Section of Microbial Pathogenesis, Yale University School of Medicine, Boyer Center for Molecular Medicine, New Haven, CT 06536, USA.
Traffic. 2010 May;11(5):587-600. doi: 10.1111/j.1600-0854.2010.01050.x. Epub 2010 Feb 15.
Biogenesis of a specialized organelle that supports intracellular replication of Legionella pneumophila involves the fusion of secretory vesicles exiting the endoplasmic reticulum (ER) with phagosomes containing this bacterial pathogen. Here, we investigated host plasma membrane SNARE proteins to determine whether they play a role in trafficking of vacuoles containing L. pneumophila. Depletion of plasma membrane syntaxins by RNA interference resulted in delayed acquisition of the resident ER protein calnexin and enhanced retention of Rab1 on phagosomes containing virulent L. pneumophila, suggesting that these SNARE proteins are involved in vacuole biogenesis. Plasma membrane-localized SNARE proteins syntaxin 2, syntaxin 3, syntaxin 4 and SNAP23 localized to vacuoles containing L. pneumophila. The ER-localized SNARE protein Sec22b was found to interact with plasma membrane SNAREs on vacuoles containing virulent L. pneumophila, but not on vacuoles containing avirulent mutants of L. pneumophila. The addition of alpha-SNAP and N-ethylmaleimide-sensitive factor (NSF) to the plasma membrane SNARE complexes formed by virulent L. pneumophila resulted in the dissociation of Sec22b, indicating functional pairing between these SNAREs. Thus, L. pneumophila stimulates the non-canonical pairing of plasma membrane t-SNAREs with the v-SNARE Sec22b to promote fusion of the phagosome with ER-derived vesicles. The mechanism by which L. pneumophila promotes pairing of plasma membrane syntaxins and Sec22b could provide unique insight into how the secretory vesicles could provide an additional membrane reserve subverted during phagosome maturation.
专门细胞器的生物发生支持肺炎军团菌的细胞内复制，涉及从内质网 (ER) 出口的分泌小泡与含有这种细菌病原体的吞噬体融合。在这里，我们研究了宿主质膜 SNARE 蛋白，以确定它们是否在含有肺炎军团菌的液泡运输中发挥作用。通过 RNA 干扰耗尽质膜 SNARE 蛋白Syntaxin 会导致驻留 ER 蛋白 calnexin 的获得延迟，并且含有毒力肺炎军团菌的吞噬体上 Rab1 的保留增强，表明这些 SNARE 蛋白参与液泡发生。质膜定位的 SNARE 蛋白 Syntaxin 2、Syntaxin 3、Syntaxin 4 和 SNAP23 定位于含有肺炎军团菌的液泡。发现 ER 定位的 SNARE 蛋白 Sec22b 与含有毒力肺炎军团菌的液泡上的质膜 SNARE 相互作用，但与含有非毒力肺炎军团菌突变体的液泡不相互作用。向含有毒力肺炎军团菌的质膜 SNARE 复合物中添加 alpha-SNAP 和 N-乙基马来酰亚胺敏感因子 (NSF) 会导致 Sec22b 解离，表明这些 SNARE 之间存在功能配对。因此，肺炎军团菌刺激质膜 t-SNARE 与 v-SNARE Sec22b 的非典型配对，以促进吞噬体与 ER 衍生小泡的融合。肺炎军团菌促进质膜 Syntaxin 和 Sec22b 配对的机制可以为分泌小泡如何在吞噬体成熟过程中提供被劫持的额外膜储备提供独特的见解。
