A CRISPR-Cas9 screen reveals a role for WD repeat-containing protein 81 (WDR81) in the entry of late penetrating viruses.
A CRISPR-Cas9 screen reveals a role for WD repeat-containing protein 81 (WDR81) in the entry of late penetrating viruses.
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DOI：
10.1371/journal.ppat.1010398
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发表时间：
2022-03
期刊：
影响因子：
6.7
通讯作者：
Danthi P
中科院分区：
文献类型：
--
作者：
Snyder AJ;Abad AT;Danthi P
许多不同病毒的成功感染需要它们被摄取到内体区室中。虽然一些病毒较早离开该隔室，但其他病毒必须到达晚期内体的降解酸性环境。哺乳动物正呼肠孤病毒（呼肠孤病毒）是一种这样的晚期穿透病毒。为了鉴定对呼肠孤病毒感染重要的宿主因子，我们进行了CRISPR-Cas9敲除（KO）筛选，该筛选靶向来自C57/BL 6小鼠胚胎的成纤维细胞中的20，000多个基因。我们鉴定了呼肠孤病毒诱导细胞死亡所需的七个基因（WDR 81、WDR 91、RAB 7、CCZ 1、CTSL、GNPTAB和SLC 35 A1）。值得注意的是，CRISPR介导的含WD重复序列的蛋白81（WDR 81）的KO使细胞对呼肠孤病毒感染具有抗性。通过用人WDR 81补充KO细胞来恢复对呼肠孤病毒感染的易感性。虽然WDR 81的缺乏不影响病毒附着效率或摄取到内体区室中进行初始拆卸，但它降低了病毒基因表达并减少了感染性病毒的产生。与WDR 81在影响内体成熟中的作用一致，WDR 81缺陷导致呼肠孤病毒颗粒在死端区室中积累。尽管WDR 81被VSV（水泡性口炎病毒）感染，其在早期阶段退出内体系统，但它是VSV-EBO GP（表达埃博拉病毒糖蛋白的VSV）所需的，其必须到达晚期内体以启动感染。这些结果揭示了WDR 81在促进通过晚期内体转运的病毒复制中的先前未被认识到的作用。病毒是专性细胞内寄生虫，需要许多宿主因子的贡献来完成病毒的生命周期。因此，宿主-病原体相互作用可以调节细胞死亡信号传导和病毒进入、复制、组装和外出。功能性遗传筛选是鉴定对建立感染重要的宿主因子的有用工具。这些信息也可以用来理解细胞生物学。值得注意的是，基因组规模的CRISPR-Cas9敲除筛选由于其特异性和宿主基因表达的丧失而具有鲁棒性。哺乳动物呼肠孤病毒是研究嗜神经性和嗜心脏性病毒致病机制的一个易处理的模型系统。使用CRISPR-Cas9筛选，我们将含有WD重复序列的蛋白81（WDR 81）鉴定为有效感染鼠细胞所需的宿主因子。WDR 81的消融阻断了病毒进入途径中的晚期步骤。此外，我们的工作表明WDR 81是表达埃博拉病毒糖蛋白的水泡性口炎病毒进入所必需的。
Successful initiation of infection by many different viruses requires their uptake into the endosomal compartment. While some viruses exit this compartment early, others must reach the degradative, acidic environment of the late endosome. Mammalian orthoreovirus (reovirus) is one such late penetrating virus. To identify host factors that are important for reovirus infection, we performed a CRISPR-Cas9 knockout (KO) screen that targets over 20,000 genes in fibroblasts derived from the embryos of C57/BL6 mice. We identified seven genes (WDR81, WDR91, RAB7, CCZ1, CTSL, GNPTAB, and SLC35A1) that were required for the induction of cell death by reovirus. Notably, CRISPR-mediated KO of WD repeat-containing protein 81 (WDR81) rendered cells resistant to reovirus infection. Susceptibility to reovirus infection was restored by complementing KO cells with human WDR81. Although the absence of WDR81 did not affect viral attachment efficiency or uptake into the endosomal compartments for initial disassembly, it reduced viral gene expression and diminished infectious virus production. Consistent with the role of WDR81 in impacting the maturation of endosomes, WDR81-deficiency led to the accumulation of reovirus particles in dead-end compartments. Though WDR81 was dispensable for infection by VSV (vesicular stomatitis virus), which exits the endosomal system at an early stage, it was required for VSV-EBO GP (VSV that expresses the Ebolavirus glycoprotein), which must reach the late endosome to initiate infection. These results reveal a previously unappreciated role for WDR81 in promoting the replication of viruses that transit through late endosomes. Viruses are obligate intracellular parasites that require the contributions of numerous host factors to complete the viral life cycle. Thus, the host-pathogen interaction can regulate cell death signaling and virus entry, replication, assembly, and egress. Functional genetic screens are useful tools to identify host factors that are important for establishing infection. Such information can also be used to understand cell biology. Notably, genome-scale CRISPR-Cas9 knockout screens are robust due to their specificity and the loss of host gene expression. Mammalian orthoreovirus (reovirus) is a tractable model system to investigate the pathogenesis of neurotropic and cardiotropic viruses. Using a CRISPR-Cas9 screen, we identified WD repeat-containing protein 81 (WDR81) as a host factor required for efficient reovirus infection of murine cells. Ablation of WDR81 blocked a late step in the viral entry pathway. Further, our work indicates that WDR81 is required for the entry of vesicular stomatitis virus that expresses the Ebolavirus glycoprotein.
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影响因子：
64.5
作者：
Daniloski Z;Jordan TX;Wessels HH;Hoagland DA;Kasela S;Legut M;Maniatis S;Mimitou EP;Lu L;Geller E;Danziger O;Rosenberg BR;Phatnani H;Smibert P;Lappalainen T;tenOever BR;Sanjana NE
通讯作者：
Sanjana NE
DOI：
10.1083/jcb.131.6.1435
发表时间：
1995-12
期刊：
The Journal of cell biology
影响因子：
--
作者：
Feng Y;Press B;Wandinger-Ness A
通讯作者：
Wandinger-Ness A
影响因子：
3.8
作者：
BORSA, J;MORASH, BD;SZEKELY, JG
通讯作者：
SZEKELY, JG
影响因子：
5.4
作者：
Connolly, JL;Rodgers, SE;Dermody, TS
通讯作者：
Dermody, TS
影响因子：
64.5
作者：
Barton, ES;Forrest, JC;Dermody, TS
通讯作者：
Dermody, TS