Li Xiang, Gulbins Erich, Zhang Yang
Department of Pharmacology & Toxicology, Medical College of Virginia Campus, Virginia Commonwealth University, Richmond, VA 23298-0613 , USA.
Cell Physiol Biochem. 2012;30(4):815-26. doi: 10.1159/000341460. Epub 2012 Aug 10.
Recent studies demonstrate that rapid translocation of the acid sphingomyelinase (ASM), a lysosomal hydrolase, to the outer leaflet of the cell membrane and concomitant release of ceramide constitute a common cellular signaling cascade to various stimuli including CD95 ligation, UV-irradiation, bacterial and viral infections. Reactive oxygen species (ROS) were shown to play a crucial role in regulating this signaling cascade at least for some bacterial infections and UV-irradiation. However, the precise role of ROS for regulation of ASM is unknown. Here, by confocal microscopy and flow cytometry analysis, we demonstrate that hydrogen peroxide (H(2)O(2)), a primary form of ROS in mammalian cells, induces very rapid translocation of ASM and formation of ceramide-enriched membrane platforms in the plasma membrane of Jurkat T cells. In parallel, H(2)O(2) triggers lysosome trafficking and fusion with the plasma membrane, i.e. lysosome exocytosis, as detected by exposure of a lysosome-associated protein, LAMP1. Depletion of intracellular Ca(2+) by cell permeable EGTA-AM inhibits H(2)O(2)-induced lysosome exocytosis, ASM translocation and formation of ceramide-enriched platforms. Pharmacological inhibition or genetic deficiency of ASM did not affect H(2)O(2)-induced lysosome exocytosis. These results indicate that ROS-induced membrane translocation of ASM is mediated by exocytosis of lysosomes, which is dependent on intracellular Ca(2+) release.
最近的研究表明，酸性鞘磷脂酶（一种溶酶体水解酶）迅速转位至细胞膜外小叶并伴随神经酰胺的释放，构成了一种常见的细胞信号级联反应，以应对包括CD95连接、紫外线照射、细菌和病毒感染在内的各种刺激。活性氧（ROS）至少在某些细菌感染和紫外线照射中，被证明在调节这种信号级联反应中起关键作用。然而，ROS对酸性鞘磷脂酶调节的确切作用尚不清楚。在此，通过共聚焦显微镜和流式细胞术分析，我们证明过氧化氢（H₂O₂），哺乳动物细胞中ROS的主要形式，可诱导酸性鞘磷脂酶在Jurkat T细胞的质膜中非常迅速地转位，并形成富含神经酰胺的膜平台。同时，H₂O₂触发溶酶体运输并与质膜融合，即溶酶体胞吐作用，这可通过溶酶体相关蛋白LAMP1的暴露检测到。细胞可渗透的EGTA-AM耗尽细胞内Ca²⁺可抑制H₂O₂诱导的溶酶体胞吐作用、酸性鞘磷脂酶转位以及富含神经酰胺平台的形成。酸性鞘磷脂酶的药理学抑制或基因缺陷并不影响H₂O₂诱导的溶酶体胞吐作用。这些结果表明，ROS诱导的酸性鞘磷脂酶膜转位是由溶酶体胞吐作用介导的，这依赖于细胞内Ca²⁺的释放。
