Novel role for ESCRT-III component CHMP4C in the integrity of the endocytic network utilized for herpes simplex virus envelopment
Novel role for ESCRT-III component CHMP4C in the integrity of the endocytic network utilized for herpes simplex virus envelopment
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ESCRT-III 组分 CHMP4C 在用于单纯疱疹病毒包膜的内吞网络完整性中的新作用
DOI：
10.1101/2020.08.19.258558
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发表时间：
2020
期刊：
影响因子：
--
通讯作者：
Russell T
中科院分区：
文献类型：
--
作者：
Russell T
包膜病毒利用细胞运输途径进行形态发生，为开发新的抗病毒疗法提供了潜在的范围。我们以前已经表明，单纯疱疹病毒1型（HSV 1）利用回收内吞膜作为其包膜的来源，在一个过程中涉及四个Rab GTP酶。为了鉴定参与HSV 1表达的新因子，我们筛选了针对80多种人类贩运蛋白的小干扰RNA（siRNA）文库，所述蛋白包括外壳蛋白、衔接蛋白、融合因子、裂变因子和Rab效应子。11个因子的耗竭使病毒产量降低20至100倍，包括3个早期分泌途径蛋白、4个晚期分泌途径蛋白和4个内吞途径蛋白，其中3个是膜分裂因子。选择11个靶标中的5个用于病毒感染中的进一步分析，其中发现仅1个，即裂变因子CHMP 4C，而不是CHMP 4A或CHMP 4 B旁系同源物的缺失，在形态发生的最后阶段减少了病毒产生。CHMP 4C耗尽，HSV 1感染的细胞的超微结构和共聚焦显微镜显示内吞膜的积累;广泛的再循环，转铁蛋白受体阳性核内体异常分裂的指示性微管;和在病毒灭活失败。未检测到对晚期内吞途径的影响，而外源性CHMP 4C显示定位于再循环内体。两者合计，这些数据揭示了一个新的角色CHMP 4C裂变因子的回收内体网络的完整性，这已经通过HSV 1对这些膜的依赖性为收购其envelopes. IMPORTANCE细胞运输途径中发挥了重要作用的分泌和膜生物合成。有包膜病毒利用这些途径将其膜蛋白引导至包膜位点，因此是解开运输因子微妙活动的有力工具，有可能精确定位治疗靶点。使用敏感的病毒生产的生物学读数，已经筛选了80多个涉及不同和定义不清的细胞过程的运输因子，以参与HSV 1的复杂过程。在11个潜在的靶点中，CHMP 4C是细胞周期终止检查点的关键组分，它是病毒在内吞小管中包裹过程所必需的。在没有CHMP 4C的情况下，再循环的内吞膜在感染的细胞中未能经历断裂，导致膜和未包裹的病毒的暂时性坏死和积累。这些数据揭示了一个新的作用，这一重要的细胞因子在回收内吞膜的生物发生。
Enveloped viruses exploit cellular trafficking pathways for their morphogenesis, providing potential scope for the development of new antiviral therapies. We have previously shown that herpes simplex virus 1 (HSV1) utilizes recycling endocytic membranes as the source of its envelope, in a process involving four Rab GTPases. To identify novel factors involved in HSV1 envelopment, we have screened a small interfering RNA (siRNA) library targeting over 80 human trafficking proteins, including coat proteins, adaptor proteins, fusion factors, fission factors, and Rab effectors. The depletion of 11 factors reduced virus yields by 20- to 100-fold, including three early secretory pathway proteins, four late secretory pathway proteins, and four endocytic pathway proteins, three of which are membrane fission factors. Five of the 11 targets were chosen for further analysis in virus infection, where it was found that the absence of only 1, the fission factor CHMP4C, but not the CHMP4A or CHMP4B paralogues, reduced virus production at the final stage of morphogenesis. Ultrastructural and confocal microscopy of CHMP4C-depleted, HSV1-infected cells showed an accumulation of endocytic membranes; extensive tubulation of recycling, transferrin receptor-positive endosomes indicative of aberrant fission; and a failure in virus envelopment. No effect on the late endocytic pathway was detected, while exogenous CHMP4C was shown to localize to recycling endosomes. Taken together, these data reveal a novel role for the CHMP4C fission factor in the integrity of the recycling endosomal network, which has been unveiled through the dependence of HSV1 on these membranes for the acquisition of their envelopes.IMPORTANCECellular transport pathways play a fundamental role in secretion and membrane biogenesis. Enveloped viruses exploit these pathways to direct their membrane proteins to sites of envelopment and, as such, are powerful tools for unraveling subtle activities of trafficking factors, potentially pinpointing therapeutic targets. Using the sensitive biological readout of virus production, over 80 trafficking factors involved in diverse and poorly defined cellular processes have been screened for involvement in the complex process of HSV1 envelopment. Out of 11 potential targets, CHMP4C, a key component in the cell cycle abscission checkpoint, stood out as being required for the process of virus wrapping in endocytic tubules, where it localized. In the absence of CHMP4C, recycling endocytic membranes failed to undergo scission in infected cells, causing transient tubulation and accumulation of membranes and unwrapped virus. These data reveal a new role for this important cellular factor in the biogenesis of recycling endocytic membranes.