Campanella Michelangelo, de Jong Arjan S, Lanke Kjerstin W H, Melchers Willem J G, Willems Peter H G M, Pinton Paolo, Rizzuto Rosario, van Kuppeveld Frank J M
Department of Experimental and Diagnostic Medicine, Section of General Pathology and Center for the Study of Inflammatory Diseases, Via Borsari 46, I-44100 Ferrara, Italy.
J Biol Chem. 2004 Apr 30;279(18):18440-50. doi: 10.1074/jbc.M309494200. Epub 2004 Feb 19.
Enteroviruses, small cytolytic RNA viruses, confer an antiapoptotic state to infected cells in order to suppress infection-limiting apoptotic host cell responses. This antiapoptotic state also lends protection against cell death induced by metabolic inhibitors like actinomycin D and cycloheximide. The identity of the viral antiapoptotic protein and the underlying mechanism are unknown. Here, we provide evidence that the coxsackievirus 2B protein modulates apoptosis by manipulating intracellular Ca(2+) homeostasis. Using fluorescent Ca(2+) indicators and organelle-targeted aequorins, we demonstrate that ectopic expression of 2B in HeLa cells decreases the Ca(2+) content of both the endoplasmic reticulum and the Golgi, resulting in down-regulation of Ca(2+) signaling between these stores and the mitochondria, and increases the influx of extracellular Ca(2+). In our studies of the physiological importance of the 2B-induced alterations in Ca(2+) signaling, we found that the expression of 2B suppressed caspase activation and apoptotic cell death induced by various stimuli, including actinomycin D and cycloheximide. Mutants of 2B that were defective in reducing the Ca(2+) content of the stores failed to suppress apoptosis. These data implicate a functional role of the perturbation of intracellular Ca(2+) compartmentalization in the enteroviral strategy to suppress intrinsic apoptotic host cell responses. The putative down-regulation of an endoplasmic reticulum-dependent apoptotic pathway is discussed.
肠道病毒是小型溶细胞性RNA病毒，它赋予受感染细胞一种抗凋亡状态，以抑制限制感染的宿主细胞凋亡反应。这种抗凋亡状态还能保护细胞免受放线菌素D和环己酰亚胺等代谢抑制剂诱导的细胞死亡。病毒抗凋亡蛋白的身份及其潜在机制尚不清楚。在此，我们提供证据表明柯萨奇病毒2B蛋白通过操纵细胞内Ca(2+)稳态来调节细胞凋亡。使用荧光Ca(2+)指示剂和细胞器靶向水母发光蛋白，我们证明在HeLa细胞中异位表达2B会降低内质网和高尔基体的Ca(2+)含量，导致这些储存库与线粒体之间的Ca(2+)信号下调，并增加细胞外Ca(2+)的内流。在我们对2B诱导的Ca(2+)信号改变的生理重要性的研究中，我们发现2B的表达抑制了由各种刺激（包括放线菌素D和环己酰亚胺）诱导的半胱天冬酶激活和凋亡细胞死亡。在降低储存库Ca(2+)含量方面存在缺陷的2B突变体无法抑制细胞凋亡。这些数据表明细胞内Ca(2+)区室化的扰动在肠道病毒抑制宿主细胞固有凋亡反应的策略中具有功能性作用。本文还讨论了内质网依赖性凋亡途径可能的下调情况。
