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At least 19 recordsLinked to original sources
PubMed1998

Examination of potential inhibitors of hepatitis A virus uncoating.

Hepatitis A virus (HAV) replication in BS-C-1 cells was studied in the presence of ten potential uncoating inhibitors. Strong inhibition of HAV replication was only observed in the presence of the phenothiazine compound chlorpromazine and the lysosomotropic agent chloroquine, but not by other lysosomotropic agents. Chlorpromazine and chloroquine were found to prevent virus uncoating. Chlorpromazine is known to inhibit endocytosis of non- clathrin-coated vesicles. Chloroquine is a weak base amine, and thought to inhibit virus replication by preventing endosomal acidification. These results therefore suggest that entry of HAV in BS-C-1 cells does not depend on the low pH encountered in the clathrin-coated endocytic entry pathway. A possible role of calcium ions in mediating viral uncoating is discussed, as calcium ions were found to destabilize HAV particles in vitro.

Animals↗
PubMed1982

Influenza virus uncoating in infected cells and effect of rimantadine.

Uncoating of influenza virus (strain WSN) in MDCK cells was studied by following the fate of the virus labelled with radioactive precursors. The accumulation of subviral components of input virus was observed in nuclear-associated cytoplasm (NAC) obtained by treatment of the nuclei with citric acid. Two types of subviral components were found there, ribonucleoproteins (RNPs) and larger subviral particles (SVP) containing RNPs in association with M protein. SVP, with different relative amounts of M protein, were revealed in NAC, suggesting that M protein was gradually released from RNPs. The released RNPs entered the nuclei while M protein accumulated within perinuclear membranes. Thus, SVP could be regarded as probable intermediates in virus uncoating. Rimantadine prevented the release of M protein from RNPs and their penetration into the nuclei provoking the accumulation of subviral components in NAC.

Adamantane↗
PubMed1994

Mechanisms of virus uncoating.

In a virus particle, the genome is highly condensed and protected by proteins and membrane bilayers. Before it can be replicated in a new host cell, uncoating must take place. Recent studies on enveloped and nonenveloped animal viruses indicate that uncoating occurs through complex, multistep processes triggered by virus-host-cell interactions.

Animals↗
PubMed1994

Role of virion M2 protein in influenza virus uncoating: specific reduction in the rate of membrane fusion between virus and liposomes by amantadine.

The anti-influenza virus drug amantadine was shown to reduce the rate of fusion of liposomes with influenza A viruses whose replication is inhibited by this drug. The fusion with amantadine-resistant viruses was unaffected. Experiments with reassortant and mutant viruses showed that this effect was linked to the M2 protein and not to the haemagglutinin of the virus. The proton ionophore monensin, on the other hand, substantially increased the rate of fusion of the viruses tested. These results indicate that the kinetics of virus-liposome fusion can be modulated by the virus M2 protein, the target of amantadine action, and it is postulated that the M2 ion channel functions by transporting protons into the virion interior and facilitating virus uncoating.

Amantadine↗
PubMed1991

Influence of different ionic and pH environments on structural alterations of poliovirus and their possible relation to virus uncoating.

Poliovirus eclipse products were totally precipitated from infected HeLa cells after different times of infection by using TCA, suggesting that cellular enzymic digestion of parental proteins was not involved in virus uncoating. In an investigation of poliovirus thermal stability in vitro, progressive degradation of native virus into 80S empty capsids occurred upon incubation at 37 degrees C in a buffer of low ionic strength containing 20 mM-Tris-HCl pH 7.5, whereas in Eagle's medium or in the presence of L cells degradation was very slow. Degradation was faster at alkaline than at acid pH. Furthermore, liberation of the viral RNA was prevented and 135S particles were produced upon treatment of virus at 37 degrees C in 20-mM-Tris-HCl pH 7.5 containing 2 mM-CaCl2. Although the poliovirus receptor is able to induce conformational alterations of the capsid, low ion concentration could contribute to virus uncoating as well.

Capsid↗
PubMed1980

Rimantadine hydrochloride blocks the second step of influenza virus uncoating.

Two steps of influenza virus A/WSN uncoating in MDCK cells are described. The products of the first step are cores which accumulate in the nuclear-associated cytoplasm. The products of the second step are ribonucleoproteins which penetrate the nuclei. Rimantadine blocks the second step without interfering with the first one.

Adamantane↗
PubMed1986

Antirhinovirus compound 44,081 R.P. inhibits virus uncoating.

44,081 R.P., or 2-[(1,5,10,10a-tetrahydro-3H-thiazolo[3,4-b]isoquinolin- 3-ylidine)amino]-4-thiazole acetic acid, is a compound which selectively inhibits rhinovirus in cell cultures. The compound, which was unable to inactivate infectivity of virions, appeared to act prior to RNA and protein synthesis without affecting adsorption or penetration of virus in MRC5 cells. In contrast, it protected intracellular [5-3H]uridine-labeled virions against the effects of RNase treatment, indicating that inhibition of virus uncoating is the mode of action of 44 081 R.P.

Adsorption↗
PubMed1999

Low temperature and pressure stability of picornaviruses: implications for virus uncoating.

The family Picornaviridae includes several viruses of great economic and medical importance. Poliovirus replicates in the human digestive tract, causing disease that may range in severity from a mild infection to a fatal paralysis. The human rhinovirus is the most important etiologic agent of the common cold in adults and children. Foot-and-mouth disease virus (FMDV) causes one of the most economically important diseases in cattle. These viruses have in common a capsid structure composed of 60 copies of four different proteins, VP1 to VP4, and their 3D structures show similar general features. In this study we describe the differences in stability against high pressure and cold denaturation of these viruses. Both poliovirus and rhinovirus are stable to high pressure at room temperature, because pressures up to 2.4 kbar are not enough to promote viral disassembly and inactivation. Within the same pressure range, FMDV particles are dramatically affected by pressure, with a loss of infectivity of more than 4 log units observed. The dissociation of polio and rhino viruses can be observed only under pressure (2.4 kbar) at low temperatures in the presence of subdenaturing concentrations of urea (1-2 M). The pressure and low temperature data reveal clear differences in stability among the three picornaviruses, FMDV being the most sensitive, polio being the most resistant, and rhino having intermediate stability. Whereas rhino and poliovirus differ little in stability (less than 10 kcal/mol at 0 degrees C), the difference in free energy between these two viruses and FMDV was remarkable (more than 200 kcal/mol of particle). These differences are crucial to understanding the different factors that control the assembly and disassembly of the virus particles during their life cycle. The inactivation of these viruses by pressure (combined or not with low temperature) has potential as a method for producing vaccines.

Adult↗
PubMed1979

Inhibition of influenza virus uncoating by rimantadine hydrochloride.

In freeze-thaw lysates of MDCK cells infected with 32P-labeled influenza virus A/WSN in the presence of added RNase, acid-precipitable radioactivity diminished to about 50% of initial values within 90 min after a 1-h virus adsorption period. A similar preparation containing rimantadine at a concentration of 50 micrograms/ml exhibited only a 10% reduction in acid-precipitable radioactivity. These findings suggest that rimantadine interferes with uncoating of influenza virus in infected cells.

Adamantane↗
PubMed1981

Proteolytic cleavage of VP1 in 'A' particles of coxsackievirus B3 does not appear to mediate virus uncoating by HeLa cells.

'A' particle of Coxsackievirus B3 were generated from native virus by heating and purified by sucrose gradient centrifugation. These particles were found to be similar to 'A' particles formed by elution from cellular receptors of HeLa cells. Electrophoretic analysis of [35S]methionine-labelled 'A' particles revealed that treatment of the particles with chymotrypsin resulted in the cleavage of VP1 and the formation of a cleavage product which migrated between VP2 and VP3. Analysis of the protease-treated material on sucrose gradients revealed a ribonuclease-sensitive particle which sedimented more slowly than an 'A' particle. This particle apparently degraded to release the viral RNA, thereby providing an in vitro model for protease-mediated uncoating of 'A' particles. The subviral particles of Coxsackievirus B3 were found to be immunoprecipitable with heterotypic Coxsackievirus group B antisera, thereby providing a method for the recovery of products produced in the cell early in infection. Infected cells which had been treated to remove unreacted virus were disrupted, an the lysates were reacted with heterotypic antisera. Analysis of the precipitated material revealed that no cleavage products were formed and no polypeptides were lost. Therefore, it appears that proteolysis is not involved in the uncoating of Coxsackievirus B3 in infected cells.

Capsid↗
PubMed2003

Structural analysis of human rhinovirus complexed with ICAM-1 reveals the dynamics of receptor-mediated virus uncoating.

Intercellular adhesion molecule 1 (ICAM-1) functions as the cellular receptor for the major group of human rhinoviruses, being not only the target of viral attachment but also the mediator of viral uncoating. The configurations of HRV3-ICAM-1 complexes prepared both at 4 degrees C and physiological temperature (37 degrees C) were analyzed by cryoelectron microscopy and image reconstruction. The particle diameters of two complexes (with and without RNA) representing uncoating intermediates generated at 37 degrees C were each 4% larger than that of those prepared at 4 degrees C. The larger virus particle arose by an expansive movement of the capsid pentamers along the fivefold axis, which loosens interprotomer contacts, particularly at the canyon region where the ICAM-1 receptor bound. Particle expansion required receptor binding and preceded the egress of the viral RNA. These observations suggest that receptor-mediated uncoating could be a consequence of restrained capsid motion, where the bound receptors maintain the viral capsid in an expanded open state for subsequent genome release.

Binding Sites↗
PubMed1999

Two determinants in the capsid of a persistent type 3 poliovirus exert different effects on mutant virus uncoating.

Mutant polioviruses (PV) have been previously found to be capable of establishing persistent infections in HEp-2c cells. Together, two amino acid substitutions in the viral capsid of a type 3 poliovirus (PV-3), at positions VP213 and VP1(290), are sufficient to confer the persistent phenotype to a normally lytic virus. When susceptible cells are infected, the double mutant T7L+2L(13)1N(290) undergoes unique conformational changes in the capsid, modifying its sedimentation coefficient from 160S to 147S. In the present study, we have further investigated mutant PV decapsidation and, in particular, the effect of each determinant independently. Our results indicate that the novel 147S form was also generated by a mutant carrying only the determinant 1N290. This form was not produced as a result of inherent capsid instability and it was generated only upon specific PV-host cell interactions. The second viral determinant, 2L13, also modified receptor-induced conformational changes, although differently from 1N290.

Capsid↗
PubMed1997

Growth of the parvovirus minute virus of mice MVMp3 in EL4 lymphocytes is restricted after cell entry and before viral DNA amplification: cell-specific differences in virus uncoating in vitro.

Two murine parvoviruses with genomic sequences differing only in 33 nucleotides (8 amino acids) in the region coding for the capsid proteins show different host cell specificities: MVMi grows in EL4 T lymphocytes and MVMp3 grows in A9 fibroblasts. In this study we compared the courses of infections with these two viruses in EL4 cells in order to investigate at which step(s) the infection process of MVMp3 is interrupted. The two viruses bound equally well to EL4 cells, and similar amounts of MVMi and MVMp3 input virion DNA appeared in the nuclear fractions of EL4 cells 1 h after infection. However, double-stranded replicative-form (RF) DNA of the two viruses appeared at different times, at 10 h postinfection with MVMi and at 24 h postinfection with MVMp3. The amount of MVMp3 RF DNA detected at 24 h was very small because it was produced only in a tiny subset of the population of EL4 cells that proved to be permissive for MVMp3. Replication of double-stranded viral DNA in EL4 cells was measured after transfection of purified RF DNA, cloned viral DNA, and cloned viral DNA with a mutation preventing synthesis of the capsid proteins. In each of these cases, DNA replication was comparable for MVMi and MVMp3. Production of virus particles also appeared to be similar after transfection of the two types of RF DNA into EL4 cells. Conversion of incoming 32P-labeled single-stranded MVM DNA to 32P-labeled double-stranded RF DNA was detected only after RF DNA amplification, indicating that few molecules serve as templates for viral DNA amplification. We showed that extracts of EL4 cells contain a factor which can destabilize MVMi virions but not MVMp3 by testing the sensitivity of viral DNA to DNase and by CsCl gradient analyses of viral particles. We therefore conclude that the MVMp3 life cycle is arrested after the transport of virions to the nucleus and prior to the replication of RF DNA, most likely at the stage of viral decapsidation.

Animals↗
PubMed1987

The assay, purification and properties of vaccinia virus-induced uncoating protein.

Vaccinia virus cores prepared in vitro can be uncoated by incubation with extracts of cells infected with vaccinia virus, as determined by the conversion of the genome to DNase susceptibility. This uncoating activity had all the characteristics of the corresponding in vivo activity and of the agent responsible for non-genetic reactivation. Thus, it was not induced by heat-inactivated virus, nor was it produced in the presence of inhibitors of RNA or protein synthesis. The uncoating protein induced by cowpox virus will uncoat vaccinia virus cores. The uncoating protein was concentrated from infected cell extracts by ultrafiltration and purified by gel filtration and ion-exchange and affinity chromatography. It was characterized as a trypsin-like protease with a mol. wt. of 23,050. Cores treated with the purified uncoating protein had an altered sedimentation rate but no differences between treated and untreated cores were detected by electron microscopy or polyacrylamide gel electrophoresis.

Carrier Proteins↗
PubMed1992

Role of ribosomes in Semliki Forest virus nucleocapsid uncoating.

The mechanism by which Semliki Forest virus nucleocapsids are uncoated was analyzed in living cells and in vitro. In BHK-21 cells, uncoating occurred with virtually complete efficiency within 1 to 2 min after the nucleocapsids entered the cytoplasm. It was inhibited by monensin, which blocks nucleocapsid penetration from endosomes. As previously shown for Sindbis virus (G. Wengler and G. Wengler, Virology 134:435-442, 1984), the capsid proteins from incoming nucleocapsids became associated with ribosomes. The ribosome-bound capsid proteins were distributed throughout the cytoplasm, while the viral RNA remained associated with vacuolar membranes. Using purified nucleocapsids and ribosomes in vitro, we established that ribosomes alone were sufficient for uncoating. Their role was to release the capsid proteins from nucleocapsids and irreversibly sequester them, in a process independent of energy and translation. The process was stoichiometric rather than catalytic, with a maximum of three to six capsid proteins bound to each ribosome. More than 80% of the capsid proteins could thus be removed from the viral RNA, resulting in the formation of nucleocapsid remnants whose sedimentation coefficients progressively decreased from 140S to 80S as uncoating proceeded.

Animals↗
PubMed1977

Genetic analysis of host range mutant viruses suggests an uncoating defect in simian virus 40-resistant monkey cells.

Host range mutations that permit simian virus 40 (SV40) to grow with increased efficiency on SV40-resistant monkey cells have been positioned within the viral B/C gene by a mapping method that relies on the coupling of specific DNA fragments. Pairs of restriction endonucleases that each cleave SV40 DNA at only one site were used to generate pairs of specific DNA fragments. Corresponding pairs of fragments were purified from host range mutant and wild-type DNA and joined in known combinations to determine the location of the host range mutations. The map position of the host range mutations was confirmed by using the same technique to generate and couple genetically marked viral DNA fragments to produce the predicted double mutants. Three different double mutants were constructed that carry both host range and temperature-sensitive A mutations. The mutations in three independently isolated host range mutant viruses are located at very close, perhaps identical, sites, because no wild type viruses were produced from the cell-mediated repair of pairwise heteroduplexes between them. The location of these host range mutations suggests that their phenotype results from mutational alteration of the major capsid protein, the product of the B/C gene. In addition it was demonstrated that monkey cells can efficiently join appropriate pairs of restriction endonuclease fragments intracellularly to produce infectious genomes. That reaction has been partially characterized. The general utility of fragment coupling (in vitro and in vivo) and heteroduplex repair for constructing and analyzing multiple mutants of SV40 is discussed.

Cell Transformation, Viral↗
PubMed1998

Inhibition of foot and mouth disease virus (FMDV) uncoating by a plant-derived peptide isolated from Melia azedarach L leaves.

Meliacine (MA), a peptide isolated from leaves of the high plant Melia azedarach L inhibited the multiplication of foot and mouth disease virus (FMDV) in BHK-21 cells. In this report, we establish that the MA-inhibitable process takes place within the first hour of the viral reproductive cycle. MA had no virucidal effect and did not affect adsorption and penetration of the virus in cells. In experiments with neutral red-labeled virus, it was found that MA significantly suppressed the development of photoresistance of the virus in infected cells. In untreated cultures nearly all virus which adsorbed to cells was uncoated within 1 h at 37 degrees C, whereas in treated cultures, even after 3 h only 3% of the virus was uncoated. Labeling of BHK-21 cells with acridine orange showed that MA affects the pH of intracellular acidic vesicles. Therefore, it is concluded that MA prevents the process of uncoating of FMDV in BHK-21 cells by inhibiting vacuolar acidification.

Animals↗