Specific Acquisition of Functional CD59 but Not CD46 or CD55 by Hepatitis C Virus

Viruses of different families encode for regulators of the complement system (RCAs) or acquire such RCAs from the host to get protection against complement-mediated lysis (CML). As hepatitis C virus (HCV) shares no genetic similarity to any known RCA and is detectable at high titers in sera of infected individuals, we investigated whether HCV has adapted host-derived RCAs to resist CML. Here we report that HCV selectively incorporates CD59 while neither CD55, nor CD46 are associated with the virus. The presence of CD59 was shown by capture assays using patient- and cell culture-derived HCV isolates. Association of CD59 with HCV was further confirmed by Western blot analysis using purified viral supernatants from infected Huh 7.5 cells. HCV captured by antibodies specific for CD59 remained infectious for Huh 7.5 cells. In addition, blocking of CD59 in the presence of active complement reduced the titer of HCV most likely due to CML. HCV produced in CD59 knock-down cells were more significantly susceptible to CML compared to wild type virus, but neither replication, assembly nor infectivity of the virus seemed to be impaired in the absence of CD59. In summary our data indicate that HCV incorporates selectively CD59 in its envelope to gain resistance to CML in serum of infected individuals.     

The Paramyxoviruses Simian Virus 5 and Mumps Virus Recruit Host Cell CD46 To Evade Complement-Mediated Neutralization

The complement system is a critical component of the innate immune response that all animal viruses must face during natural infections. Our previous results have shown that treatment of the paramyxovirus simian virus 5 (SV5) with human serum results in deposition of complement C3-derived polypeptides on virion particles. Here, we show that the virion-associated C3 component includes the inactive form iC3b, suggesting that SV5 may have mechanisms to evade the host complement system. Electron microscopy, gradient centrifugation, and Western blot analysis indicated that purified SV5 virions derived from human A549 cells contained CD46, a plasma membrane-expressed regulator of complement that acts as a cofactor for cleavage and inactivation of C3b into iC3b. In vitro cleavage assays with purified complement components showed that SV5 virions had C3b cofactor activity, resulting in specific factor I-mediated cleavage of C3b into inactive iC3b. SV5 particles generated in CHO cells, which do not express CD46, did not have cofactor activity. Conversely, virions derived from a CHO cell line that was engineered to overexpress human CD46 contained elevated levels of virion-associated CD46 and displayed enhanced C3b cofactor activity. In comparison with C3b, purified SV5 virions had very low cofactor activity against C4b, consistent with the known preference of CD46 for C3b versus C4b. Similar results were obtained for the closely related mumps virus (MuV), except that MuV particles derived from CHO-CD46 cells had higher C4b cofactor activity than SV5 virions. In neutralization assays with human serum, SV5 and MuV containing CD46 showed slower kinetics and more resistance to neutralization than SV5 and MuV that lacked CD46. Our results support a model in which the rubulaviruses SV5 and MuV incorporate cell surface complement inhibitors into progeny virions as a mechanism to limit complement-mediated neutralization.The complement system constitutes a complex group of both soluble and cell-associated proteins that together form an integral part of the innate host defense against pathogens (reviewed in references 7, 9, 11, and 31). Complement can serve to link innate and adaptive immunity through a large number of activities, including recognition of viruses, direct neutralization of infectivity, recruitment and stimulation of leukocytes, opsonization by immune cells, and activation of T- and B-cell responses (9, 11, 27). Complement activation and the ability of viruses to counteract complement can play important roles in viral pathogenesis, as well as the design of more effective vaccines and therapeutic vectors (6, 9, 17, 36, 43). The overall goal of the work described here was to determine the mechanism by which the paramyxoviruses simian virus 5 (SV5) and mumps virus (MuV) limit activation of complement pathways.The complement cascade can be initiated through three main pathways: the classical pathway, the lectin pathway, and the alternative pathway (11, 40). These three pathways converge on a central component, C3, which is activated by cleavage into C3a and C3b. C3a serves as an anaphylatoxin to promote inflammation. C3b can bind covalently to viral components to aid in opsonization and phagocytosis. In addition, C3b can associate with other factors, such as factor B, to form the C3 convertase (e.g., C3bBb), and this functions to amplify the initially deposited C3b signal by further cleavage of C3 molecules in a feedback loop. Likewise, C4 can be activated by cleavage into the anaphylatoxin C4a and the C4b fragment, which links the classical and lectin pathways with the alternative pathway. The association of C3b with components further downstream, such as C6 through C9, can lead to formation of the membrane attack complex, which is capable of lysing virus particles or infected cells (reviewed in references 7, 11, and 28).The complement system needs to be highly regulated to prevent inappropriate activation and potential damage to normal cells and healthy tissues (3). Self-regulation of complement pathways involves the highly concerted actions of a family of soluble and cell-associated proteins called regulators of complement activation (RCA). RCA proteins can limit inappropriate complement activation through two major mechanisms: (i) by accelerating the disassociation of C3 or C5 convertase or (ii) by acting as a cofactor to promote proteolytic cleavage of C3b or C4b by the complement protease factor I. Examples of RCA proteins are factor H, CD46, complement receptor 1 (CR1, or CD35), and C4 binding protein (14, 19, 24, 45).CD46, or membrane cofactor protein, is an integral membrane RCA protein that is expressed on a wide range of tissues and cell types (32). CD46 is an N- and O-linked glycosylated protein expressed at the plasma membrane as multiple isoforms that are derived from alternative splicing (32, 33, 39, 41). CD46 selectively binds to both C3b and C4b on cell surfaces, where it acts as a cofactor to promote efficient cleavage by complement protease factor I (44; reviewed in references 5 and 32). For C3b, CD46 and factor I combine to mediate inactivation to iC3b, and this is a major mechanism for limiting the amplification of low basal levels of C3b that arise from spontaneous alternative-pathway activation. CD46 also serves as a cofactor for factor I-mediated cleavage of C4b into C4c and C4d, but this cofactor activity is less efficient than that seen for C3b cleavage (35, 45).Viruses have evolved a number of mechanisms to inhibit or to delay the neutralizing effects of complement (9, 16). Large DNA viruses have coding capacities that allow them to encode a variety of mimics of host cell RCA proteins, and these viral homologs often function to inactivate complement components by supplying cofactor activity or by accelerating the decay of convertases (reviewed in references 2, 7, 29, and 31). For example, herpesvirus saimiri expresses a complement control protein that inhibits the C3 convertase (20). As an alternative mechanism to counteract complement, a number of enveloped DNA viruses and retroviruses have been shown to recruit cell-associated RCA proteins into budding particles (15, 47, 48). Examples of this are vaccinia virus and human immunodeficiency virus type 1, which incorporate CD55, CD59, and CD46 into progeny virions (16, 42, 48).In contrast to retroviruses and large DNA viruses, mechanisms that are employed to limit or evade host cell complement pathways have not been described for the paramyxovirus family of negative-strand RNA viruses (30). It has been known for many years that complement is an important factor in paramyxovirus neutralization (18, 23, 34, 49, 50). For example, the closely related paramyxoviruses SV5 and MuV preferentially activate the complement alternative pathway in vitro, and this activation can contribute to the efficiency of neutralization by human serum (23, 26). These findings raised the question of whether negative-strand RNA viruses have mechanisms to limit activation and/or amplification of the complement cascade.We have previously shown that treatment of SV5 and MuV particles with normal human serum led to deposition of C3-derived components on virions, but the virion-associated C3 molecules had properties of the inactive form, iC3b, and not the intact C3b (26). In this study, we tested the hypothesis that SV5 and MuV incorporate host cell RCA proteins into budding virions as a mechanism to limit complement activity through inactivation of C3b. CD46 was found associated with purified SV5 and MuV virions that were derived from cells expressing CD46, and these particles displayed C3b cofactor activity in vitro. Consistent with this inactivation of C3b, CD46-containing virus was more resistant to in vitro neutralization by human serum than virus derived from CD46-deficient cells. Our results support a model in which these closely related paramyxoviruses incorporate at least one cell surface RCA protein into progeny virions as a mechanism to evade complement-mediated neutralization.     

Incorporation of Host Complement Regulatory Proteins into Newcastle Disease Virus Enhances Complement Evasion

Newcastle disease virus (NDV), an avian paramyxovirus, is inherently tumor selective and is currently being considered as a clinical oncolytic virus and vaccine vector. In this study, we analyzed the effect of complement on the neutralization of NDV purified from embryonated chicken eggs, a common source for virus production. Fresh normal human serum (NHS) neutralized NDV by multiple pathways of complement activation, independent of neutralizing antibodies. Neutralization was associated with C3 deposition and the activation of C2, C3, C4, and C5 components. Interestingly, NDV grown in mammalian cell lines was resistant to complement neutralization by NHS. To confirm whether the incorporation of regulators of complement activity (RCA) into the viral envelope afforded complement resistance, we grew NDV in CHO cells stably transfected with CD46 or HeLa cells, which strongly express CD46 and CD55. NDV grown in RCA-expressing cells was resistant to complement by incorporating CD46 and CD55 on virions. Mammalian CD46 and CD55 molecules on virions exhibited homologous restriction, since chicken sera devoid of neutralizing antibodies to NDV were able to effectively neutralize these virions. The incorporation of chicken RCA into NDV produced in embryonated eggs similarly provided species specificity toward chicken sera.     

Morbillivirus Downregulation of CD46

There is evidence that CD46 (membrane cofactor protein) is a cellular receptor for vaccine and laboratory-passaged strains of measles virus (MV). Following infection with these MV strains, CD46 is downregulated from the cell surface, and consequent complement-mediated lysis has been shown to occur upon infection of a human monocytic cell line. The MV hemagglutinin (H) protein alone is capable of inducing this downregulation. Some wild-type strains of MV fail to downregulate CD46, despite infection being prevented by anti-CD46 antibodies. In this study we show that CD46 is also downregulated to the same extent by wild-type, vaccine, and laboratory-passaged strains of rinderpest virus (RPV), although CD46 did not appear to be the receptor for RPV. Expression of the RPV H protein by a nonreplicating adenovirus vector was also found to cause this downregulation. A vaccine strain of peste des petits ruminants virus caused slight downregulation of CD46 in infected Vero cells, while wild-type and vaccine strains of canine distemper virus and a wild-type strain of dolphin morbillivirus failed to downregulate CD46. Downregulation of CD46 can, therefore, be a function independent of the use of this protein as a virus receptor.     

Cellular Proteins Associated with the Interior and Exterior of Vesicular Stomatitis Virus Virions

Virus particles (virions) often contain not only virus-encoded but also host-encoded proteins. Some of these host proteins are enclosed within the virion structure, while others, in the case of enveloped viruses, are embedded in the host-derived membrane. While many of these host protein incorporations are likely accidental, some may play a role in virus infectivity, replication and/or immunoreactivity in the next host. Host protein incorporations may be especially important in therapeutic applications where large numbers of virus particles are administered. Vesicular stomatitis virus (VSV) is the prototypic rhabdovirus and a candidate vaccine, gene therapy and oncolytic vector. Using mass spectrometry, we previously examined cell type dependent host protein content of VSV virions using intact (“whole”) virions purified from three cell lines originating from different species. Here we aimed to determine the localization of host proteins within the VSV virions by analyzing: i) whole VSV virions; and ii) whole VSV virions treated with Proteinase K to remove all proteins outside the viral envelope. A total of 257 proteins were identified, with 181 identified in whole virions and 183 identified in Proteinase K treated virions. Most of these proteins have not been previously shown to be associated with VSV. Functional enrichment analysis indicated the most overrepresented categories were proteins associated with vesicles, vesicle-mediated transport and protein localization. Using western blotting, the presence of several host proteins, including some not previously shown in association with VSV (such as Yes1, Prl1 and Ddx3y), was confirmed and their relative quantities in various virion fractions determined. Our study provides a valuable inventory of virion-associated host proteins for further investigation of their roles in the replication cycle, pathogenesis and immunoreactivity of VSV.     

Reduced sensitivity to human serum inactivation of enveloped viruses produced by pig cells transgenic for human CD55 or deficient for the galactosyl-alpha(1-3) galactosyl epitope

Complement activation mediated by the major xenogeneic epitope in the pig, galactosyl-alpha(1-3) galactosyl sugar structure (alpha-Gal), and human natural antibodies could cause hyperacute rejection (HAR) in pig-to-human xenotransplantation. The same reaction on viruses bearing alpha-Gal may serve as a barrier to zoonotic infection. Expressing human complement regulatory proteins or knocking out alpha-Gal epitopes in pig in order to overcome HAR may therefore pose an increased risk in xenotransplantation with regard to zoonosis. We investigated whether amphotropic murine leukemia virus, porcine endogenous retrovirus, and vesicular stomatitis virus (VSV) budding from primary transgenic pig aortic endothelial (TgPAE) cells expressing human CD55 (hCD55 or hDAF) was protected from human-complement-mediated inactivation. VSV propagated through the ST-IOWA pig cell line, in which alpha-galactosyl-transferase genes were disrupted (Gal null), was also tested for sensitivity to human complement. The TgPAE cells were positive for hCD55, and all pig cells except the Gal-null ST-IOWA expressed alpha-Gal epitopes. Through antibody binding, we were able to demonstrate the incorporation of hCD55 onto VSV particles. Viruses harvested from TgPAE cells were relatively resistant to complement-mediated inactivation by the three sources of human sera tested. Additionally, VSV from Gal-null pig cells was resistant to human complement inactivation. Such protection of enveloped viruses may increase the risk of zoonosis from pigs genetically modified for pig-to-human xenotransplantation.     

Protection of xenogeneic cells from human complement-mediated lysis by the expression of human DAF, CD59 and MCP

CD59 and membrane cofactor protein (MCP, CD46) are widely expressed cell surface glycoproteins that protect host cells from the effect of homologous complement attack. cDNAs encoding human CD59 and MCP cloned from Chinese human embryo were separately transfected into NIH/3T3 cells resulting in the expression of human CD59 and MCP protein on the cell surface. The functional properties of expressed proteins were studied. When the transfected cells were exposed to human serum as a source of complement and naturally occurring anti-mouse antibody, they were resistant to human complement-mediated cell killing. However, the cells remained sensitive to rabbit and guinea pig complement. Human CD59 and MCP can only protect NIH/3T3 cells from human complement-mediated lysis. These results demonstrated that complement inhibitory activity of these proteins is species-selective. The cDNAs of CD59 and MCP were also separately transfected into the endothelial cells (ECs) of the pigs transgenic for the human DAF gene to investigate a putative synergistic action. The ECs expressing both DAF and MCP proteins or both DAF and CD59 proteins exhibited more protection against cytolysis by human serum compared to the cells with only DAF expressed alone.     

Incorporation of CD4 into virions by a recombinant herpes simplex virus.

Two herpes simplex virus type 1 (HSV-1) recombinants were constructed by inserting the human CD4 gene into the HSV-1 genome between the gC promoter and the gC structural gene. These viruses, designated K delta T/CD4 and K082/CD4, synthesized a significant quantity of CD4. CD4 was expressed on the surface of infected cells at levels substantially higher than on the surface of HUT78 cells, a CD4+ cell line. Most significantly, a small but detectable quantity of CD4 was incorporated into virions produced by the recombinant viruses. This was demonstrated both by immunoprecipitation of CD4 from purified virions and by neutralization of the recombinant virions by OKT4 and complement. These results suggest that specific virion incorporation signals are not strictly required for inclusion of glycoproteins into HSV-1 virions. It may be possible to utilize this ability to alter the host range or tissue specificity of HSV-1.     

Ablation of Nectin4 Binding Compromises CD46 Usage by a Hybrid Vesicular Stomatitis Virus/Measles Virus

Measles virus (MV) immunosuppression is due to infection of SLAM-positive immune cells, whereas respiratory shedding and virus transmission are due to infection of nectin4-positive airway epithelial cells. The vaccine lineage MV strain Edmonston (MV-Edm) acquired an additional tropism for CD46 which is the basis of its oncolytic specificity. VSVFH is a vesicular stomatitis virus (VSV) encoding the MV-Edm F and H entry proteins in place of G. The virus spreads faster than MV-Edm and is highly fusogenic and a potent oncolytic. To determine whether ablating nectin4 tropism from VSVFH might prevent shedding, increasing its safety profile as an oncolytic, or might have any effect on CD46 binding, we generated VSVFH viruses with H mutations that disrupt attachment to SLAM and/or nectin4. Disruption of nectin4 binding reduced release of VSVFH from the basolateral side of differentiated airway epithelia composed of Calu-3 cells. However, because nectin4 and CD46 have substantially overlapping receptor binding surfaces on H, disruption of nectin4 binding compromised CD46 binding and greatly diminished the oncolytic potency of these viruses on human cancer cells. Thus, our results support continued preclinical development of VSVFH without ablation of nectin4 binding.     

Comparison of the Ribonucleic Acid Polymerases of Two Rhabdoviruses, Kern Canyon Virus and Vesicular Stomatitis Virus

A ribonucleic acid (RNA)-dependent RNA polymerase has been demonstrated in Kern Canyon virus (KCV) particles. The RNA product of the KCV polymerase hybridizes to KCV viral RNA. The properties of this viral enzyme have been characterized and compared with those of vesicular stomatitis virus (VSV). RNA polymerases from both viruses require similar conditions of temperature, pH, and detergent and magnesium concentrations for maximal synthesis of RNA. The RNA polymerase contained in the virion of KCV was more dependent on the presence of a sulfhydryl agent than was the VSV enzyme. Under optimal conditions, the specific activity of the VSV polymerase is about twenty-five times as great as that of KCV.     

Exceptional resistance of human H2 glioblastoma cells to complement-mediated killing by expression and utilization of factor H and factor H-like protein 1

Of over 20 nucleated cell lines we have examined to date, human H2 glioblastoma cells have turned out to be the most resistant to complement-mediated cytolysis in vitro. H2 cells expressed strongly the membrane attack complex inhibitor protectin (CD59), moderately CD46 (membrane cofactor protein) and CD55 (decay-accelerating factor), but no CD35 (complement receptor 1). When treated with a polyclonal anti-H2 Ab, anti-CD59 mAb, and normal human serum, only 5% of H2 cells became killed. Under the same conditions, 70% of endothelial-like EA.hy 926 cells and 40% of U251 control glioma cells were killed. A combined neutralization of CD46, CD55, and CD59 increased H2 lysis only minimally, demonstrating that these complement regulators are not enough to account for the resistance of H2 cells. After treatment with Abs and serum, less C5b-9 was deposited on H2 than on U251 and EA.hy 926 cell lines. A reason for the exceptional resistance of H2 cells was revealed when RT-PCR and protein biochemical methods showed that the H2 cells, unlike the other cell lines tested, actively produced the soluble complement inhibitors factor H and factor H-like protein 1. H2 cells were also capable of binding human factor H from the fluid phase to their cell surface and promoted the cleavage of C3b to its inactive form iC3b more efficiently than U251 and EA.hy 926 cells. In accordance, anti-factor H mAbs enhanced killing of H2 glioblastoma cells. Taken together, our results show that production and binding of factor H and factor H-like protein 1 is a novel mechanism that these malignant cells utilize to escape complement-mediated killing.     

Proteins of Vesicular Stomatitis Virus and of Phenotypically Mixed Vesicular Stomatitis Virus-Simian Virus 5 Virions

The identity of the glycoprotein of vesicular stomatitis virus (VSV) as the spike protein has been confirmed by the removal of the spikes with a protease from Streptomyces griseus, leaving bullet-shaped particles bounded by a smooth membrane. This treatment removes the glycoprotein but does not affect the other virion proteins, apparently because they are protected from the enzyme by the lipids in the viral membrane. The proteins of phenotypically mixed, bullet-shaped virions produced by cells mixedly infected with VSV and the parainfluenza virus simian virus 5 (SV5) have been analyzed by polyacrylamide gel electrophoresis. These virions contain all the VSV proteins plus the two SV5 spike proteins, both of which are glycoproteins. The finding of the SV5 spike glycoproteins on virions with the typical morphology of VSV indicates that there is not a stringent requirement that only the VSV glycoprotein can be used to form the bullet-shaped virion. On the other hand, the SV5 nucleocapsid protein and the major non-spike protein of the SV5 envelope were not detected in the phenotypically mixed virions, and this suggests that a specific interaction between the VSV nucleocapsid and regions of the cell membrane which contain the nonglycosylated VSV envelope protein is necessary for assembly of the bullet-shaped virion.     

An ssDNA virus infecting archaea: a new lineage of viruses with a membrane envelope

Archaeal organisms are generally known as diverse extremophiles, but they play a crucial role also in moderate environments. So far, only about 50 archaeal viruses have been described in some detail. Despite this, unusual viral morphotypes within this group have been reported. Interestingly, all isolated archaeal viruses have a double-stranded DNA (dsDNA) genome. To further characterize the diversity of archaeal viruses, we screened highly saline water samples for archaea and their viruses. Here, we describe a new haloarchaeal virus, Halorubrum pleomorphic virus 1 (HRPV-1) that was isolated from a solar saltern and infects an indigenous host belonging to the genus Halorubrum . Infection does not cause cell lysis, but slightly retards growth of the host and results in high replication of the virus. The sequenced genome (7048 nucleotides) of HRPV-1 is single-stranded DNA (ssDNA), which makes HRPV-1 the first characterized archaeal virus that does not have a dsDNA genome. In spite of this, similarities to another archaeal virus were observed. Two major structural proteins were recognized in protein analyses, and by lipid analyses it was shown that the virion contains a membrane. Electron microscopy studies indicate that the enveloped virion is pleomorphic (approximately 44 × 55 nm). HRPV-1 virion may represent commonly used virion architecture, and it seems that structure-based virus lineages may be extended to non-icosahedral viruses.     

Dual Function of CD81 in Influenza Virus Uncoating and Budding

As an obligatory pathogen, influenza virus co-opts host cell machinery to harbor infection and to produce progeny viruses. In order to characterize the virus-host cell interactions, several genome-wide siRNA screens and proteomic analyses have been performed recently to identify host factors involved in influenza virus infection. CD81 has emerged as one of the top candidates in two siRNA screens and one proteomic study. The exact role played by CD81 in influenza infection, however, has not been elucidated thus far. In this work, we examined the effect of CD81 depletion on the major steps of the influenza infection. We found that CD81 primarily affected virus infection at two stages: viral uncoating during entry and virus budding. CD81 marked a specific endosomal population and about half of the fused influenza virus particles underwent fusion within the CD81-positive endosomes. Depletion of CD81 resulted in a substantial defect in viral fusion and infection. During virus assembly, CD81 was recruited to virus budding site on the plasma membrane, and in particular, to specific sub-viral locations. For spherical and slightly elongated influenza virus, CD81 was localized at both the growing tip and the budding neck of the progeny viruses. CD81 knockdown led to a budding defect and resulted in elongated budding virions with a higher propensity to remain attached to the plasma membrane. Progeny virus production was markedly reduced in CD81-knockdown cells even when the uncoating defect was compensated. In filamentous virus, CD81 was distributed at multiple sites along the viral filament. Taken together, these results demonstrate important roles of CD81 in both entry and budding stages of the influenza infection cycle.     

Role of CD40 ligand and CD28 in induction and maintenance of antiviral CD8+ effector T cell responses

The primary aim of this report was to evaluate the immune responses of CD40 ligand-deficient (CD40L-/-) mice infected with two viruses known to differ markedly in their capacity to replicate in the host. Lymphocytic choriomeningitis virus (LCMV) is a natural mouse pathogen that replicates widely and extensively, whereas vesicular stomatitis virus (VSV) spreads poorly. We found that the primary response of CD40L-/- mice toward VSV is significantly impaired; proliferation of both CD4+ and CD8+ cells is reduced 2- to 3-fold, few CD8+ cells acquire an activated phenotype, and little functional activity is induced. Very similar results were obtained in VSV-infected, CD28-deficient mice. In contrast, neither CD40L nor CD28 was required for induction of a primary CD8+ response toward LCMV. Surprisingly, lack of CD4+ T cells had no impact on the primary immune response toward any of the viruses, even though the CD40 ligand dependence demonstrated for VSV would be expected to be associated with CD4 dependence. Upon coinfection of VSV-infected mice with LCMV, the requirement for CD40 ligand (but not CD28) could be partially bypassed, as evidenced by a 3-fold increase in the frequency of VSV-specific CD8+ T cells on day 6 postinfection. Finally, despite the fact that the primary LCMV-specific CD8+ response is virtually unimpaired in CD40L-/- mice, their capacity to maintain CD8+ effector activity and to permanently control the infection is significantly reduced. Thus, our results demonstrate that the importance of CD40/CD40L interaction for activation of CD8+ T cells varies between viruses and over time.     

腮腺炎病毒F蛋白七肽重复序列的表达、纯化及特性分析

副粘病毒F蛋白的两段七肽重复序列（HR1和HR2）在病毒侵染细胞的过程中相互作用形成热稳定的富含α螺旋的异源二聚体,此结构的形成引起病毒囊膜与细胞膜的并置而最终导致膜融合的发生。腮腺炎病毒（Mumps virus, MuV）属于副粘病毒科,腮腺炎病毒属,可能利用与其他副粘病毒相似的侵染机制。本研究对MuV 融合蛋白的HR区进行了计算机程序预测,并利用大肠杆菌GST融合表达系统对MuV F蛋白HR1和HR2两段多肽进行了表达和纯化,通过GST pull_down 实验证实HR1和HR2多肽在体外能够相互作用,凝胶过滤层析证明HR1、HR2多肽能够形成多聚体,说明MuV F蛋白的HR区的相互作用可能是其发挥融合功能的关键因素。     

VSV Pseudotype Particles with the Coat of Avian Myeloblastosis Virus

PSEUDOTYPES of vesicular stomatitis virus (VSV) with the coat of avian myeloblastosis (AMV) or murine leukaemia viruses—VSV(AMV) and VSV(MLV)—can be produced by growing VSV in chick cells preinfected with AMV or in mouse cells preinfected with MLV¹. The VSV particles carrying their own neutralization antigen and double-neutralizable particles may be inactivated with antiserum against VSV. The surviving pseudotypes possess neutralization, host-range and interference specificities corresponding to the tumour virus donating their coat. It has also been shown that a conditional lethal mutant of VSV in which a structural protein is affected is complemented under restrictive conditions with AMV. This mutant, ts-45, when complemented with AMV again predominantly produces the pseudotype VSV(AMV).     

The entry into host cells of Sindbis virus, vesicular stomatitis virus and Sendai virus.

We have compared the mechanisms of entry into host cells of three enveloped viruses: Sendai virus, vesicular stomatitis virus (VSV) and Sindbis virus. Virus entry by membrane fusion should antigenically modify the surface of a newly infected cell in such a way that it will be killed by anti-viral antibody and complement. On the other hand, virus entry by a mechanism involving uptake by the cell of the whole virion should not make cells sensitive to antibody and complement. As expected, cells newly infected with Sendai virus were readily and completely lysed by anti-Sendai antibody and complement. In marked contrast, however, cells newly infected with either Sindbis virus or VSV were killed by anti-viral antibody and complement only when infected at an extremely high multiplicity of infection, in excess of 1000 plaque-forming units per cell. We favor the following explanation for these results with Sindbis virus and VSV: a very large majority of the Sindbis and VSV virions entered the infected cells by some means other than membrane fusion, presumably engulfment of the whole particle. Efficient entry by way of membrane fusion may therefore not be a general characteristic of enveloped viruses.