Dissecting the Cell Entry Pathway of Baculovirus by Single-Particle Tracking and Quantitative Electron Microscopic Analysis
Dissecting the Cell Entry Pathway of Baculovirus by Single-Particle Tracking and Quantitative Electron Microscopic Analysis
复制标题
通过单粒子追踪和定量电子显微镜分析剖析杆状病毒的细胞进入途径
DOI：
10.1128/jvi.00033-19
复制
发表时间：
2019-04-01
影响因子：
5.4
通讯作者：
Sun, Xiulian
中科院分区：
文献类型：
--
作者：
Qin, Fujun;Xu, Congrui;Sun, Xiulian
杆状病毒广泛用作环境友好型杀虫剂、蛋白质表达系统和潜在的哺乳动物基因传递载体。尽管具有重要的应用价值，但人们对杆状病毒的细胞进入和内吞运输途径知之甚少。在这项研究中，我们证明甲杆病毒 AcMNPV 表现出依赖于肌动蛋白和微管的核衣壳释放运输，主要从早期内涵体内释放。与哺乳动物细胞中的 AcMNPV 转导相反，其在宿主昆虫细胞中的感染是通过肌动蛋白聚合进行内化和早期内体中进行内吞运输的微管促进的，这意味着 AcMNPV 在细胞骨架网络的需求中表现出细胞类型特异性。此外，微管的实验性解聚损害了早期内体扩大以外的感染进展。这是第一项在单颗粒水平上剖析杆状病毒进入宿主细胞的细胞途径的研究，这增进了我们对杆状病毒进入细胞早期步骤的理解。摘要 苜蓿银纹夜蛾多核多角体病毒（AcMNPV）的出芽病毒主要通过网格蛋白介导的内吞作用感染昆虫细胞。然而，AcMNPV 进入细胞的途径仍不清楚。在本研究中，通过使用基于群体的单病毒追踪分析和电子显微镜，我们研究了 AcMNPV 的内化、融合行为和内吞运输。肌动蛋白聚合和动力促进 AcMNPV 内化到宿主昆虫细胞中。掺入早期内涵体后，AcMNPV 包膜与早期内涵体的膜融合，允许核衣壳释放到细胞质中。微管参与含病毒内体的双向和长程运输。此外，微管解聚降低了携带病毒的早期内体的运动性，从而损害了扩大的早期内体之外的感染进展。这些发现表明，肌动蛋白聚合以动力蛋白依赖性方式促进了 AcMNPV 内化，并且核衣壳释放以微管依赖性方式发生在早期内涵体中。这项研究提供了对 AcMNPV 感染的机制和动力学见解，并增强了我们对杆状病毒感染途径的理解。重要性杆状病毒被广泛用作环境友好型杀虫剂、蛋白质表达系统和潜在的哺乳动物基因传递载体。尽管具有重要的应用价值，但人们对杆状病毒的细胞进入和内吞运输途径知之甚少。在这项研究中，我们证明甲杆病毒 AcMNPV 表现出依赖于肌动蛋白和微管的核衣壳释放运输，主要从早期内涵体内释放。与哺乳动物细胞中的 AcMNPV 转导相反，其在宿主昆虫细胞中的感染是通过肌动蛋白聚合进行内化和早期内体中进行内吞运输的微管促进的，这意味着 AcMNPV 在细胞骨架网络的需求中表现出细胞类型特异性。此外，微管的实验性解聚损害了早期内体扩大以外的感染进展。这是第一项在单颗粒水平上剖析杆状病毒进入宿主细胞的细胞途径的研究，这增进了我们对杆状病毒进入细胞早期步骤的理解。
Baculoviruses are used widely as environmentally benign pesticides, protein expression systems, and potential mammalian gene delivery vectors. Despite the significant application value, little is known about the cell entry and endocytic trafficking pathways of baculoviruses. In this study, we demonstrated that the alphabaculovirus AcMNPV exhibited actin- and microtubule-dependent transport for nucleocapsid release predominantly from within early endosomes. In contrast to AcMNPV transduction in mammalian cells, its infection in host insect cells is facilitated by actin polymerization for internalization and microtubules for endocytic trafficking within early endosomes, implying that AcMNPV exhibits cell type specificity in the requirement of the cytoskeleton network. In addition, experimental depolymerization of microtubules impaired the progression of infection beyond enlarged early endosomes. This is the first study that dissects the cell entry pathway of baculoviruses in host cells at the single-particle level, which advances our understanding of the early steps of baculovirus entry. ABSTRACT The budded virus of Autographa californica multiple nucleopolyhedrovirus (AcMNPV) infects insect cells through mainly clathrin-mediated endocytosis. However, the cell entry pathway of AcMNPV remains unclear. In this study, by using population-based analysis of single-virus tracking and electron microscopy, we investigated the internalization, fusion behavior, and endocytic trafficking of AcMNPV. AcMNPV internalization into host insect cells was facilitated by actin polymerization and dynamin. After incorporation into early endosomes, the AcMNPV envelope fused with the membranes of early endosome, allowing for nucleocapsid release into the cytoplasm. Microtubules were implicated in the bidirectional and long-range transport of virus-containing endosomes. In addition, microtubule depolymerization reduced the motility of virus-bearing early endosomes, impairing the progression of infection beyond enlarged early endosomes. These findings demonstrated that AcMNPV internalization was facilitated by actin polymerization in a dynamin-dependent manner, and nucleocapsid release occurred in early endosomes in a microtubule-dependent manner. This study provides mechanistic and kinetic insights into AcMNPV infection and enhance our understanding of the infection pathway of baculoviruses. IMPORTANCE Baculoviruses are used widely as environmentally benign pesticides, protein expression systems, and potential mammalian gene delivery vectors. Despite the significant application value, little is known about the cell entry and endocytic trafficking pathways of baculoviruses. In this study, we demonstrated that the alphabaculovirus AcMNPV exhibited actin- and microtubule-dependent transport for nucleocapsid release predominantly from within early endosomes. In contrast to AcMNPV transduction in mammalian cells, its infection in host insect cells is facilitated by actin polymerization for internalization and microtubules for endocytic trafficking within early endosomes, implying that AcMNPV exhibits cell type specificity in the requirement of the cytoskeleton network. In addition, experimental depolymerization of microtubules impaired the progression of infection beyond enlarged early endosomes. This is the first study that dissects the cell entry pathway of baculoviruses in host cells at the single-particle level, which advances our understanding of the early steps of baculovirus entry.