The Experts below are selected from a list of 22092 Experts worldwide ranked by ideXlab platform
Geoffrey L Smith - One of the best experts on this subject based on the ideXlab platform.
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vΔ169 generates better protection after challenge.
2015Co-Authors: Pavla Strnadova, Hongwei Ren, Robert Valentine, Michela Mazzon, Trevor R. Sweeney, Ian Brierley, Geoffrey L SmithAbstract:(A) BALB/c mice (n = 5) were infected i.n. with 5 × 103 PFU of the purified Viruses and at day 28 the mice were challenged i.n. with 5 × 106 PFU of wild type Virus and the weight change was determined daily. (B) BALB/c mice were infected as in (A) and sera were collected 28 days p.i. and assayed for neutralization of intracellular Mature Virus of VACV strain WR. The median value for each population is represented by a horizontal black bar. (C, D) BALB/c mice were infected as in (A) and at 28 days p.i. splenic lymphocytes were harvested and their ability to lyse uninfected YAC-1 (C) or VACV-infected P815 (D) cells was determined by chromium release assay. Data are presented as the percentage cell lysis at various effector to target (E:T) cell ratios. Data shown are from one representative experiment (n = 2) and results are expressed as the average ± SEM. Statistical analysis for all panels was performed using a two-tailed Student’s t-test, * p < 0.05.
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serological responses in humans to the smallpox vaccine lc16m8
Journal of General Virology, 2011Co-Authors: Benjamin F Johnson, Tomoya Saito, Hiroyuki Yokote, Yasuhiro Kanatani, Tatsuya Fujii, Geoffrey L SmithAbstract:In response to potential bioterrorism with smallpox, members of the Japanese Self-Defense Forces were vaccinated with vaccinia Virus (VACV) strain LC16m8, an attenuated smallpox vaccine derived from VACV strain Lister. The serological response induced by LC16m8 to four virion-surface proteins and the intracellular Mature Virus (IMV) and extracellular enveloped Virus (EEV) was investigated. LC16m8 induced antibody response against the IMV protein A27 and the EEV protein A56. LC16m8 also induced IMV-neutralizing antibodies, but unlike the VACV strain Lister, did not induce either EEV-neutralizing antibody or antibody to EEV protein B5, except after revaccination. Given that B5 is the only target for EEV-neutralizing antibody and that neutralization of both IMV and EEV give optimal protection against orthopoxVirus challenge, these data suggest that immunity induced by LC16m8 might be less potent than that deriving from strain Lister. This potential disadvantage should be balanced against the advantage of the greater safety of LC16m8.
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vaccinia Virus strain nyvac induces substantially lower and qualitatively different human antibody responses compared with strains lister and dryvax
Journal of General Virology, 2008Co-Authors: Claire M Midgley, Mike M Putz, Jonathan Weber, Geoffrey L SmithAbstract:The antibody responses elicited by immunization of humans with vaccinia Virus (VACV) strains Lister, Dryvax and NYVAC have been determined and compared. Neutralizing antibodies against intracellular Mature Virus (IMV) and extracellular enveloped Virus (EEV), and binding antibody titres (ELISA) against the EEV protein B5, the IMV proteins A27 and H3, and VACV-infected cell lysate were measured. Lister and Dryvax induced broadly similar antibody titres, consistent with the fact that these vaccines each protected against smallpox. In contrast, antibody titres induced by NYVAC were significantly lower than those induced by both Lister and Dryvax. Moreover, there were qualitative differences with NYVAC-immunized subjects failing to induce A27-specific antibodies. These observations suggest that although NYVAC is a safer VACV strain, it does not induce an optimal VACV-specific antibody response. However, NYVAC strains engineered to express antigens from other pathogens remain promising candidate vaccines for immunization against other diseases.
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short communication vaccinia Virus strain nyvac induces substantially lower and qualitatively different human antibody responses compared with strains lister and dryvax
2008Co-Authors: Claire M Midgley, Mike M Putz, Jonathan Weber, Geoffrey L SmithAbstract:The antibody responses elicited by immunization of humans with vaccinia Virus (VACV) strains Lister, Dryvax and NYVAC have been determined and compared. Neutralizing antibodies against intracellular Mature Virus (IMV) and extracellular enveloped Virus (EEV), and binding antibody titres (ELISA) against the EEV protein B5, the IMV proteins A27 and H3, and VACV-infected cell lysate were measured. Lister and Dryvax induced broadly similar antibody titres, consistent with the fact that these vaccines each protected against smallpox. In contrast, antibody titres induced by NYVAC were significantly lower than those induced by both Lister and Dryvax. Moreover, there were qualitative differences with NYVAC-immunized subjects failing to induce A27-specific antibodies. These observations suggest that although NYVAC is a safer VACV strain, it does not induce an optimal VACV-specific antibody response. However, NYVAC strains engineered to express antigens from other pathogens remain promising candidate vaccines for immunization against other diseases.
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quantification of antibody responses against multiple antigens of the two infectious forms of vaccinia Virus provides a benchmark for smallpox vaccination
Nature Medicine, 2006Co-Authors: Mike M Putz, Claire M Midgley, Mansun Law, Geoffrey L SmithAbstract:Smallpox was eradicated without an adequate understanding of how vaccination induced protection. In response to possible bioterrorism with smallpox, the UK government vaccinated ∼300 health care workers with vaccinia Virus (VACV) strain Lister. Antibody responses were analyzed using ELISA for multiple surface antigens of the extracellular enveloped Virus (EEV) and the intracellular Mature Virus (IMV), plaque reduction neutralization and a fluorescence-based flow cytometric neutralization assay. Antibody depletion experiments showed that the EEV surface protein B5 is the only target responsible for EEV neutralization in vaccinated humans, whereas multiple IMV surface proteins, including A27 and H3, are targets for IMV-neutralizing antibodies. These data suggest that it would be unwise to exclude the B5 protein from a future smallpox vaccine. Repeated vaccination provided significantly higher B5-specific and thus EEV-neutralizing antibody responses. These data provide a benchmark against which new, safer smallpox vaccines and residual immunity can be compared.
Wen Chang - One of the best experts on this subject based on the ideXlab platform.
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differential innate immune signaling in macrophages by wild type vaccinia Mature Virus and a mutant Virus with a deletion of the a26 protein
Journal of Virology, 2017Co-Authors: Siti Khadijah Kasani, Huei Yin Cheng, Shu Yun Tung, Shujung Chang, Kunhai Yeh, Paul Weiche Hsu, Chungtiang Liang, Wen ChangAbstract:ABSTRACT The Western Reserve (WR) strain of Mature vaccinia Virus contains an A26 envelope protein that mediates Virus binding to cell surface laminin and subsequent endocytic entry into HeLa cells. Removal of the A26 protein from the WR strain Mature Virus generates a mutant, WRΔA26, that enters HeLa cells through plasma membrane fusion. Here, we infected murine bone marrow-derived macrophages (BMDM) with wild-type strain WR and the WRΔA26 mutant and analyzed viral gene expression and cellular innate immune signaling. In contrast to previous studies, in which both HeLa cells infected with WR and HeLa cells infected with WRΔA26 expressed abundant viral late proteins, we found that WR expressed much less viral late protein than WRΔA26 in BMDM. Microarray analysis of the cellular transcripts in BMDM induced by Virus infection revealed that WR preferentially activated type 1 interferon receptor (IFNAR)-dependent signaling but WRΔA26 did not. We consistently detected a higher level of soluble beta interferon secretion and phosphorylation of the STAT1 protein in BMDM infected with WR than in BMDM infected with WRΔA26. When IFNAR-knockout BMDM were infected with WR, late viral protein expression increased, confirming that IFNAR-dependent signaling was differentially induced by WR and, in turn, restricted viral late gene expression. Finally, wild-type C57BL/6 mice were more susceptible to mortality from WRΔA26 infection than to that from WR infection, whereas IFNAR-knockout mice were equally susceptible to WR and WRΔA26 infection, demonstrating that the ability of WRΔA26 to evade IFNAR signaling has an important influence on viral pathogenesis in vivo. IMPORTANCE The vaccinia Virus A26 protein was previously shown to mediate Virus attachment and to regulate viral endocytosis. Here, we show that infection with strain WR induces a robust innate immune response that activates type 1 interferon receptor (IFNAR)-dependent cellular genes in BMDM, whereas infection with the WRΔA26 mutant does not. We further demonstrated that the differential activation of IFNAR-dependent cellular signaling between WR and WRΔA26 not only is important for differential host restriction in BMDM but also is important for viral virulence in vivo. Our study reveals a new property of WRΔA26, which is in regulating host antiviral innate immunity in vitro and in vivo.
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vaccinia Mature Virus fusion regulator a26 protein binds to a16 and g9 proteins of the viral entry fusion complex and dissociates from Mature virions at low ph
Journal of Virology, 2012Co-Authors: Shujung Chang, Aochun Shih, Yinliang Tang, Wen ChangAbstract:Vaccinia Mature Virus enters cells through either endocytosis or plasma membrane fusion, depending on Virus strain and cell type. Our previous results showed that vaccinia Virus Mature virions containing viral A26 protein enter HeLa cells preferentially through endocytosis, whereas Mature virions lacking A26 protein enter through plasma membrane fusion, leading us to propose that A26 acts as an acid-sensitive fusion suppressor for Mature Virus (S. J. Chang, Y. X. Chang, R. Izmailyan R, Y. L. Tang, and W. Chang, J. Virol. 84:8422-8432, 2010). In the present study, we investigated the fusion suppression mechanism of A26 protein. We found that A26 protein was coimmunoprecipitated with multiple components of the viral entry-fusion complex (EFC) in infected HeLa cells. Transient expression of viral EFC components in HeLa cells revealed that vaccinia Virus A26 protein interacted directly with A16 and G9 but not with G3, L5 and H2 proteins of the EFC components. Consistently, a glutathione S-transferase (GST)-A26 fusion protein, but not GST, pulled down A16 and G9 proteins individually in vitro. Together, our results supported the idea that A26 protein binds to A16 and G9 protein at neutral pH contributing to suppression of vaccinia Virus-triggered membrane fusion from without. Since vaccinia Virus extracellular envelope proteins A56/K2 were recently shown to bind to the A16/G9 subcomplex to suppress Virus-induced fusion from within, our results also highlight an evolutionary convergence in which vaccinia viral fusion suppressor proteins regulate membrane fusion by targeting the A16 and G9 components of the viral EFC complex. Finally, we provide evidence that acid (pH 4.7) treatment induced A26 protein and A26-A27 protein complexes of 70 kDa and 90 kDa to dissociate from Mature virions, suggesting that the structure of A26 protein is acid sensitive.
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disulfide bond formation at the c termini of vaccinia Virus a26 and a27 proteins does not require viral redox enzymes and suppresses glycosaminoglycan mediated cell fusion
Journal of Virology, 2009Co-Authors: Yaocheng Ching, Yinliang Tang, Chesheng Chung, Chengyen Huang, Yu Hsia, Wen ChangAbstract:Vaccinia Virus A26 protein is an envelope protein of the intracellular Mature Virus (IMV) of vaccinia Virus. A mutant A26 protein with a truncation of the 74 C-terminal amino acids was expressed in infected cells but failed to be incorporated into IMV (W. L. Chiu, C. L. Lin, M. H. Yang, D. L. Tzou, and W. Chang, J. Virol 81: 2149-2157, 2007). Here, we demonstrate that A27 protein formed a protein complex with the full-length form but not with the truncated form of A26 protein in infected cells as well as in IMV. The formation of the A26-A27 protein complex occurred prior to virion assembly and did not require another A27-binding protein, A17 protein, in the infected cells. A26 protein contains six cysteine residues, and in vitro mutagenesis showed that Cys441 and Cys442 mediated intermolecular disulfide bonds with Cys71 and Cys72 of viral A27 protein, whereas Cys43 and Cys342 mediated intramolecular disulfide bonds. A26 and A27 proteins formed disulfide-linked complexes in transfected 293T cells, showing that the intermolecular disulfide bond formation did not depend on viral redox pathways. Finally, using cell fusion from within and fusion from without, we demonstrate that cell surface glycosaminoglycan is important for Virus-cell fusion and that A26 protein, by forming complexes with A27 protein, partially suppresses fusion.
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vaccinia Virus 4c a26l protein on intracellular Mature Virus binds to the extracellular cellular matrix laminin
Journal of Virology, 2007Co-Authors: Wenling Chiu, Derlii M Tzou, Chilong Lin, Minhsiang Yang, Wen ChangAbstract:Vaccinia Virus intracellular Mature Virus (IMV) binds to glycosaminoglycans (GAGs) on cells via three virion proteins, H3L, A27L, and D8L. In this study, we demonstrated that binding of IMV to BSC40 cells was competitively inhibited by soluble laminin but not by fibronectin or collagen V, suggesting that this cell surface extracellular matrix (ECM) protein may play a role in vaccinia Virus entry. Moreover, IMV infection of GAG− sog9 cells was also inhibited by laminin, demonstrating that virion binding to laminin does not involve a prior interaction with GAGs. Furthermore, comparative envelope protein analyses of wild-type vaccinia Virus strain Western Reserve, which binds to laminin, and of a mutant Virus, IA27L, which does not, showed that the A26L open reading frame (ORF), encoding an envelope protein, was mutated in IA27L, resulting in A26L being absent from the IMV. Expression of the wild-type A26L ORF in IA27L resulted in laminin binding activity. Moreover, recombinant A26L protein bound to laminin in vitro with a high affinity, providing direct evidence that A26L is the laminin binding protein on IMV. In summary, these results reveal a novel role for the vaccinia viral envelope protein A26L in binding to the ECM protein laminin, an association that is proposed to facilitate IMV entry.
Mansun Law - One of the best experts on this subject based on the ideXlab platform.
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quantification of antibody responses against multiple antigens of the two infectious forms of vaccinia Virus provides a benchmark for smallpox vaccination
Nature Medicine, 2006Co-Authors: Mike M Putz, Claire M Midgley, Mansun Law, Geoffrey L SmithAbstract:Smallpox was eradicated without an adequate understanding of how vaccination induced protection. In response to possible bioterrorism with smallpox, the UK government vaccinated ∼300 health care workers with vaccinia Virus (VACV) strain Lister. Antibody responses were analyzed using ELISA for multiple surface antigens of the extracellular enveloped Virus (EEV) and the intracellular Mature Virus (IMV), plaque reduction neutralization and a fluorescence-based flow cytometric neutralization assay. Antibody depletion experiments showed that the EEV surface protein B5 is the only target responsible for EEV neutralization in vaccinated humans, whereas multiple IMV surface proteins, including A27 and H3, are targets for IMV-neutralizing antibodies. These data suggest that it would be unwise to exclude the B5 protein from a future smallpox vaccine. Repeated vaccination provided significantly higher B5-specific and thus EEV-neutralizing antibody responses. These data provide a benchmark against which new, safer smallpox vaccines and residual immunity can be compared.
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entry of the vaccinia Virus intracellular Mature virion and its interactions with glycosaminoglycans
Journal of General Virology, 2005Co-Authors: Gemma C Carter, Mansun Law, Michael Hollinshead, Geoffrey L SmithAbstract:Vaccinia Virus (VACV) produces two distinct enveloped virions, the intracellular Mature Virus (IMV) and the extracellular enveloped Virus (EEV), but the entry mechanism of neither virion is understood. Here, the binding and entry of IMV particles have been investigated. The cell receptors for IMV are unknown, but it was proposed that IMV can bind to glycosaminoglycans (GAGs) on the cell surface and three IMV surface proteins have been implicated in this. In this study, the effect of soluble GAGs on IMV infectivity was reinvestigated and it was demonstrated that GAGs affected IMV infectivity partially in some cells, but not at all in others. Therefore, binding of IMV to GAGs is cell type-specific and not essential for IMV entry. By using electron microscopy, it is demonstrated that IMV from strains Western Reserve and modified Virus Ankara enter cells by fusion with the plasma membrane. After an IMV particle bound to the cell, the IMV membrane fused with the plasma membrane and released the Virus core into the cytoplasm. IMV surface antigen became incorporated into the plasma membrane and was not left outside the cell, as claimed in previous studies. Continuity between the IMV membrane and the plasma membrane was confirmed by tilt-series analysis to orientate membranes perpendicularly to the beam of the electron microscope. This analysis shows unequivocally that IMV is surrounded by a single lipid membrane and enters by fusion at the cell surface.
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the formation and function of extracellular enveloped vaccinia Virus
Journal of General Virology, 2002Co-Authors: Geoffrey L Smith, Alain Vanderplasschen, Mansun LawAbstract:Vaccinia Virus produces four different types of virion from each infected cell called intracellular Mature Virus (IMV), intracellular enveloped Virus (IEV), cell-associated enveloped Virus (CEV) and extracellular enveloped Virus (EEV). These virions have different abundance, structure, location and roles in the Virus life-cycle. Here, the formation and function of these virions are considered with emphasis on the EEV form and its precursors, IEV and CEV. IMV is the most abundant form of Virus and is retained in cells until lysis; it is a robust, stable virion and is well suited to transmit infection between hosts. IEV is formed by wrapping of IMV with intracellular membranes, and is an intermediate between IMV and CEV/EEV that enables efficient Virus dissemination to the cell surface on microtubules. CEV induces the formation of actin tails that drive CEV particles away from the cell and is important for cell-to-cell spread. Lastly, EEV mediates the long-range dissemination of Virus in cell culture and, probably, in vivo. Seven Virus-encoded proteins have been identified that are components of IEV, and five of them are present in CEV or EEV. The roles of these proteins in Virus morphogenesis and dissemination, and as targets for neutralizing antibody are reviewed. The production of several different Virus particles in the VV replication cycle represents a coordinated strategy to exploit cell biology to promote Virus spread and to aid Virus evasion of antibody and complement.
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antibody sensitive and antibody resistant cell to cell spread by vaccinia Virus role of the a33r protein in antibody resistant spread
Journal of General Virology, 2002Co-Authors: Mansun Law, Ruth Hollinshead, Geoffrey L SmithAbstract:The roles of vaccinia Virus (VV) intracellular Mature Virus (IMV), intracellular enveloped Virus (IEV), cell-associated enveloped Virus (CEV) and extracellular enveloped Virus (EEV) and their associated proteins in Virus spread were investigated. The plaques made by VV mutants lacking individual IEV- or EEV-specific proteins (vΔA33R, vΔA34R, vΔA36R, vΔA56R, vΔB5R, vΔF12L and vΔF13L) were compared in the presence of IMV- or EEV-neutralizing antibodies (Ab). Data presented show that for long-range spread, the comet-shaped plaques of VV were caused by the unidirectional spread of EEV probably by convection currents, and for cell-to-cell spread, VV uses a combination of Ab-resistant and Ab-sensitive pathways. Actin tails play a major role in the Ab-resistant pathway, but mutants such as vΔA34R and vΔA36R that do not make actin tails still spread from cell to cell in the presence of Ab. Most strikingly, the Ab-resistant pathway was abolished when the A33R gene was deleted. This effect was not due to alterations in the efficiency of neutralization of EEV made by this mutant, nor due to a deficiency in IMV wrapping to form IEV, which was indispensable for EEV formation by vΔA33R and vΔA34R. We suggest a role for A33R in promoting Ab-resistant cell-to-cell spread of Virus. The roles of the different Virus forms in the VV life-cycle are discussed.
Michael Hollinshead - One of the best experts on this subject based on the ideXlab platform.
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entry of the vaccinia Virus intracellular Mature virion and its interactions with glycosaminoglycans
Journal of General Virology, 2005Co-Authors: Gemma C Carter, Mansun Law, Michael Hollinshead, Geoffrey L SmithAbstract:Vaccinia Virus (VACV) produces two distinct enveloped virions, the intracellular Mature Virus (IMV) and the extracellular enveloped Virus (EEV), but the entry mechanism of neither virion is understood. Here, the binding and entry of IMV particles have been investigated. The cell receptors for IMV are unknown, but it was proposed that IMV can bind to glycosaminoglycans (GAGs) on the cell surface and three IMV surface proteins have been implicated in this. In this study, the effect of soluble GAGs on IMV infectivity was reinvestigated and it was demonstrated that GAGs affected IMV infectivity partially in some cells, but not at all in others. Therefore, binding of IMV to GAGs is cell type-specific and not essential for IMV entry. By using electron microscopy, it is demonstrated that IMV from strains Western Reserve and modified Virus Ankara enter cells by fusion with the plasma membrane. After an IMV particle bound to the cell, the IMV membrane fused with the plasma membrane and released the Virus core into the cytoplasm. IMV surface antigen became incorporated into the plasma membrane and was not left outside the cell, as claimed in previous studies. Continuity between the IMV membrane and the plasma membrane was confirmed by tilt-series analysis to orientate membranes perpendicularly to the beam of the electron microscope. This analysis shows unequivocally that IMV is surrounded by a single lipid membrane and enters by fusion at the cell surface.
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the vaccinia Virus a27l protein is needed for the microtubule dependent transport of intracellular Mature Virus particles
Journal of General Virology, 2000Co-Authors: Christopher M Sanderson, Michael Hollinshead, Geoffrey L SmithAbstract:The vaccinia Virus (VV) A27L gene encodes a 14 kDa protein that is required for the formation of intracellular enveloped Virus (IEV) and, consequently, normal sized plaques. Data presented here show that A27L plays an additional role in VV assembly. When cells were infected with the VV WR32-7/Ind 14K, under conditions that repress A27L expression, transport of intracellular Mature Virus (IMV) from Virus factories was inhibited and some IMV was found in aberrant association with Virus crescents. In contrast, other VV mutants (vΔB5R and vΔF13L) that are defective in IEV formation produce IMV particles that are transported out of Virus factories. This indicated a specific role for A27L in IMV transport. Induction of A27L expression at 10 h post-infection promoted the dispersal of clustered IMV particles, but only when microtubules were intact. Formation of IEV particles was also impaired when cells were infected with WR32-7/14K, a VV strain expressing a mutated form of the A27L protein; however, this mutation did not inhibit intracellular transport of IMV particles. Collectively, these data define two novel aspects of VV morphogenesis. Firstly, A27L is required for both IMV transport and the process of envelopment that leads to IEV formation. Secondly, movement of IMV particles between the Virus factory and the site of IEV formation is microtubule-dependent.
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the extracellular domain of vaccinia Virus protein b5r affects plaque phenotype extracellular enveloped Virus release and intracellular actin tail formation
Journal of Virology, 1998Co-Authors: Elizabeth C Mathew, Christopher M Sanderson, Michael Hollinshead, Geoffrey L SmithAbstract:Vaccinia Virus produces two morphologically distinct forms of infectious Virus, termed intracellular Mature Virus (IMV) and extracellular enveloped Virus (EEV). EEV is important for Virus dissemination within a host and has different surface proteins which bind to cell receptors different from those used by IMV. Six genes are known to encode EEV-specific proteins. One of these, B5R, encodes a 42-kDa glycoprotein with amino acid similarity to members of the complement control protein superfamily and contains four copies of a 50- to 70-amino-acid repeat called the short consensus repeat (SCR). Deletion of B5R causes a small-plaque phenotype, a 10-fold reduction in EEV formation, and Virus attenuation in vivo. In this study, we inserted mutated versions of the B5R gene lacking different combinations of the SCRs into a Virus deletion mutant lacking the B5R gene. The resultant Viruses each formed small plaques only slightly larger than those of the deletion mutant; however, the Virus containing only SCR 1 formed plaques slightly larger than those of Viruses with SCRs 1 and 2 or SCRs 1, 2, and 3. All of these Viruses produced approximately 50-fold more infectious EEV than wild-type Virus and formed comet-shaped plaques under liquid overlay. Despite producing more EEV, the mutant Viruses were unable to induce the polymerization of actin on intracellular Virus particles. The implications of these results for our understanding of EEV formation, release, and infectivity are discussed.
Michael G. Rossmann - One of the best experts on this subject based on the ideXlab platform.
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FlaviViruses have imperfect icosahedral symmetry
Proceedings of the National Academy of Sciences of the United States of America, 2018Co-Authors: Matthew D. Therkelsen, Michael G. Rossmann, Thomas Klose, Frank S. Vago, Wen Jiang, Richard J. KuhnAbstract:FlaviViruses assemble initially in an imMature, noninfectious state and undergo extensive conformational rearrangements to generate Mature Virus. Previous cryo-electron microscopy (cryo-EM) structural studies of flaviViruses assumed icosahedral symmetry and showed the concentric organization of the external glycoprotein shell, the lipid membrane, and the internal nucleocapsid core. We show here that when icosahedral symmetry constraints were excluded in calculating the cryo-EM reconstruction of an imMature flaviVirus, the nucleocapsid core was positioned asymmetrically with respect to the glycoprotein shell. The core was positioned closer to the lipid membrane at the proximal pole, and at the distal pole, the outer glycoprotein spikes and inner membrane leaflet were either perturbed or missing. In contrast, in the asymmetric reconstruction of a Mature flaviVirus, the core was positioned concentric with the glycoprotein shell. The deviations from icosahedral symmetry demonstrated that the core and glycoproteins have varied interactions, which likely promotes viral assembly and budding.
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implications of the picornaVirus capsid structure for polyprotein processing protein nucleic acid catalysis Virus structure polyprotein proteolysis virion morphogenesis maturation
2016Co-Authors: Edward Arnold, Michael G. Rossmann, Ming Luo, Gerrit Vriend, Ann C Palmenbergt, Griffith D Parkst, Martin J H NicklintAbstract:Mature picornaviral proteins are derived by progressive, posttranslational cleavage of a precursor polypro- tein. These cleavages play a role in the control of Virus functions. Although the processed termini are separated by as much as 75 A in the native Virus capsid, the fold and arrangement of polypeptide chains in a protomer before proteolysis are likely to be similar to that found in the Mature Virus. The three-dimensional structures of rhinoVirus and Mengo Virus suggest that the cleavage sites within the proto- meric precursor are in structurally flexible regions. The final proteolytic processing event, maturation of the virion peptide VPO (also called peptide 1AB) appears to occur by an unusual autocatalytic serine protease-type mechanism possibly involv- ing viral RNA basic groups that would serve as proton- abstractors during the cleavage reaction.
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Structure of imMature West Nile Virus.
Journal of virology, 2007Co-Authors: Ying Zhang, Paul R. Chipman, Richard Kuhn, Bärbel Kaufmann, Michael G. RossmannAbstract:The structure of imMature West Nile Virus particles, propagated in the presence of ammonium chloride to block Virus maturation in the low-pH environment of the trans-Golgi network, was determined by cryo-electron microscopy (cryo-EM). The structure of these particles was similar to that of imMature West Nile Virus particles found as a minor component of Mature Virus samples (naturally occurring imMature particles [NOIPs]). The structures of Mature infectious flaviViruses are radically different from those of the imMature particles. The similarity of the ammonium chloride-treated particles and NOIPs suggests either that the NOIPs have not undergone any conformational change during maturation or that the conformational change is reversible. Comparison with the cryo-EM reconstruction of imMature dengue Virus established the locations of the N-linked glycosylation sites of these Viruses, verifying the interpretation of the reconstructions of the imMature flaviViruses.
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structural changes of bacteriophage φ29 upon dna packaging and release
The EMBO Journal, 2006Co-Authors: Ye Xiang, Marc C Morais, Anthony J Battisti, Shelley Grimes, Paul J Jardine, Dwight L Anderson, Michael G. RossmannAbstract:Cryo‐electron microscopy three‐dimensional reconstructions have been made of Mature and of emptied bacteriophage ϕ29 particles without making symmetry assumptions. Comparisons of these structures with each other and with the ϕ29 prohead indicate how conformational changes might initiate successive steps of assembly and infection. The 12 adsorption capable ‘appendages’ were found to have a structure homologous to the bacteriophage P22 tailspikes. Two of the appendages are extended radially outwards, away from the long axis of the Virus, whereas the others are around and parallel to the phage axis. The appendage orientations are correlated with the symmetry‐mismatched positions of the five‐fold related head fibers, suggesting a mechanism for partial cell wall digestion upon rotation of the head about the tail when initiating infection. The narrow end of the head‐tail connector is expanded in the Mature Virus. Gene product 3, bound to the 5′ ends of the genome, appears to be positioned within the expanded connector, which may potentiate the release of DNA‐packaging machine components, creating a binding site for attachment of the tail.
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Visualization of membrane protein domains by cryo-electron microscopy of dengue Virus
Nature Structural & Molecular Biology, 2003Co-Authors: Wei Zhang, Paul R. Chipman, Michael G. Rossmann, Ying Zhang, Suchetana Mukhopadhyay, Jeroen Corver, Peter R Johnson, Timothy S Baker, James H Strauss, Richard J. KuhnAbstract:Improved technology for reconstructing cryo-electron microscopy (cryo-EM) images has now made it possible to determine secondary structural features of membrane proteins in enveloped Viruses. The structure of Mature dengue Virus particles was determined to a resolution of 9.5 Å by cryo-EM and image reconstruction techniques, establishing the secondary structural disposition of the 180 envelope (E) and 180 membrane (M) proteins in the lipid envelope. The α-helical 'stem' regions of the E molecules, as well as part of the N-terminal section of the M proteins, are buried in the outer leaflet of the viral membrane. The 'anchor' regions of E and the M proteins each form antiparallel E-E and M-M transmembrane α-helices, leaving their C termini on the exterior of the viral membrane, consistent with the predicted topology of the unprocessed polyprotein. This is one of only a few determinations of the disposition of transmembrane proteins in situ and shows that the nucleocapsid core and envelope proteins do not have a direct interaction in the Mature Virus.