SARS-CoV-2入侵细胞的研究进展

新型冠状病毒(SARS-CoV-2)感染引发的肺炎疫情已蔓延全球,尽快认清病毒感染规律和致病机制是做好疫情防控的基础。SARS-CoV-2表面的刺突蛋白(Spike,S)识别靶细胞受体并与之结合,诱导病毒与细胞的膜融合,是病毒侵入宿主细胞的第一步,也是预防和治疗病毒感染的关键靶点。大量研究揭示了病毒进入细胞的分子机制,本文将主要对SARS-CoV-2入侵细胞的研究成果进行总结,并简要叙述以该环节为靶点的药物和疫苗研发现状。     

丙型肝炎病毒感染中的T细胞衰竭

本文简要综述丙型肝炎病毒感染中T细胞衰竭的形成机制,提出在急性和慢性丙型肝炎病毒感染时T细胞衰竭的事实,并就丙型肝炎病毒感染时的T细胞功能与其他持续性病毒感染进行比较,从而探讨急性和慢性丙型肝炎病毒感染中T细胞衰竭的机制,解释病毒持续性形成的原因,在了解病毒的致病机制和设计有效疫苗上具有实际意义.     

小RNA病毒受体的研究进展

病毒与宿主间的相互作用位点为病毒感染细胞提供可能性,位于宿主细胞上的结合位点称作受体。目前在宿主细胞的免疫球蛋白超家族、低密度脂蛋白受体家族、补体家族和整联蛋白细胞粘附分子家族等中陆续发现了小RNA病毒的受体。了解各个受体的利用情况,对理解病毒感染机制、宿主嗜性和抗病毒机理有重要意义。本文主要就已证实的小RNA病毒受体进行分类概述,以期为深入研究小RNA病毒的致病机理及其防控提供参考。     

趋化因子在病毒感染中的作用及意义

趋化因子是近年发现的一大类结构相似、功能多样的新型分子。其不仅与淋巴细胞迁移、炎症反应、新生血管生成、造血、肿瘤有关,越来越多的证据表明趋化因子在病毒感染的发生、发展中扮演着重要角色。病毒感染宿主细胞后可导致组成性和诱导性趋化因子表达谱的改变,这些表达水平改变了趋化因子通过直接和间接的方式参与了病毒的致病过程;同时某些病毒可编码趋化因子样或趋化因子受体样分子,从而干扰趋化因子网络功能。研究趋化因子在病毒感染中的作用及意义,将有利于阐明在病毒性疾病的发病机制和抗病毒感染所策略的制定,具有重要的理论和应用价值。     

口蹄疫病毒入侵宿主细胞研究进展

口蹄疫病毒(FMDV)是小RNA病毒科,口蹄疫病毒属的典型成员,是一种基因组大约含有8 400个核苷酸的无囊膜单股正链RNA病毒。大量研究发现识别细胞表面受体并侵入细胞是FMDV感染宿主细胞非常重要的环节;对FMDV而言,利用哪种受体就决定了利用哪种內吞路径。近年来在口蹄疫病毒入侵宿主细胞方面进行了大量研究,在一定程度上解释了口蹄疫病毒感染机制方面的问题,为解决实际生产问题提供了重要依据。对前期工作进行阶段性总结,为后期深入研究口蹄疫病毒致病机制和探索更有效的防治措施提供参考。     

丙型肝炎病毒感染机制的研究进展

丙型3肝炎病毒感染的机制目前不明了,大量的证据表明CD81是该病毒感染的最初受体,也有实验认为病毒与低密度脂蛋白结合后通过低密度脂蛋白受体的介导进入细胞。本文对该病毒感染的受体及它们的多态性研究作一综述。     

丙型肝炎病毒受体及其感染机制

病毒与宿主细胞的吸附是病毒感染的第一步,其相互作用决定了病毒的宿主范围和组织、细胞的易感性。丙型肝炎病毒(HCV)受体的研究,对于阐明HCV的感染机制及感染模型具有重要意义。     

整联蛋白与口蹄疫病毒感染

整联蛋白是一类分布广泛的细胞表面受体家族,它不仅在细胞的生长、移行、增殖和分化等许多方面发挥重要生物学功能,而且在多种病理过程中发挥重要作用。许多病毒都能利用整联蛋白分子作为病毒受体或共受体进入宿主细胞。本文主要就整联蛋白的特征及其在口蹄疫病毒感染宿主细胞过程中的作用机制进行综述。     

硫酸乙酰肝素与口蹄疫病毒感染

受体是病毒宿主嗜性和致病机制的主要决定因素.硫酸乙酰肝素(HS)是一种多聚阴离子碳水化合物,广泛存在于真核细胞的细胞膜和细胞基质.HS是许多病毒在细胞膜上的特异受体或辅助受体.目前发现口蹄疫病毒可利用HS和整联蛋白(ανβ3、ανβ6、ανβ1、ανβ8)作为病毒受体.口蹄疫病毒可能在不同的感染阶段利用不同类型的受体与宿主细胞相互作用中.研究病毒受体的结构和功能对理解病毒与宿主细胞的关系具有重要意义.本文主要论述了HS的生物学特性及其与口蹄疫病毒感染的关系.     

病毒感染蛋白质组学研究进展

病毒的侵入会导致宿主细胞蛋白表达模式的改变,这种改变将影响宿主细胞的正常生理功能并决定病毒的致病进程和结果.因此,病毒感染蛋白质组学研究有助于揭示病毒与宿主的相互作用机制和病毒的分子致病机制,以及寻找病毒早期感染的分子标记、建立早期诊断方法、评价治疗效果和预后.本文介绍了病毒感染蛋白质组学研究技术、病毒诱导宿主细胞蛋白质组改变和病毒感染宿主血清差异蛋白质组等方面的研究进展.     

新型冠状病毒感染与复制的进展

冠状病毒是一类具有囊膜包裹的线性单股正链RNA病毒,在自然界广泛存在,可引起不同程度的呼吸性传染病。新型冠状病毒是一种新发突发病毒,对各类人群均易感。截止目前,该病已经在世界范围内广泛流行,对公共卫生安全构成极大的威胁。文中从冠状病毒及新型冠状病毒的基因组特征、关键蛋白、对宿主的感染和复制的角度加以综述,旨在为获得病毒侵染宿主细胞致病机制的探究提供理论依据,也为特异的抗病毒药物的研发提供基础支持。     

Regulation of cell death during infection by the severe acute respiratory syndrome coronavirus and other coronaviruses

Both apoptosis and necrosis have been observed in cells infected by various coronaviruses, suggesting that the regulation of cell death is important for viral replication and/or pathogenesis. Expeditious research on the severe acute respiratory syndrome (SARS) coronavirus, one of the latest discovered coronaviruses that infect humans, has provided valuable insights into the molecular aspects of cell-death regulation during infection. Apoptosis was observed in vitro, while both apoptosis and necrosis were observed in tissues obtained from SARS patients. Viral proteins that can regulate apoptosis have been identified, and many of these also have the abilities to interfere with cellular functions. Occurrence of cell death in host cells during infection by other coronaviruses, such as the mouse hepatitis virus and transmissible porcine gastroenteritis virus, has also being extensively studied. The diverse cellular responses to infection revealed the complex manner by which coronaviruses affect cellular homeostasis and modulate cell death. As a result of the complex interplay between virus and host, infection of different cell types by the same virus does not necessarily activate the same cell-death pathway. Continuing research will lead to a better understanding of the regulation of cell death during viral infection and the identification of novel antiviral targets.     

PLP2, a potent deubiquitinase from murine hepatitis virus, strongly inhibits cellular type I interferon production

Infections by coronaviruses such as severe acute respiratory syndrome (SARS) coronavirus (SCoV) and mouse hepatitis virus A59 (MHV-A59) result in very little type I interferon (IFN) production by host cells, which is potentially responsible for the rapid viral growth and severe immunopathology associated with SARS. However, the molecular mechanisms for the low IFN production in cells infected with coronaviruses remain unclear. Here, we provide evidence that Papain-like protease domain 2 (PLP2), a catalytic domain of the nonstructural protein 3 (nsp3) of MHV-A59, can bind to IRF3, cause its deubiquitination and prevent its nuclear translocation. As a consequence, co-expression of PLP2 strongly inhibits CARDIF-, TBK1- and IRF3-mediated IFNbeta reporter activities. In addition, we show that wild-type PLP2 but not the mutant PLP2 lacking the deubiquitinase (DUB) activity can reduce IFN induction and promote viral growth in cells infected with VSV. Thus, our study uncovered a viral DUB which coronaviruses may use to escape from the host innate antiviral responses.     

猪流行性腹泻病毒通过病毒蛋白酶激活Caspase-3诱导细胞凋亡

冠状病毒是一大类能够引起呼吸系统疾病,从而威胁人类健康的病毒．目前,对冠状病毒诱导细胞凋亡及其机制研究甚少.本研究以动物冠状病毒 猪流行性腹泻病毒(PEDV) 为模型探讨冠状病毒诱导细胞凋亡效应及其可能作用机制. 通过流式细胞术检测发现感染PEDV病毒后细胞凋亡率明显升高,且PEDV诱导细胞凋亡呈时间和剂量依赖性(P<0.05或P<0.01);进一步研究发现,冠状病毒木瓜样蛋白酶（PLP）在病毒引起凋亡过程中起重要作用.实验发现,转染PEDV-PLP质粒后,caspase-3活化体表达水平明显升高. 提示冠状病毒PLP蛋白酶通过激活caspase-3在病毒诱导细胞凋亡过程中起着关键作用. 以上结果为研究人类冠状病毒PLP蛋白功能及其通过细胞凋亡调节宿主抗病毒天然免疫机制提供重要基础.     

Feline aminopeptidase N serves as a receptor for feline, canine, porcine, and human coronaviruses in serogroup I.

Two members of coronavirus serogroup I, human respiratory coronavirus HCV-229E and porcine transmissible gastroenteritis virus (TGEV), use aminopeptidase N (APN) as their cellular receptors. These viruses show marked species specificity in receptor utilization, as HCV-229E can utilize human but not porcine APN, while TGEV can utilize porcine but not human APN. To determine whether feline APN could serve as a receptor for two feline coronaviruses in serogroup I, feline infectious peritonitis virus (FIPV) and feline enteric coronavirus (FeCV), we cloned the cDNA encoding feline APN (fAPN) by PCR from cDNA isolated from a feline cell line and stably expressed it in FIPV- and FeCV-resistant mouse and hamster cells. The predicted amino acid sequence of fAPN shows 78 and 77% identity with human and porcine APN, respectively. When inoculated with either of two biologically different strains of FIPV or with FeCV, fAPN-transfected mouse and hamster cells became infected and viral antigens developed in the cytoplasm. Infectious FIPV was released from hamster cells stably transfected with fAPN. The fAPN-transfected mouse and hamster cells were challenged with other coronaviruses in serogroup I including canine coronavirus, porcine coronavirus TGEV, and human coronavirus HCV-229E. In addition to serving as a receptor for the feline coronaviruses, fAPN also served as a functional receptor for each of these serogroup I coronaviruses as shown by development of viral antigens in the cytoplasm of infected mouse or hamster cells stably transfected with fAPN. In contrast, fAPN did not serve as a functional receptor for mouse hepatitis virus (MHV-A59), which is in serogroup II and utilizes mouse biliary glycoprotein receptors unrelated to APN. Thus, fAPN serves as a receptor for a much broader range of group I coronaviruses than human and porcine APNs. The human, porcine, and canine coronaviruses in serogroup I that are able to use fAPN as a receptor have previously been shown to infect cats without causing disease. Therefore, host factors in addition to receptor specificity apparently affect the virulence and transmissibility of nonfeline serogroup I coronaviruses in the cat.     

Coronavirus infection of polarized epithelial cells

Epithelial cells are the first host cells to be infected by incoming coronaviruses. Recent observations in vitro show that coronaviruses are released from a specific side of these polarized cells, and this polarized release might be important for the spread of the infection in vivo. Mechanisms for the directional sorting of coronaviruses might be similar to those governing the polar release of secretory proteins.     

Comparison of lentiviruses pseudotyped with S proteins from coronaviruses and cell tropisms of porcine coronaviruses

The surface glycoproteins of coronaviruses play an important role in receptor binding and cell entry. Different coronaviruses interact with their specific receptors to enter host cells. Lentiviruses pseudotyped with their spike proteins (S) were compared to analyze the entry efficiency of various coronaviruses. Our results indicated that S proteins from different coronaviruses displayed varied abilities to mediate pseudotyped virus infection. Furthermore, the cell tropisms of porcine epidemic diarrhea virus (PEDV) and transmissible gastroenteritis virus (TGEV) have been characterized by live and pseudotyped viruses. Both live and pseudoviruses could infected Vero- CCL-81 (monkey kidney), Huh-7 (human liver), and PK-15 (pig kidney) cells efficiently. CCL94 (cat kidney) cells could be infected efficiently by TGEV but not PEDV. Overall, our study provides new insights into the mechanisms of viral entry and forms a basis for antiviral drug screening.

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Differential Sensitivity of Bat Cells to Infection by Enveloped RNA Viruses: Coronaviruses,Paramyxoviruses, Filoviruses,and Influenza Viruses

Bats (Chiroptera) host major human pathogenic viruses including corona-, paramyxo, rhabdo- and filoviruses. We analyzed six different cell lines from either Yinpterochiroptera (including African flying foxes and a rhinolophid bat) or Yangochiroptera (genera Carollia and Tadarida) for susceptibility to infection by different enveloped RNA viruses. None of the cells were sensitive to infection by transmissible gastroenteritis virus (TGEV), a porcine coronavirus, or to infection mediated by the Spike (S) protein of SARS-coronavirus (SARS-CoV) incorporated into pseudotypes based on vesicular stomatitis virus (VSV). The resistance to infection was overcome if cells were transfected to express the respective cellular receptor, porcine aminopeptidase N for TGEV or angiotensin-converting enzyme 2 for SARS-CoV. VSV pseudotypes containing the S proteins of two bat SARS-related CoV (Bg08 and Rp3) were unable to infect any of the six tested bat cell lines. By contrast, viral pseudotypes containing the surface protein GP of Marburg virus from the family Filoviridae infected all six cell lines though at different efficiency. Notably, all cells were sensitive to infection by two paramyxoviruses (Sendai virus and bovine respiratory syncytial virus) and three influenza viruses from different subtypes. These results indicate that bat cells are more resistant to infection by coronaviruses than to infection by paramyxoviruses, filoviruses and influenza viruses. Furthermore, these results show a receptor-dependent restriction of the infection of bat cells by CoV. The implications for the isolation of coronaviruses from bats are discussed.     

SARS virus: The beginning of the unraveling of a new coronavirus

Severe acute respiratory syndrome (SARS) virus caused a severe outbreak in several regions of the world in 2003. The virus is a novel coronavirus, which may have an origin in wild animals such as civet cats in southern China. Its genome structure, gene expression pattern and protein profiles are similar to those of other coronaviruses. However, distinct patterns of several open reading frames in the SARS virus genome may contribute to its severe virulence. The potential mutability of the coronavirus genome may pose problems in the control of future SARS outbreaks. The mechanism of SARS pathogenesis may involve both direct viral cytocidal effects on the target cells and immune-mediated mechanisms. The life cycle of the SARS virus is largely unknown; however, based on the analogy with other coronaviruses, several potential targets for antiviral development are identified. Vaccines offer an important preventive measure for possible future recurrences of SARS, but the prospect for their development is still unknown because of the uncertainty regarding the role of immune responses in SARS virus pathogenesis. The comparative studies of other coronaviruses offer insights into the understanding of SARS virus.     

Identification of host factors involved in coronavirus replication by quantitative proteomics analysis

In this study, we applied a quantitative proteomic approach, based on SILAC, to investigate the interactions of coronaviruses with the secretory pathway of the host cell, with the aim to identify host factors involved in coronavirus replication. Comparison of the protein profiles of Golgi-enriched fractions of cells that were either mock infected or infected with mouse hepatitis virus revealed the significant depletion or enrichment of 116 proteins. Although ribosomal/nucleic acid binding proteins were enriched in the Golgi-fractions of mouse hepatitis virus-infected cells, proteins annotated to localize to several organelles of the secretory pathway were overrepresented among the proteins that were depleted from these fractions upon infection. We hypothesized that proteins, of which the abundance or distribution is affected by infection, are likely to be involved in the virus life cycle. Indeed, depletion of a small subset of the affected proteins by using small interfering RNAs identified several host factors involved in coronavirus infection. Transfection of small interfering RNAs targeting either C11orf59 or Golgi apparatus glycoprotein 1 resulted in increased virus replication, whereas depletion of vesicle-trafficking protein vesicle-trafficking protein sec22b enhanced the release of infectious progeny virus. Overexpression of these proteins, on the other hand, had a negative effect on virus replication. Overall, our study shows that the SILAC approach is a suitable tool to study host-pathogen interactions and to identify host proteins involved in virus replication.