scholarly journals Cell Surface Proteoglycans Are Necessary for A27L Protein-Mediated Cell Fusion: Identification of the N-Terminal Region of A27L Protein as the Glycosaminoglycan-Binding Domain

1998 ◽  
Vol 72 (10) ◽  
pp. 8374-8379 ◽  
Author(s):  
Jye-Chian Hsiao ◽  
Che-Sheng Chung ◽  
Wen Chang
Keyword(s):  
Virus Infection ◽  
Cell Surface ◽  
Vaccinia Virus ◽  
Heparan Sulfate ◽  
Cell Fusion ◽  
Binding Domain ◽  
Terminal Region ◽  
Virus Binding ◽  
Vaccinia Virus Infection

ABSTRACT We previously showed that vaccinia virus infection of BSC40 cells was blocked by soluble heparin, suggesting that cell surface heparan sulfate mediates vaccinia virus binding (C.-S. Chung, J.-C. Hsiao, Y.-S. Chang, and W. Chang, J. Virol. 72:1577–1585, 1998). In this study, we extended our previous work and demonstrated that soluble A27L protein bound to heparan sulfate on cells and interfered with vaccinia virus infection at a postbinding step. In addition, we investigated the structure of A27L protein that provides for its binding to heparan sulfate on cells. A mutant of A27L protein, named D-A27L, devoid of a cluster of 12 amino acids rich in basic residues, was constructed. In contrast to the soluble A27L protein, purified D-A27L protein was inactive in all of our assays, including binding to heparin in vitro, binding to heparan sulfate on cells, and the ability to block virus infection. These data demonstrated that the N-terminal region acts as a glycosaminoglycan (GAG)-binding domain critical for A27L protein binding to cells. Previously A27L protein was thought to be involved in fusion of virus-infected cells induced by acid treatment. When we investigated whether cell surface GAGs also participate in A27L-dependent fusion, our results indicated that soluble A27L protein blocked cell fusion, whereas D-A27L protein did not. Taken together, the results therefore demonstrated that A27L-mediated cell fusion is triggered by its interaction with cell surface GAGs through the N-terminal domain.

scholarly journals A27L Protein Mediates Vaccinia Virus Interaction with Cell Surface Heparan Sulfate

1998 ◽  
Vol 72 (2) ◽  
pp. 1577-1585 ◽  
Author(s):  
Che-Sheng Chung ◽  
Jye-Chian Hsiao ◽  
Yuan-Shau Chang ◽  
Wen Chang
Keyword(s):  
Virus Infection ◽  
Cell Surface ◽  
Vaccinia Virus ◽  
Heparan Sulfate ◽  
Mammalian Cells ◽  
Myxoma Virus ◽  
Cowpox Virus ◽  
Virus Binding ◽  
Vaccinia Virus Infection

ABSTRACT Vaccinia virus has a wide host range and infects mammalian cells of many different species. This suggests that the cell surface receptors for vaccinia virus are ubiquitously expressed and highly conserved. Alternatively, different receptors are used for vaccinia virus infection of different cell types. Here we report that vaccinia virus binds to heparan sulfate, a glycosaminoglycan (GAG) side chain of cell surface proteoglycans, during virus infection. Soluble heparin specifically inhibits vaccinia virus binding to cells, whereas other GAGs such as condroitin sulfate or dermantan sulfate have no effect. Heparin also blocks infections by cowpox virus, rabbitpox virus, myxoma virus, and Shope fibroma virus, suggesting that cell surface heparan sulfate could be a general mediator of the entry of poxviruses. The biochemical nature of the heparin-blocking effect was investigated. Heparin analogs that have acetyl groups instead of sulfate groups also abolish the inhibitory effect, suggesting that the negative charges on GAGs are important for virus infection. Furthermore, BSC40 cells treated with sodium chlorate to produce undersulfated GAGs are more refractory to vaccinia virus infection. Taken together, the data support the notion that cell surface heparan sulfate is important for vaccinia virus infection. Using heparin-Sepharose beads, we showed that vaccinia virus virions bind to heparin in vitro. In addition, we demonstrated that the recombinant A27L gene product binds to the heparin beads in vitro. This recombinant protein was further shown to bind to cells, and such interaction could be specifically inhibited by soluble heparin. All the data together indicated that A27L protein could be an attachment protein that mediates vaccinia virus binding to cell surface heparan sulfate during viral infection.

scholarly journals Vaccinia Virus Envelope H3L Protein Binds to Cell Surface Heparan Sulfate and Is Important for Intracellular Mature Virion Morphogenesis and Virus Infection In Vitro and In Vivo

2000 ◽  
Vol 74 (7) ◽  
pp. 3353-3365 ◽  
Author(s):  
Chi-Long Lin ◽  
Che-Sheng Chung ◽  
Hans G. Heine ◽  
Wen Chang
Keyword(s):  
Cell Surface ◽  
Vaccinia Virus ◽  
Heparan Sulfate ◽  
Mutant Virus ◽  
Wild Type Virus ◽  
Type Virus ◽  
Wild Type ◽  
Virion Morphogenesis

ABSTRACT An immunodominant antigen, p35, is expressed on the envelope of intracellular mature virions (IMV) of vaccinia virus. p35 is encoded by the viral late gene H3L, but its role in the virus life cycle is not known. This report demonstrates that soluble H3L protein binds to heparan sulfate on the cell surface and competes with the binding of vaccinia virus, indicating a role for H3L protein in IMV adsorption to mammalian cells. A mutant virus defective in expression of H3L (H3L−) was constructed; the mutant virus has a small plaque phenotype and 10-fold lower IMV and extracellular enveloped virion titers than the wild-type virus. Virion morphogenesis is severely blocked and intermediate viral structures such as viral factories and crescents accumulate in cells infected with the H3L− mutant virus. IMV from the H3L− mutant virus are somewhat altered and less infectious than wild-type virions. However, cells infected by the mutant virus form multinucleated syncytia after low pH treatment, suggesting that H3L protein is not required for cell fusion. Mice inoculated intranasally with wild-type virus show high mortality and severe weight loss, whereas mice infected with H3L− mutant virus survive and recover faster, indicating that inactivation of the H3L gene attenuates virus virulence in vivo. In summary, these data indicate that H3L protein mediates vaccinia virus adsorption to cell surface heparan sulfate and is important for vaccinia virus infection in vitro and in vivo. In addition, H3L protein plays a role in virion assembly.

scholarly journals LEUKOCYTES AND INTERFERON IN THE HOST RESPONSE TO VIRAL INFECTIONS

1965 ◽  
Vol 121 (6) ◽  
pp. 1001-1018 ◽  
Author(s):  
Lowell A. Glasgow
Keyword(s):  
Virus Infection ◽  
Vaccinia Virus ◽  
Viral Infections ◽  
Complete Control ◽  
Degree Of Protection ◽  
Vaccinia Virus Infection ◽  
Primary Mouse ◽  
Vitro Model ◽  
Spread Of Infection

1. Investigation of the role of leukocytes in vaccinia virus infection is reported in an in vitro model, in the absence of an immune response. 2. Mouse leukocytes were shown to be capable of inhibiting the progression of vaccinia virus infection in primary mouse embryo fibroblast cultures. The degree of protection varied from slowing of spread of infection to complete control of the infection with eventual elimination of detectable virus and recovery of the culture. 3. Interferon production by leukocytes is thought to be an important factor in the observed protective effect.

scholarly journals Profile of natural killer cells after a previous natural Vaccinia virus infection in an in vitro viral re-exposure

2014 ◽  
Vol 184 ◽  
pp. 20-29 ◽  
Author(s):  
Eduardo Augusto dos Santos Moreira-Silva ◽  
Daniela Carla Medeiros-Silva ◽  
Juliana de Assis Silva Gomes ◽  
Flávio Guimarães da Fonseca ◽  
Rodrigo Correa-Oliveira
Keyword(s):  
Natural Killer Cells ◽  
Virus Infection ◽  
Vaccinia Virus ◽  
Natural Killer ◽  
Killer Cells ◽  
Vaccinia Virus Infection

scholarly journals A Genome-Wide Haploid Genetic Screen Identifies Heparan Sulfate-Associated Genes and the Macropinocytosis Modulator TMED10 as Factors Supporting Vaccinia Virus Infection

2019 ◽  
Vol 93 (13) ◽  
Author(s):  
Rutger D. Luteijn ◽  
Ferdy van Diemen ◽  
Vincent A. Blomen ◽  
Ingrid G. J. Boer ◽  
Saravanan Manikam Sadasivam ◽  
...  
Keyword(s):  
Virus Infection ◽  
Vaccinia Virus ◽  
Heparan Sulfate ◽  
Host Cell ◽  
Host Factors ◽  
Virus Infectivity ◽  
Sequencing Analysis ◽  
Vaccinia Virus Infection ◽  
Genome Wide ◽  
A Genome

ABSTRACTVaccinia virus is a promising viral vaccine and gene delivery candidate and has historically been used as a model to study poxvirus-host cell interactions. We employed a genome-wide insertional mutagenesis approach in human haploid cells to identify host factors crucial for vaccinia virus infection. A library of mutagenized HAP1 cells was exposed to modified vaccinia virus Ankara (MVA). Deep-sequencing analysis of virus-resistant cells identified host factors involved in heparan sulfate synthesis, Golgi organization, and vesicular protein trafficking. We validated EXT1, TM9SF2, and TMED10 (TMP21/p23/p24δ) as important host factors for vaccinia virus infection. The critical roles of EXT1 in heparan sulfate synthesis and vaccinia virus infection were confirmed. TM9SF2 was validated as a player mediating heparan sulfate expression, explaining its contribution to vaccinia virus infection. In addition, TMED10 was found to be crucial for virus-induced plasma membrane blebbing and phosphatidylserine-induced macropinocytosis, presumably by regulating the cell surface expression of the TAM receptor Axl.IMPORTANCEPoxviruses are large DNA viruses that can infect a wide range of host species. A number of these viruses are clinically important to humans, including variola virus (smallpox) and vaccinia virus. Since the eradication of smallpox, zoonotic infections with monkeypox virus and cowpox virus are emerging. Additionally, poxviruses can be engineered to specifically target cancer cells and are used as a vaccine vector against tuberculosis, influenza, and coronaviruses. Poxviruses rely on host factors for most stages of their life cycle, including attachment to the cell and entry. These host factors are crucial for virus infectivity and host cell tropism. We used a genome-wide knockout library of host cells to identify host factors necessary for vaccinia virus infection. We confirm a dominant role for heparin sulfate in mediating virus attachment. Additionally, we show that TMED10, previously not implicated in virus infections, facilitates virus uptake by modulating the cellular response to phosphatidylserine.

scholarly journals A genome-wide haploid genetic screen for essential factors in vaccinia virus infection identifies TMED10 as regulator of macropinocytosis

2018 ◽  
Author(s):  
Rutger David Luteijn ◽  
Ferdy R van Diemen ◽  
Vincent A Blomen ◽  
Ingrid GJ Boer ◽  
Saravanan Manikam Sadasivam ◽  
...  
Keyword(s):  
Virus Infection ◽  
Vaccinia Virus ◽  
Heparan Sulfate ◽  
Critical Role ◽  
Host Factors ◽  
Sequencing Analysis ◽  
Vaccinia Virus Infection ◽  
Genome Wide ◽  
A Genome ◽  
Viral Vaccine

Vaccinia virus is a promising viral vaccine and gene delivery candidate, and has historically been used as a model to study poxvirus-host cell interactions. We employed a genome-wide insertional mutagenesis approach in human haploid cells to identify host factors crucial for vaccinia virus infection. A library of mutagenized HAP1 cells was exposed to Modified Vaccinia Virus Ankara (MVA). Deep-sequencing analysis of virus-resistant cells identified host factors involved in heparan sulfate synthesis, Golgi organization, and vesicular protein trafficking. We validated EXT1, TM9SF2 and TMED10 TMP21/p23/p24δ) as important host factors for vaccinia virus infection. The critical role of EXT1 in heparan sulfate synthesis and vaccinia virus infection was confirmed. TM9SF2 was validated as a player mediating heparan sulfate expression, explaining its contribution to vaccinia virus infection. In addition, TMED10 was found to be crucial for virus-induced plasma membrane blebbing and phosphatidylserine-induced macropinocytosis, suggesting that TMED10 regulates actin cytoskeleton remodelling necessary for virus infection.

scholarly journals Pharmacodynamics of Cidofovir for Vaccinia Virus Infection in an In Vitro Hollow-Fiber Infection Model System

2008 ◽  
Vol 53 (1) ◽  
pp. 129-135 ◽  
Author(s):  
James J. McSharry ◽  
Mark R. Deziel ◽  
Kris Zager ◽  
Qingmei Weng ◽  
George L. Drusano
Keyword(s):  
Virus Infection ◽  
Vaccinia Virus ◽  
Hollow Fiber ◽  
Antiviral Treatment ◽  
Drug Application ◽  
Model System ◽  
Smallpox Vaccination ◽  
Infection Model ◽  
Vaccinia Virus Infection

ABSTRACT Variola major virus remains a potent weapon of bioterror. There is currently an investigational-new-drug application for cidofovir for the therapy of variola major virus infections. Stittelaar and colleagues compared the levels of effectiveness of postexposure smallpox vaccination (Elstree-RIVM) and antiviral treatment with cidofovir or an acyclic nucleoside phosphonate analogue {6-[2-(phosphonomethoxy)alkoxy]-2,4-diaminopyrimidine (HPMPO-DAPy)} after lethal intratracheal infection of cynomolgus monkeys with monkeypox virus, a variola virus surrogate. Their results demonstrated that either compound was more effective than vaccination with the Ellstree vaccine (K. J. Stittelaar et al., Nature 439:745-748, 2006). An unanswered question is how to translate this information into therapy for poxvirus infections in people. In a proof-of-principle study, we used a novel in vitro hollow-fiber infection model system to determine the pharmacodynamics of vaccinia virus infection of HeLa-S3 cells treated with cidofovir. Our results demonstrate that the currently licensed dose of cidofovir of 5 mg/kg of body weight weekly with probenecid (which ameliorates nephrotoxicity) is unlikely to provide protection for patients intentionally exposed to Variola major virus. We further demonstrate that the antiviral effect is independent of the schedule of drug administration. Exposures (area under the concentration-time curve) to cidofovir that will have a robust protective effect will require doses that are 5 to 10 times that currently administered to humans. Such doses may cause nephrotoxicity, and therefore, approaches that include probenecid administration as well as schedules of administration that will help ameliorate the uptake of cidofovir into renal tubular epithelial cells need to be considered when addressing such treatment for people.

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