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At least 19 recordsLinked to original sources
PubMed1987

Difference in capacities for virion-to-virion fusion of young and aged HVJ (Sendai virus): a model of membrane fusion.

Young and aged HVJ virions differ structurally and morphologically due to changes that occur during aging in vitro or in ovo. Young virions soon after their budding off are rod-shaped, rigid and relatively uniform in size, whereas virions that have aged in vitro after their formation are round, nonrigid and variable in size. These changes during aging seem to be due to the variation of M protein, a "skeletal" protein that is associated with both the envelope membrane proteins and nucleocapsid strands in the virions. The capacities for virion-to-virion fusion of young and aged virions were compared to clarify the relation between the membrane fusion and membrane-associating skeletal proteins. On treatment with polyethylene glycol (PEG), aged virions readily fused, forming large virion vesicles, but young virions were resistant to fusion. Further, aged virions fused even on incubation at 37 degrees C without the fusogen. Thus the capacity for virion-to-virion fusion evidently increases during aging of virions. This result suggests that skeletal proteins associating with the biological membrane are important for preventing membrane fusion, and that virion-to-virion fusion is a good model system for use in studies on the mechanism of membrane fusion.

Animals↗
PubMed2001

RNAs extracted from herpes simplex virus 1 virions: apparent selectivity of viral but not cellular RNAs packaged in virions.

Following the lead of recent studies on the presence of RNA in virions of human cytomegalovirus, we investigated the presence and identity of RNAs from purified virions of herpes simple virus 1. To facilitate these studies, we designed primers for all known open reading frames (ORFs) and also constructed cDNA arrays containing probes designed to detect all known transcripts. In the first series of experiments, labeled DNA made by reverse transcription of poly(A)(+) RNA extracted from infected HEp-2 or rabbit skin cells hybridized to all but two of the probes in the cDNA array. A similar analysis of the RNA extracted from purified extracellular virions derived from infected HEp-2 cells hybridized to probes representing 24 of the ORFs. In the second series of analyses, we reverse transcribed and amplified by PCR RNAs from purified intracellular or extracellular virions derived from infected HEp-2 or Vero cell lines. The positive RNAs were retested by PCR with and without prior reverse transcription to ensure that the samples tested were free of contaminating DNA. The results were as follows. (i) Only a fraction of viral ORF transcripts were represented in virion RNA, and only nine RNAs (U(L)10, U(L)34/U(L)35, U(L)36, U(L)42, U(L)48, U(L)51, U(S)1/U(S)1.5, U(S)8.5, and U(S)10/U(S)11) were positive in all RT PCR assays. Of these, seven were positive by hybridization to cDNA arrays. (ii) RNA extracted from cells infected with a mutant virus lacking the U(S)8 to U(S)12 genes yielded results similar to those described above, indicating that U(S)11, a known RNA binding protein, does not play a role in packaging RNA in virions. (iii) Cellular RNAs detected in virions were representative of the abundant cellular RNAs. Last, RNA extracted from virions was translated in vitro and the translation products were reacted with antibody to alphaTIF (VIP16). The immune precipitate contained a labeled protein with the apparent molecular weight of alphaTIF, indicating that at least one mRNA packaged in virions was intact and capable of being translated. The basis for the apparent selectivity in the packaging of the viral RNAs packaged in virions is unknown.

Animals↗
PubMed2004

Modulation of Env content in virions of simian immunodeficiency virus: correlation with cell surface expression and virion infectivity.

Specific mutations were created in the cytoplasmic domain of the gp41 transmembrane protein of simian immunodeficiency virus strain 239 (SIV239). The resultant strains included a mutant in which Env residue 767 was changed to a stop codon, a double mutant in which positions 738 and 739 were changed to stop codons, another mutant in which a prominent endocytosis motif was changed from YRPV to GRPV by the substitution of tyrosine 721, and a final combination mutant bearing Q738stop, Q739stop, and Y721G mutations. The effects of these mutations on cell surface expression, on Env incorporation into virions, and on viral infectivity were examined. The molar ratio of Gag to gp120 of 54:1 that we report here for SIV239 virions agrees very well with the ratio of 60:1 reported previously by Chertova et al. (E. Chertova, J. W. Bess, Jr., B. J. Crise, R. C. Sowder II, T. M. Schaden, J. M. Hilburn, J. A. Hoxie, R. E. Benveniste, J. D. Lifson, L. E. Henderson, and L. O. Arthur, J. Virol. 76:5315-5325, 2002), although they were determined by very different methodologies. Assuming 1,200 to 2,500 Gag molecules per virion, this corresponds to 7 to 16 Env trimers per SIV239 virion particle. Although all of the mutations increased Env levels in virions, E767stop had the most dramatic effect, increasing the Env content per virion 25- to 50-fold. Increased levels of Env content in virions correlated strictly with higher levels of Env expression on the cell surface. The increased Env content with the E767stop mutation also correlated with an increased infectivity, but the degree of change was not proportional: the 25- to 50-fold increase in Env content only increased infectivity 2- to 3-fold. All of the mutants replicated efficiently in the CEMx174 and Rh221-89 cell lines. Although some of these findings have been reported previously, our findings show that the effects of the cytoplasmic domain of gp41 on the Env content in virions can be dramatic, that the Env content in virions correlates strictly with the levels of cell surface expression, and that the Env content in virions can determine infectivity; furthermore, our results define a particular change with the most dramatic effects.

Animals↗
PubMed2004

The raft-promoting property of virion-associated cholesterol, but not the presence of virion-associated Brij 98 rafts, is a determinant of human immunodeficiency virus type 1 infectivity.

Lipid rafts are enriched in cholesterol and sphingomyelin and are isolated on the basis of insolubility in detergents, such as Brij 98 and Triton X-100. Recent work by Holm et al. has shown that rafts insoluble in Brig 98 can be found in human immunodeficiency virus type 1 (HIV-1) virus-like particles, although it is not known whether raft-like structures are present in authentic HIV-1 and it is unclear whether a virion-associated raft-like structure is required for HIV replication. Independently, it was previously reported that virion-associated cholesterol is critical for HIV-1 infectivity, although the specific requirement of virion cholesterol in HIV-1 was not examined. In the present study, we have demonstrated that infectious wild-type HIV-1 contains Brij 98 rafts but only minimal amounts of Triton X-100 rafts. To directly assess the functional requirement of virion-associated rafts and various features of cholesterol on HIV-1 replication, we replaced virion cholesterol with exogenous cholesterol analogues that have demonstrated either raft-promoting or -inhibiting capacity in model membranes. We observed that variable concentrations of exogenous analogues are required to replace a defined amount of virion-associated cholesterol, showing that structurally diverse cholesterol analogues have various affinities toward HIV-1. We found that replacement of 50% of virion cholesterol with these exogenous cholesterol analogues did not eliminate the presence of Brij 98 rafts in HIV-1. However, the infectivity levels of the lipid-modified HIV-1s directly correlate with the raft-promoting capacities of these cholesterol analogues. Our data provide the first direct assessment of virion-associated Brij 98 rafts in retroviral replication and illustrate the importance of the raft-promoting property of virion-associated cholesterol in HIV-1 replication.

Centrifugation, Density Gradient↗
PubMed2026

New form of HPV18 L1 found in native virions contributes to virion stability and infectivity.

UNLABELLED: Current dogma states that papillomavirus virions consist of only one form of the L1 major capsid protein. Human papillomaviruses (HPVs) have been studied using recombinant particles, thereby bypassing the need to grow the virus in a model of differentiating epithelium. Expression vectors engineered to produce L1 utilize a consensus methionine found in many HPV types. However, HPV18 has two additional in-frame methionine residues located 61 and 26 amino acids upstream of the consensus methionine. These methionine residues are excluded from the L1 expression vector used to create the recombinant virus. On the other hand, HPV produced from organotypic raft culture allows virions to be assembled in a differentiating epithelium in the presence of its native promoters and complete genome. Here, we have utilized this system to show that the wild-type HPV18, produced in a more natural setting, translates a larger form of L1 from the upstream methionine, which is 61 amino acid residues upstream from the consensus methionine, and a smaller form from the consensus methionine, with both sizes assembled in the newly formed virion. Ablation of the upstream methionine residue at position 61 altered the virion capsid conformation, and also decreased virion stability and infectivity. IMPORTANCE: The present study investigates whether the papillomavirus virions contain more than one isoform of the L1 major capsid protein. By using organotypic raft cultures that mimic the naturally differentiating epithelium, we revealed that wild-type HPV18 expresses and incorporates two distinct L1 isoforms into the virion, including a previously unrecognized longer form translated from an upstream methionine. Our study has further shown that disruption of the upstream methionine impairs virion stability and infectivity, demonstrating that the longer L1 isoform contributes to proper capsid architecture and viral function. These findings offer a new understanding of HPV capsid biology and have significant implications for the future development of HPV vaccines, diagnostics, and antiviral therapies.

HPV18↗
PubMed2006

Labeling HIV-1 virions with two fluorescent proteins allows identification of virions that have productively entered the target cell.

GFP-Vpr labeled HIV-1 virions have provided a method to visually examine the interactions between the virus and target cell during infection. However, existing methods to discriminate between virions that have been non-specifically endocytosed from those that have productively entered the host cell cytoplasm have remained problematic. Therefore, we examined the ability of a series of membrane-targeted fluorescent fusion protein constructs to be incorporated into virions. We find that a fluorescent protein fusion targeted to the plasma membrane by the addition of the N-terminal 15 amino acid sequence of c-Src (S15) is efficiently packaged into HIV virions. Using fluorescent proteins fused to this sequence, we have generated virions dually labeled with S15-mCherry and GFP-Vpr. Importantly, we can detect the loss of this S15-mCherry membrane signal following fusion. After infection with VSV-g pseudotyped HIV virions, we find a measurable, specific loss of membrane label during infection. This loss of fluorescence is not observed when fusion is prevented using bafilomycin A. This increased ability to discriminate between non-productively endocytosed virions and those actively undergoing steps in the infectious process will facilitate efforts to examine early steps in infection microscopically.

Cell Line↗
PubMed2006

Packaging of the virion host shutoff (Vhs) protein of herpes simplex virus: two forms of the Vhs polypeptide are associated with intranuclear B and C capsids, but only one is associated with enveloped virions.

The virion host shutoff (Vhs) protein (UL41) is a minor component of herpes simplex virus virions which, following penetration, accelerates turnover of host and viral mRNAs. Infected cells contain 58-kDa and 59.5-kDa forms of Vhs, which differ in the extent of phosphorylation, yet only a 58-kDa polypeptide is incorporated into virions. In pulse-chase experiments, the primary Vhs translation product comigrated in sodium dodecyl sulfate-polyacrylamide gel electrophoresis with the 58-kDa virion polypeptide, and could be chased to 59.5 kDa. While both 59.5-kDa and 58-kDa forms were found in nuclear and cytoplasmic fractions, the 59.5-kDa form was significantly enriched in the nucleus. Both forms were associated with intranuclear B and C capsids, yet only the 58-kDa polypeptide was found in enveloped cytoplasmic virions. A 58-kDa form, but not the 59.5-kDa form, was found in L particles, noninfectious particles that contain an envelope and tegument but no capsid. The data suggest that virions contain two populations of Vhs that are packaged by different pathways. In the first pathway, the primary translation product is processed to 59.5 kDa, is transported to the nucleus, binds intranuclear capsids, and is converted to 58 kDa at some stage prior to final envelopment. The second pathway does not involve the 59.5-kDa form or interactions between Vhs and capsids. Instead, the primary translation product is phosphorylated to the 58-kDa virion form and packaged through interactions with other tegument proteins in the cytoplasm or viral envelope proteins at the site of final envelopment.

Animals↗
PubMed2006

Vaccinia virus A6L encodes a virion core protein required for formation of mature virion.

Vaccinia virus A6L is a previously uncharacterized gene that is conserved in all sequenced vertebrate poxviruses. Here, we constructed a recombinant vaccinia virus encoding A6 with an epitope tag and showed that A6 was expressed in infected cells after viral DNA replication and packaged in the core of the mature virion. Furthermore, we showed that A6 was essential for vaccinia virus replication by performing clustered charge-to-alanine mutagenesis on A6, which resulted in two vaccinia virus mutants (vA6L-mut1 and vA6L-mut2) that displayed a temperature-sensitive phenotype. At 31 degrees C, both mutants replicated efficiently; however, at 40 degrees C, vA6L-mut1 grew to a low titer, while vA6L-mut2 failed to replicate. The A6 protein expressed by vA6L-mut2 exhibited temperature-dependent instability. At the nonpermissive temperature, vA6L-mut2 was normal at viral gene expression and viral factory formation, but it was defective for proteolytic processing of the precursors of several major virion proteins, a defect that is characteristic of a block in virion morphogenesis. Electron microscopy further showed that the morphogenesis of vA6L-mut2 was arrested before the formation of immature virion with nucleoid and mature virion. Taken together, our data show that A6 is a virion core protein that plays an essential role in virion morphogenesis.

Cell Line↗
PubMed1996

The cytoplasmic tail of influenza A virus neuraminidase (NA) affects NA incorporation into virions, virion morphology, and virulence in mice but is not essential for virus replication.

In this study, we investigated the role of the conserved neuraminidase (NA) cytoplasmic tail residues in influenza virus replication. Mutants of influenza A virus (A/WSN/33 [H1N1]) with deletions of the NA cytoplasmic tail region were generated by reverse genetics. The resulting viruses, designated NOTAIL, contain only the initiating methionine of the conserved six amino-terminal residues. The mutant viruses grew much less readily and produced smaller plaques than did the wild-type virus. Despite similar levels of NA cell surface expression by the NOTAIL mutants and wild-type virus, incorporation of mutant NA molecules into virions was decreased by 86%. This reduction resulted in less NA activity per virion, leading to the formation of large aggregates of progeny mutant virions on the surface of infected cells. A NOTAIL virus containing an additional mutation (Ser-12 to Pro) in the transmembrane domain incorporated three times more NA molecules into virions than did the NOTAIL parent but approximately half of the amount incorporated by the wild-type virus. However, aggregation of the progeny virions still occurred at the cell surface. All NOTAIL viruses were attenuated in mice. We conclude that the cytoplasmic tail of NA is not absolutely essential for virus replication but exerts important effects on the incorporation of NA into virions and thus on the aggregation and virulence of progeny virus. In addition, the relative abundance of long filamentous particles formed by the NOTAIL mutants, compared with the largely spherical wild-type particles, indicates a role for the NA cytoplasmic tail in virion morphogenesis.

Amino Acid Sequence↗
PubMed1998

Comparison of the native CCMV virion with in vitro assembled CCMV virions by cryoelectron microscopy and image reconstruction.

Cryoelectron microscopy and three-dimensional image reconstruction analysis has been used to determine the structure of native and in vitro assembled cowpea chlorotic mottle virus (CCMV) virions and capsids to 25-A resolution. Purified CCMV coat protein was used in conjunction with in vitro transcribed viral RNAs to assemble RNA 1 only, RNA 2 only, RNA 3/4 only, and empty (RNA lacking) virions. The image reconstructions demonstrate that the in vitro assembled CCMV virions are morphologically indistinguishable from native virions purified from infected plants. The viral RNA (vRNA) is packaged similarly within the different types of virions. The centers of all assembled particles are generally devoid of density and the vRNA packs against the interior surface of the virion shell. The vRNA appears to adopt an ordered conformation at each of the quasi-threefold axes.

Bromovirus↗
PubMed2000

Packageable antiviral therapeutics against human immunodeficiency virus type 1: virion-targeted virus inactivation by incorporation of a single-chain antibody against viral integrase into progeny virions.

To determine their activities as an antiviral agent packageable within virions and suitable for continued expression in cells, we tested a single-chain antibody (scAb) against human immunodeficiency virus type 1 (HIV-1) integrase and its three fusion proteins: fused to viral protein R (scab-Vpr), a double-cassette of the WXXF motif binding to Vpr (scAb-WXXF), and viral major capsid protein (scAb-CA), respectively. Cotransfection of human 293T cells with expression plasmid for scAb-Vpr or -WXXF along with HIV-1 clone pLAI resulted in the production of a normal amount of progeny virions with infectivity decreased by more than 10(3)-fold. Immunoblot analyses showed that scAb-Vpr or -WXXF was associated with virions, whereas scAb or scAb-CA was not, suggesting that scAb-Vpr or -WXXF was incorporated into virions. The incorporation of scAb-WXXF appeared to be Vpr dependent, because the fusion protein was associated with the wild-type but not with Vpr-truncated HIV-1 virions. Since G418-selected HeLa clones carrying expression plasmid for scAb-WXXF were obtained much more frequently than those for scAb-Vpr, scAb-WXXF was inferred to be less toxic to cells than scAb-Vpr. These results suggest that scAb-WXXF may serve as a novel class of antiviral therapeutic that inactivates progeny HIV virions from within.

Antibodies↗
PubMed2003

Cholesterol depletion of human immunodeficiency virus type 1 and simian immunodeficiency virus with beta-cyclodextrin inactivates and permeabilizes the virions: evidence for virion-associated lipid rafts.

Recent evidence suggests that human immunodeficiency virus type 1 (HIV-1) particles assemble and bud selectively through areas in the plasma membrane of cells that are highly enriched with glycosylphosphatidylinositol-anchored proteins and cholesterol, called lipid rafts. Since cholesterol is required to maintain lipid raft structure and function, we proposed that virion-associated cholesterol removal with the compound 2-hydroxy-propyl-beta-cyclodextrin (beta-CD) might be disruptive to HIV-1 and simian immunodeficiency virus (SIV). We examined the effect of beta-CD on the structure and infectivity of cell-free virions. We found that beta-CD inactivated HIV-1 and SIV in a dose-dependent manner and permeabilized the viral membranes, resulting in the loss of mature Gag proteins (capsid, matrix, nucleocapsid, p1, and p6) without loss of the envelope glycoproteins. SIV also lost reverse transcriptase (RT), integrase (IN), and viral RNA. IN appeared to be only slightly diminished in HIV-1, and viral RNA, RT, matrix, and nucleocapsid proteins were retained in HIV-1 but to a much lesser degree. Host proteins located internally in the virus (actin, moesin, and ezrin) and membrane-associated host proteins (major histocompatibility complex classes I and II) remained associated with the treated virions. Electron microscopy revealed that under conditions that permeabilized the viruses, holes were present in the viral membranes and the viral core structure was perturbed. These data provide evidence that an intact viral membrane is required to maintain mature virion core integrity. Since the viruses were not fixed before beta-CD treatment and intact virion particles were recovered, the data suggest that virions may possess a protein scaffold that can maintain overall structure despite disruptions in membrane integrity.

2-Hydroxypropyl-beta-cyclodextrin↗
PubMed2006

Epstein-Barr virus protein kinase BGLF4 is a virion tegument protein that dissociates from virions in a phosphorylation-dependent process and phosphorylates the viral immediate-early protein BZLF1.

Epstein-Barr virus (EBV) BGLF4 is a viral protein kinase that is expressed in the lytic phase of infection and is packaged in virions. We report here that BGLF4 is a tegument protein that dissociates from the virion in a phosphorylation-dependent process. We also present evidence that BGLF4 interacts with and phosphorylates BZLF1, a key viral regulator of lytic infection. These conclusions are based on the following observations. (i) In in vitro tegument release assays, a significant fraction of BGLF4 was released from virions in the presence of physiological NaCl concentrations. (ii) Addition of physiological concentrations of ATP and MgCl(2) to virions enhanced BGLF4 release, but phosphatase treatment of virions significantly reduced BGLF4 release. (iii) A recombinant protein containing a domain of BZLF1 was specifically phosphorylated by purified recombinant BGLF4 in vitro, and BGLF4 altered BZLF1 posttranslational modification in vivo. (iv) BZLF1 was specifically coimmunoprecipitated with BGLF4 in 12-O-tetradecanoylphorbol-13-acetate-treated B95-8 cells and in COS-1 cells transiently expressing both of these viral proteins. (v) BGLF4 and BZLF1 were colocalized in intranuclear globular structures, resembling the viral replication compartment, in Akata cells treated with anti-human immunoglobulin G. Our results suggest that BGLF4 functions not only in lytically infected cells by phosphorylating viral and cellular targets but also immediately after viral penetration like other herpesvirus tegument proteins.

Cell Line↗
PubMed1980

Herpes simplex virus phosphoproteins. II. Characterization of the virion protein kinase and of the polypeptides phosphorylated in the virion.

The protein kinase associated with purified herpes simplex virus 1 and 2 virions partitioned with the capsid-tegument structures and was not solubilized by non-ionic detergents and low, non-inhibitory concentrations of urea. The enzyme required Mg2+ or Mn2+ and utilized ATP or GTP. The activity was enhanced by non-ionic detergents and by Na+ even in the presence of high concentrations of of Mg2+, but not by cyclic nucleotides. The enzyme associated with capsid-tegument structures phosphorylated virion polypeptides only; exogenously added substrates (acidic and basic histones, casein, phosphovitin, protamine, and bovine serum albumin) were not phosphorylated. The major phosphorylated species were virion polypeptides (VP) 1-2, 4, 11-12, 13-14, 18.7, 18.8 and 23. VP 18.7 and VP 18.8 have not been previously detected, but may be phosphorylated forms of polypeptides co-migrating with VP 19. Of the remainder, only VP 23 has been previously identified as a capsid protein; the others are constituents of the tegument or of the under surface of the virion envelope. The distribution of the phosphate bound to viral polypeptides varied depending on the Mg2+ concentration and pH. In the absence of dithiothreitol, in vitro phosphate exchange was demonstrable in VP 23 and to a lesser extent in two other polypeptides on sequential phosphorylation frist with saturating amounts off unlabeled ATP and then with [gamma-32P]ATP. Analysis of the virion polypeptides specified by herpes simplex virus 1 X herpes simplex virus 2 recombinants indicates that the genes specifying the polypeptides which serve as a substrate for the protein kinase map in the unique sequences near the left and right reinterated DNA sequences of the L component.

Cations↗
PubMed1983

Interferon induces the production of membrane protein-deficient and infectivity-defective vesicular stomatitis virions through interference in the virion assembly process.

The reduced rate of synthesis, maturation and degradation as well as the level of accumulation of the intracellular virus proteins in VSV-infected cells may account for the overall reduction (less than 10-fold) of progeny virion yield due to interferon (IFN); however, the deficiency of the virions proteins, G and M, which apparently caused a drastic loss of infectivity of these progeny virions (about 1000-fold) cannot be easily explained, because the concentrations of G and M proteins relative to other virus proteins were not reduced in the cell. In fact, intracellular M protein was significantly increased. Moreover, the virus proteins in IFN-treated and control cells were synthesized and accumulated in large excess of the amount incorporated into the released virions. The reduction in the intracellular activity of GlcNac-P-P-Dol transferase did not appear to play a direct role in the antiviral mechanism in this system. Our results, however, do suggest that the deficiency of G and M proteins in the virion is related to specific inhibition of the incorporation of either or both of these proteins in the virus assembly process.

Interferons↗
PubMed1974

Sendai virion transcriptase complex: polyeptide composition and inhibition by virion envelope proteins.

Sendai virions, disrupted in 2% Triton X-100 in 1 M KCl, were separated into nucleocapsids and envelope proteins by centrifugation. The nucleocapsids, representing 46% of the virion proteins, had a buoyant density of 1.29 gm/cm(3) in D(2)O sucrose. RNA-dependent transcriptase activity associated with them had a ninefold greater specific activity than transcriptase assayed in unfractionated detergent-disrupted virions. These enzyme-active nucleocapsids contained only two polypeptides, the largest virion polypeptide (molecular weight 75,000) and the nucleocapsid structure unit (molecular weight 60,000). Virion envelope proteins, either glycoproteins or nonglycosylated matrix protein, inhibited nucleocapsid-associated polymerase activity; brief heat denaturation abolished their inhibitory activity. Yeast RNA stimulated nucleocapsid-associated enzyme, suggesting that stimulatory polyanions act at the enzyme-template level.

Cell-Free System↗
PubMed2002

Inhibition of host ER glucosidase activity prevents Golgi processing of virion-associated bovine viral diarrhea virus E2 glycoproteins and reduces infectivity of secreted virions.

Recently, it was shown that replication of bovine viral diarrhea virus (BVDV) is sensitive to inhibitors of host ER glucosidases. Consistent with these findings, we report that incubation of BVDV-infected MDBK cells with the glucosidase inhibitor n-butyl-deoxynojirimycin (nB-DNJ) reduced BVDV yields by 70- to 100-fold (n = 27), while having no effect on MDBK cell viability. However, the 70- to 100-fold reduction in infectious virus was associated with only a 2-fold reduction in genomic RNA synthesis and secretion of enveloped virus particles. Analysis of secreted virions showed that in the absence of glucosidase inhibitor, approximately 50% of the virion-associated BVDV E2 glycoprotein was resistant to endoglycosidase H (endo H) digestion, whereas intracellular E2 was completely sensitive to endo H digestion. In the presence of glucosidase inhibitor, virion-associated E2 and intracellular E2 were completely sensitive to endo H digestion. Taken together, these results suggest that BVDV is secreted through a Golgi-mediated pathway and that host ER glucosidase activity is required for production of infectious virions and Golgi processing of envelope E2 protein during virus egress.

1-Deoxynojirimycin↗
PubMed2004

Temperature-sensitive mutants in the vaccinia virus 4b virion structural protein assemble malformed, transcriptionally inactive intracellular mature virions.

Two noncomplementing vaccinia virus temperature-sensitive mutants, Cts8 and Cts26, were mapped to the A3L gene, which encodes the major virion structural protein, 4b. The two ts mutants display normal patterns of gene expression, DNA replication, telomere resolution, and protein processing during infection. Morphogenesis during mutant infections is normal through formation of immature virions with nucleoids (IVN) but appears to be defective in the transition from IVN to intracellular mature virus (IMV). In mutant infections, aberrant particles that have the appearance of malformed IMV accumulate. The mutant particles are wrapped in Golgi-derived membranes and exported from cells. Purified mutant particles are indistinguishable from wt particles in protein and DNA composition; however, they are defective in a permeabilized-virion-directed transcription reaction despite containing significant (Cts8) or even normal (Cts26) levels of specific transcription enzymes. These results indicate that the 4b protein is required for proper metamorphosis of IMV from IVN and that proper organization of the IMV structure is required to produce a transcriptionally active virion particle.

Animals↗