We hypothesize that intracellular trafficking pathways are altered in chlamydial infected cells to maximize the ability of Chlamydia to scavenge nutrients while not overtly stressing the host cell. Previous data demonstrated the importance of two eukaryotic SNARE proteins, VAMP4 and syntaxin 10 (St × 10), in chlamydial growth and development. Although, the mechanism for these effects is still unknown. To interrogate whether chlamydial infection altered these proteins' networks, we created BirA*-VAMP4 and BirA*-St × 10 fusion constructs to use the BioID proximity labeling system. While we identified a novel eukaryotic protein-protein interaction between St × 10 and VAPB, we also identified caveats in using the BioID system to study the impact of infection by an obligate intracellular pathogen on SNARE protein networks. The addition of the BirA* altered the localization of VAMP4 and St × 10 during infection with C. trachomatis serovars L2 and D and C. burnetii Nine Mile Phase II. We also discovered that BirA* traffics to and biotinylates Coxiella-containing vacuoles and, in general, has a propensity for labeling membrane or membrane-associated proteins. While the BioID system identified a novel association for St × 10, it is not a reliable methodology to examine intracellular trafficking pathway dynamics during infection with intracellular pathogens.

中文翻译：

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转移蛋白质组：使用 BioID 邻近标记系统研究细胞内病原体感染期间 SNARE 蛋白运输的局限性

我们假设衣原体感染细胞中的细胞内运输途径发生了改变，以最大限度地提高衣原体清除营养物质的能力，同时不会对宿主细胞造成明显的压力。先前的数据证明了两种真核 SNARE 蛋白 VAMP4 和突触蛋白 10 (St × 10) 在衣原体生长和发育中的重要性。虽然，这些影响的机制仍然未知。为了询问衣原体感染是否改变了这些蛋白质的网络，我们创建了 BirA*-VAMP4 和 BirA*-St × 10 融合构建体以使用 BioID 邻近标记系统。虽然我们确定了 St × 10 和 VAPB 之间的新型真核蛋白质-蛋白质相互作用，但我们还发现了使用 BioID 系统研究专性细胞内病原体感染对 SNARE 蛋白质网络的影响的注意事项。添加 BirA* 在感染沙眼衣原体 L2 和 D 和 C. burnetii 九英里二期期间改变了 VAMP4 和 St × 10 的定位。我们还发现 BirA* 运输并生物素化含有 Coxiella 的液泡，并且通常具有标记膜或膜相关蛋白的倾向。虽然 BioID 系统确定了 St × 10 的新关联，但它不是一种可靠的方法来检查细胞内病原体感染期间的细胞内运输途径动态。