单分子视踪技术揭示SARS-CoV-2病毒利用动态的丝状伪足入侵细胞  

Single particle tracking reveals SARS-CoV-2 regulating and utilizing dynamic filopodia for viral invasion

作　　者：张悦 张晓玮 李钟艺 赵唯淞 杨慧 赵双双 汤代骄 张倩 李总红 刘会生 李浩宇 李博 Pekka Lappalainen 徐涛 崔宗强 酒亚明 Yue Zhang;Xiaowei Zhang;Zhongyi Li;Weisong Zhao;Hui Yang;Shuangshuang Zhao;Daijiao Tang;Qian Zhang;Zonghong Li;Huisheng Liu;Haoyu Li;Bo Li;Pekka Lappalainen;Tao Xu;Zongqiang Cui;Yaming Jiu(Unit of Cell Biology and Imaging Study of Pathogen Host Interaction,The Center for Microbes,Development and Health,Key Laboratory of Molecular Virology and Immunology,Shanghai Institute of Immunity and Infection,Chinese Academy of Sciences,Shanghai 200031,China;University of Chinese Academy of Sciences,Beijing 100049,China;State Key Laboratory of Virology,Wuhan Institute of Virology,Center for Biosafety Mega-Science,Chinese Academy of Sciences,Wuhan 430071,China;Institute of Biomechanics and Medical Engineering,Applied Mechanics Laboratory,Department of Engineering Mechanics,Tsinghua University,Beijing 100084,China;Innovation Photonics and Imaging Center,School of Instrumentation Science and Engineering,Harbin Institute of Technology,Harbin 150080,China;Guangzhou Laboratory,Guangzhou 510005,China;Institute of Biotechnology and Helsinki Institute of Life Science,University of Helsinki,Helsinki 00014,Finland)

机构地区：[1]Unit of Cell Biology and Imaging Study of Pathogen Host Interaction,The Center for Microbes,Development and Health,Key Laboratory of Molecular Virology and Immunology,Shanghai Institute of Immunity and Infection,Chinese Academy of Sciences,Shanghai 200031,China [2]University of Chinese Academy of Sciences,Beijing 100049,China [3]State Key Laboratory of Virology,Wuhan Institute of Virology,Center for Biosafety Mega-Science,Chinese Academy of Sciences,Wuhan 430071,China [4]Institute of Biomechanics and Medical Engineering,Applied Mechanics Laboratory,Department of Engineering Mechanics,Tsinghua University,Beijing 100084,China [5]Innovation Photonics and Imaging Center,School of Instrumentation Science and Engineering,Harbin Institute of Technology,Harbin 150080,China [6]Guangzhou Laboratory,Guangzhou 510005,China [7]Institute of Biotechnology and Helsinki Institute of Life Science,University of Helsinki,Helsinki 00014,Finland

出　　处：《Science Bulletin》2023年第19期2210-2224,M0004,共16页科学通报（英文版）

基　　金：the Key Research and Development Program,Ministry of Science and Technology of China(2022YFC2303502 and 2021YFC2300204);the Chinese Academy of Sciences-Vice Presidency Science and Technology Silk Road Science Fund(GJHZ2021138);the National Natural Science Foundation of China(32222022,92054104,31970660,31925025,and 22293033);the Strategic Priority Research Program of the Chinese Academy of Sciences(XDB29050201);the R&D Program of Guangzhou Laboratory(SRPG22-002)。

摘　　要：SARS-CoV-2病毒侵袭细胞的分子机制已被初步探索,但关于其如何调节亚细胞结构重塑从而侵袭多种器官及细胞类型尚不清楚.本文利用活细胞实时成像技术揭示了SARS-CoV-2病毒颗粒通过宿主细胞丝状伪足进入靶细胞的动态过程.利用荧光标记的SARS-CoV-2病毒样颗粒(VLP)和稀疏去卷积算法成像技术,发现VLP利用丝状伪足以“冲浪”和“抓取”两种模式到达入侵点,以避免病毒在细胞膜上随机搜索入侵位点,此外,结合力学模拟实验,阐明了病毒诱导丝状伪足的形成和丝状伪足的回缩速度分别取决于细胞骨架动力学和病毒重力引起的底物表面摩擦阻力.进一步研究发现SARS-CoV-2通过丝状伪足进入细胞的过程依赖于小G蛋白Cdc42(细胞分裂周期蛋白42)活性和肌动蛋白相关蛋白fasin(肌动蛋白结合蛋白)、formin(成核蛋白)和Arp2/3(肌动蛋白相关蛋白2/3复合物).综上所述,本文结果强调了SARS-CoV-2感染能对宿主细胞的微丝骨架进行时空调节,使细胞形成丝状伪足作为病毒进入的“高速轨道”,丝状伪足形成相关蛋白及其信号通路可作为潜在的抗病毒靶标.Although severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)entry mechanism has been explored,little is known about how SARS-CoV-2 regulates the subcellular structural remodeling to invade multiple organs and cell types.Here,we unveil how SARS-CoV-2 boosts and utilizes filopodia to enter the target cells by real-time imaging.Using SARS-CoV-2 single virus-like particle(VLP)tracking in live cells and sparse deconvolution algorithm,we uncover that VLPs utilize filopodia to reach the entry site in two patterns,“surfing”and“grabbing”,which avoid the virus from randomly searching on the plasma membrane.Moreover,combining mechanical simulation,we elucidate that the formation of virus-induced filopodia and the retraction speed of filopodia depend on cytoskeleton dynamics and friction resistance at the substrate surface caused by loading-virus gravity,respectively.Further,we discover that the entry process of SARS-CoV-2 via filopodia depends on Cdc42 activity and actin-associated proteins fascin,formin,and Arp2/3.Together,our results highlight that the spatial–temporal regulation of actin cytoskeleton by SARS-CoV-2 infection makes filopodia as a highway for virus entry and potentiates it as an antiviral target.

关 键 词：力学模拟 亚细胞结构 随机搜索 单分子 病毒样颗粒 表面摩擦阻力 病毒侵袭 病毒颗粒 

分 类 号：R373[医药卫生—病原生物学]