌动蛋白动力学。这些RalF效应物活性的差异是由一个c端结构域介导的，该结构域指导定位和与宿主细胞决定因子的相互作用。这些数据揭示了RalF蛋白的重要特性，为军团菌和立克次体感染期间Arf的不同作用提供了见解，也为研究结构相似的Arf gef的细胞内定位如何使蛋白质调节功能不同的宿主细胞过程提供了模型。
The Legionella pneumophila effector protein RalF functions as a guanine nucleotide exchange factor (GEF) that activates the host small GTPase protein ADP-ribosylation factor (Arf), and recruits this host protein to the vacuoles in which this pathogen resides. GEF activity is conferred by the Sec7 domain located in the N-terminal region of RalF. Structural studies indicate that the C-terminal region of RalF makes contacts with residues in the Sec7 domain important for Arf interactions. Theoretically, the C-terminal region of RalF could prevent nucleotide exchange activity by blocking the ability of Arf to interact with the Sec7 domain. For this reason, the C-terminal region of RalF has been termed a capping domain. Here, the role of the RalF capping domain was investigated by comparing biochemical and effector activities mediated by this domain in both the Legionella RalF protein (LpRalF) and in a RalF ortholog isolated from the unrelated intracellular pathogen Rickettsia prowazekii (RpRalF). These data indicate that both RalF proteins contain a functional Sec7 domain and that the capping domain regulates RalF GEF activity. The capping domain has intrinsic determinants that mediate localization of the RalF protein inside of host cells and confer distinct effector activities. Localization mediated by the capping domain of LpRalF enables the GEF to modulate membrane transport in the secretory pathway, whereas, the capping domain of RpRalF enables this bacterial GEF to modulate actin dynamics occurring near the plasma membrane. Thus, these data reveal that divergence in the function of the C-terminal capping domain alters the in vivo functions of the RalF proteins. Legionella pneumophila and Rickettsia prowazekii are two pathogenic intracellular bacteria, phylogenetically distant and presenting different intracellular lifestyles. Interestingly, both organisms encode a protein called RalF, which in Legionella has been shown to be an effector protein that functions as a guanine nucleotide exchange factor (GEF) for the Arf family of eukaryotic GTPases. We show that the Sec7 domains in both proteins have a similar enzymatic ability to activate Arf1, however, the two proteins differ in their effector functions inside mammalian cells. The Legionella RalF protein targets secretory transport functions, whereas, the Rickettsia RalF protein modulates actin dynamics at the plasma membrane. These differences in RalF effector activities are mediated by a C-terminal domain that directs localization and interaction with host cell determinants. These data reveal important properties of the RalF protein that provide insight into differential roles for Arf during infection by Legionella and Rickettsia, and also provide a model for investigating how intracellular localization of structurally similar Arf GEFs enables a protein to regulate functionally distinct host cell processes.
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DOI：
10.1083/jcb.200806041
发表时间：
2008-09-22
期刊：
The Journal of cell biology
影响因子：
--
作者：
Kahn RA;Bruford E;Inoue H;Logsdon JM Jr;Nie Z;Premont RT;Randazzo PA;Satake M;Theibert AB;Zapp ML;Cassel D
通讯作者：
Cassel D
影响因子：
3.7
作者：
Gillespie JJ;Ammerman NC;Dreher-Lesnick SM;Rahman MS;Worley MJ;Setubal JC;Sobral BS;Azad AF
通讯作者：
Azad AF
影响因子：
3.3
作者：
Cohen, Lee Ann;Honda, Akira;Donaldson, Julie G.
通讯作者：
Donaldson, Julie G.
影响因子：
4.8
作者：
Macia, E;Chabre, M;Franco, M
通讯作者：
Franco, M
影响因子：
16
作者：
DiNitto, Jonathan P.;Delprato, Anna;Lambright, David G.
通讯作者：
Lambright, David G.