Department of Molecular and Cell Biology, Centro Nacional de Biotecnología, Consejo Superior de Investigaciones Científicas, Campus Universidad Autónoma, Madrid 28049, Spain.
Cell Microbiol. 2011 Jun;13(6):846-58. doi: 10.1111/j.1462-5822.2011.01583.x. Epub 2011 Mar 4.
As an enveloped virus, replication of human cytomegalovirus (HCMV) is dependent on interaction with cellular membrane systems. Its final envelopment occurs into intracellular membranes prior to its secretion. However the mechanisms underlying these processes are poorly understood. Here, we show that HCMV infection induces expression of the soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) syntaxin 3 (STX3), a component of the cellular machinery for membrane fusion. STX3 was located at the plasma membrane and at the assembly site where it was found associated with virus wrapping membranes by immunogold labelling. Depletion of STX3 using RNA interference reduced HCMV production, while expression of a STX3 construct resistant to RNAi inhibition enhanced virus production. Ultrastructural examination of the assembly site in HCMV-infected STX3-depleted cells showed fewer mature virions and more viruses undergoing final envelopment. In contrast, silencing of STX3 did not affect herpes simplex virus type 1 production. The mechanism through which STX3 affected HCMV morphogenesis likely involved late endosomes/lysosomes since STX3 depletion reduced the expression of lysosomal membrane glycoproteins. Our results demonstrate a function for STX3 in HCMV morphogenesis, and unravel a new role for this SNARE protein in late endosomes/lysosomes compartments.
作为一种包膜病毒，人巨细胞病毒（HCMV）的复制依赖于与细胞膜系统的相互作用。其最终的包膜发生在其分泌之前，进入细胞内膜。然而，这些过程的机制还知之甚少。在这里，我们表明 HCMV 感染诱导可溶性 N-乙基马来酰亚胺敏感因子附着蛋白受体（SNARE）突触融合蛋白 3（STX3）的表达，这是细胞融合的膜融合机制的组成部分。STX3 位于质膜上，在组装部位被发现与病毒包裹膜结合，通过免疫金标记。使用 RNA 干扰耗尽 STX3 会降低 HCMV 的产生，而表达对 RNAi 抑制有抗性的 STX3 构建体则增强了病毒的产生。在 STX3 耗尽的 HCMV 感染细胞的组装部位的超微结构检查显示成熟病毒颗粒较少，更多的病毒正在进行最终包膜。相比之下，沉默 STX3 并不影响单纯疱疹病毒 1 的产生。STX3 影响 HCMV 形态发生的机制可能涉及晚期内体/溶酶体，因为 STX3 耗竭降低了溶酶体膜糖蛋白的表达。我们的结果表明 STX3 在 HCMV 形态发生中的功能，并揭示了这种 SNARE 蛋白在晚期内体/溶酶体区室中的新作用。
