Cell Biology and Metabolism Program, Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, Maryland, USA.
Infect Immun. 2013 Jun;81(6):2226-35. doi: 10.1128/IAI.00157-13. Epub 2013 Apr 8.
When the bacterium Legionella pneumophila, the causative agent of Legionnaires' disease, is phagocytosed by alveolar macrophages, it delivers a large number of effector proteins through its Dot/Icm type IV secretion system into the host cell cytosol. Among those proteins is LidA, an effector that interacts with several host GTPases of the Rab family, including Rab6A', a regulator of retrograde vesicle trafficking within eukaryotic cells. The effect of LidA on Rab6A' function and the role of Rab6A' for L. pneumophila growth within host cells has been unclear. Here, we show that LidA preferentially binds Rab6A' in the active GTP-bound conformation. Rab6 binding occurred through the central region of LidA and followed a stoichiometry for LidA and Rab6A' of 1:2. LidA maintained Rab6A' in the active conformation by efficiently blocking the hydrolysis of GTP by Rab6A', even in the presence of cellular GTPase-activating proteins, suggesting that the function of Rab6A' must be important for efficient intracellular replication of L. pneumophila. Accordingly, we found that production of constitutively inactive Rab6A'(T27N) but not constitutively active Rab6A'(Q72L) significantly reduced the ability of L. pneumophila to initiate intracellular replication in human macrophages. Thus, the presence of an active pool of Rab6 within host cells early during infection is required to support efficient intracellular growth of L. pneumophila.
当肺炎军团菌（Legionnaires' disease 的病原体）被肺泡巨噬细胞吞噬时，它会通过其 Dot/Icm 型 IV 型分泌系统将大量效应蛋白输送到宿主细胞质中。在这些蛋白中，LidA 是一种与 Rab 家族的几种宿主 GTPase 相互作用的效应蛋白，包括 Rab6A'，这是真核细胞内逆行囊泡运输的调节剂。LidA 对 Rab6A'功能的影响以及 Rab6A'对宿主细胞内肺炎军团菌生长的作用尚不清楚。在这里，我们表明 LidA 优先结合活性 GTP 结合构象中的 Rab6A'。Rab6 结合发生在 LidA 的中心区域，LidA 和 Rab6A'的结合比为 1:2。LidA 通过有效地阻止 Rab6A'的 GTP 水解，使 Rab6A'保持在活性构象中，即使存在细胞 GTPase 激活蛋白，这表明 Rab6A'的功能对肺炎军团菌的有效细胞内复制至关重要。因此，我们发现，持续失活的 Rab6A'（T27N）的产生而不是持续激活的 Rab6A'（Q72L）显著降低了肺炎军团菌在人巨噬细胞中起始细胞内复制的能力。因此，宿主细胞内早期存在活性 Rab6 池对于支持肺炎军团菌的有效细胞内生长是必需的。
