Transbilayer Movement of Sphingomyelin Precedes Catastrophic Breakage of Enterobacteria-Containing Vacuoles
Transbilayer Movement of Sphingomyelin Precedes Catastrophic Breakage of Enterobacteria-Containing Vacuoles
复制标题
DOI：
10.1016/j.cub.2020.05.083
复制
发表时间：
2020-08-03
期刊：
影响因子：
9.2
通讯作者：
Randow, Felix
中科院分区：
文献类型：
--
作者：
Ellison, Cara J.;Kukulski, Wanda;Randow, Felix
病原菌通过摄取含有细菌的空泡(BCV)和随后的空泡膜破裂进入宿主细胞的胞浆[1]。细菌入侵者可以直接通过细菌配体的细胞质模式识别受体直接感知，也可以通过危险受体间接感知，这些受体结合了在异常情况下显示的宿主分子，例如受损BCV上的多糖[2-4]。与革兰氏阳性菌单核细胞增生性李斯特菌造成的损害不同，革兰氏阴性病原体如福氏志贺氏菌或鼠伤寒沙门氏菌导致的BCV破裂仍然不完全清楚[5，6]。后者可能直接导致膜损伤，当它们的3型分泌针插入宿主膜时，或通过转位的细菌效应蛋白间接造成膜损伤[7-9]。在这里，我们报告了BCV膜管腔小叶中的一种丰富的脂类神经鞘磷脂，通常不存在于胞浆中，暴露于胞浆中，作为革兰氏阴性菌破裂BCV的早期预测标志。Tomonvisor亚细胞鞘磷脂分布，我们从Lysenin(一种来自Eisenia Fetida的鞘磷脂特异性毒素)中产生了一个活的SphingMarelin报告程序[10，11]。利用超分辨率活体成像和相关的光学和电子显微镜(CLEM)，我们发现BCV的破裂经历了两个不同的连续阶段：首先，鞘磷脂逐渐转移到BCV的胞浆小叶中，紧随其后的是胞液中的葡聚糖暴露，它招募Galectin-8，表明细菌进入胞浆。因此，暴露在BCV上的鞘磷脂可能会作为早期危险信号，提醒细胞注意即将到来的细菌入侵。
Pathogenic bacteria enter the cytosol of host cells through uptake into bacteria-containing vacuoles (BCVs) and subsequent rupture of the vacuolar membrane [1]. Bacterial invaders are sensed either directly, through cytosolic pattern-recognition receptors specific for bacterial ligands, or indirectly, through danger receptors that bind host molecules displayed in an abnormal context, for example, glycans on damaged BCVs [2-4]. In contrast to damage caused by Listeria monocytogenes, a Gram-positive bacterium, BCV rupture by Gram-negative pathogens such as Shigella flexneri or Salmonella Typhimurium remains incompletely understood [5, 6]. The latter may cause membrane damage directly, when inserting their Type Three Secretion needles into host membranes, or indirectly through translocated bacterial effector proteins [7-9]. Here, we report that sphingomyelin, an abundant lipid of the luminal leaflet of BCV membranes, and normally absent from the cytosol, becomes exposed to the cytosol as an early predictive marker of BCV rupture by Gram-negative bacteria. Tomonitor subcellular sphingomyelin distribution, we generated a live sphingomyelin reporter from Lysenin, a sphingomyelin-specific toxin from the earthworm Eisenia fetida [10, 11]. Using super resolution live imaging and correlative light and electron microscopy (CLEM), we discovered that BCV rupture proceeds through two distinct successive stages: first, sphingomyelin is gradually translocated into the cytosolic leaflet of the BCV, invariably followed by cytosolic exposure of glycans, which recruit galectin-8, indicating bacterial entry into the cytosol. Exposure of sphingomyelin on BCVs may therefore act as an early danger signal alerting the cell to imminent bacterial invasion.