Binding of SycH chaperone to YscM1 and YscM2 activates effector yop expression in Yersinia enterocolitica
Binding of SycH chaperone to YscM1 and YscM2 activates effector yop expression in Yersinia enterocolitica
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DOI：
10.1128/jb.186.3.829-841.2004
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发表时间：
2004-02-01
影响因子：
3.2
通讯作者：
Schneewind, O
中科院分区：
文献类型：
--
作者：
Cambronne, ED;Sorg, JA;Schneewind, O
小肠结肠炎耶尔森氏菌通过 III 型途径转运 YscM1 和 YscM2，这是建立细菌感染所需的机制。在接触宿主细胞之前，YscM1 和 YscM2 发挥转录后调节作用，抑制效应 yop 基因的表达，该基因编码通过 III 型途径进入巨噬细胞细胞质的毒力因子。据预测，抑制的缓解是通过 YscM1 和 YscM2 向细胞外介质中的 III 型分泌而发生的，从而导致细菌细胞质中调节分子的耗尽。使用洋地黄皂苷分级分离和耶尔森氏菌感染细胞中 FlAsH 标记多肽的荧光显微镜，我们将 YscM1 和 YscM2 定位在宿主细胞细胞质内。 YscM1 和 YscM2 的 III 型注射需要 SycH 伴侣。 YscM1 和 YscM2 与新霉素磷酸转移酶报告基因的 C 端融合表达揭示了在 SycH 不存在的情况下调节活性和分泌所需的序列。 SycH 和谷胱甘肽 S-转移酶 (GST)-YscM1 或 GST-YscM2（不能由 III 型装置转运的杂合 GST 变体）的共表达也缓解了对 Yop 合成的抑制。 GST-SycH 与 YscM1 和 YscM2 结合并激活效应子 yop 表达，而不启动结合的调节分子进入 III 型途径。此外，YscM1、YscM2 和 SycH 对 yop 表达的调节显示出独立于调节分泌的因素的作用，并且凝胶过滤色谱法揭示了在 Yop 合成受到抑制的条件下不与 SycH 结合的 YscM1 和 YscM2 群体。总而言之，这些结果表明，YscM1 和 YscM2 介导的抑制可以通过在 III 型注射到宿主细胞之前与细胞质伴侣 SycH 结合而缓解。
Yersinia enterocolitica transports YscM1 and YscM2 via the type III pathway, a mechanism that is required for the establishment of bacterial infections. Prior to host cell contact, YscM1 and YscM2 exert posttranscriptional regulation to inhibit expression of effector yop genes, which encode virulence factors that travel the type III pathway into the cytoplasm of macrophages. Relief from repression has been predicted to occur via the type III secretion of YscM1 and YscM2 into the extracellular medium, resulting in the depletion of regulatory molecules from the bacterial cytoplasm. Using digitonin fractionation and fluorescence microscopy of FlAsH-labeled polypeptides in Yersinia-infected cells, we have localized YscM1 and YscM2 within the host cell cytoplasm. Type III injection of YscM1 and YscM2 required the SycH chaperone. Expression of C-terminal fusions of YscM1 and YscM2 to the neomycin phosphotransferase reporter revealed sequences required for regulatory activity and for secretion in the absence of SycH. Coexpression of SycH and glutathione S-transferase (GST)-YscM1 or GST-YscM2, hybrid GST variants that cannot be transported by the type III apparatus, also relieved repression of Yop synthesis. GST-SycH bound to YscM1 and YscM2 and activated effector yop expression without initiation of the bound regulatory molecules into the type III pathway. Further, regulation of yop expression by YscM1, YscM2, and SycH is shown to act independently of factors that regulate secretion, and gel filtration chromotography revealed populations of YscM1 and YscM2 that are not bound to SycH under conditions where Yop synthesis is repressed. Taken together, these results suggest that YscM1- and YscM2-mediated repression may be relieved through binding to the cytoplasmic chaperone SycH prior to their type III injection into host cells.