Brucella intracellular replication requires trafficking through the late endosomal/lysosomal compartment
Brucella intracellular replication requires trafficking through the late endosomal/lysosomal compartment
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DOI：
10.1111/j.1600-0854.2008.00718.x
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发表时间：
2008-05-01
期刊：
影响因子：
4.5
通讯作者：
Celli, Jean
中科院分区：
文献类型：
--
作者：
Starr, Tregei;Ng, Tony W.;Celli, Jean
在进入哺乳动物细胞后，细胞内病原体流产布鲁氏菌居住在一个膜结合的腔室中，即含布鲁氏菌液泡（BCV），其成熟由细菌控制，产生来自内质网（ER）的复制细胞器。在到达内质网之前，布鲁氏菌被认为通过抑制中间BCV与晚期内体和溶酶体的融合来确保其细胞内存活，尽管这些BCV呈酸性并积累溶酶体相关膜蛋白（LAMP-1）。在这里，我们使用共聚焦显微镜和活细胞成像进一步研究了中间bcv的性质。我们发现，bcv迅速获得了几种晚期内噬标记物，包括鸟苷三磷酸酶Rab7及其效应物rab -相互作用溶酶体蛋白（RILP），并且可以被传递到整个内噬途径或预加载到溶酶体的液相标记物所接近，这表明bcv与晚期内噬体和溶酶体相互作用。与此一致的是，中间bcv呈酸性，在感染后12小时内表现出蛋白水解活性。Rab7的显性阴性表达或RILP的过表达显著削弱了细菌将液泡转化为er来源的细胞器和复制的能力，这表明BCV的成熟需要与功能性晚期内体/溶酶体隔室相互作用。在表达显性阴性Rab7[T22N]的细胞中，bcv仍呈酸性，但与溶酶体的融合减少。综上所述，这些结果表明，bcv沿着内吞途径运输并与溶酶体融合，这种融合事件是bcv进一步成熟为er来源的复制细胞器所必需的。
Upon entry into mammalian cells, the intracellular pathogen Brucella abortus resides within a membrane-bound compartment, the Brucella-containing vacuole (BCV), the maturation of which is controlled by the bacterium to generate a replicative organelle derived from the endoplasmic reticulum (ER). Prior to reaching the ER, Brucella is believed to ensure its intracellular survival by inhibiting fusion of the intermediate BCV with late endosomes and lysosomes, although such BCVs are acidic and accumulate the lysosomal-associated membrane protein (LAMP-1). Here, we have further examined the nature of intermediate BCVs using confocal microscopy and live cell imaging. We show that BCVs rapidly acquire several late endocytic markers, including the guanosine triphosphatase Rab7 and its effector Rab-interacting lysosomal protein (RILP), and are accessible to fluid-phase markers either delivered to the whole endocytic pathway or preloaded to lysosomes, indicating that BCVs interact with late endosomes and lysosomes. Consistently, intermediate BCVs are acidic and display proteolytic activity up to 12 h post-infection. Expression of dominant-negative Rab7 or overexpression of RILP significantly impaired the ability of bacteria to convert their vacuole into an ER-derived organelle and replicate, indicating that BCV maturation requires interactions with functional late endosomal/lysosomal compartments. In cells expressing dominant-negative Rab7[T22N], BCVs remained acidic, yet displayed decreased fusion with lysosomes. Taken together, these results demonstrate that BCVs traffic along the endocytic pathway and fuse with lysosomes, and such fusion events are required for further maturation of BCVs into an ER-derived replicative organelle.