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Results for “Virus Replication”

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At least 469 records · Page 26Linked to original sources

PubMed1993

Topoisomerase I and II activities are required for Epstein-Barr virus replication.

The roles of topoisomerases I and II in Epstein-Barr virus (EBV) replication were investigated using Raji cells infected with EBV. The topoisomerase II inhibitor ellipticine inhibited the synthesis of EBV polypeptides at concentrations which did not affect total protein synthesis. Slot blot analysis of total cellular DNA showed that camptothecin and ellipticine inhibited replication of progeny EBV DNA in superinfected Raji cells at concentrations which did not inhibit synthesis of EBV early polypeptides prerequisite for EBV DNA replication. Analysis of the structure of EBV DNA termini demonstrated that both inhibitors affected the replicating EBV DNA. Gardella gel electrophoresis showed that both inhibitors affected the formation of the linear form of EBV DNA. However, restriction analysis of EBV DNA in superinfected Raji cells demonstrated that both inhibitors degraded neither endogenous nor exogenous EBV DNA. Cell viability was not affected by either inhibitor at the concentrations tested. These findings suggest that topoisomerase II is required for expression of the EBV genome and that both topoisomerases I and II are involved in replication of the EBV genome during the lytic phase of the life cycle. The effects of topoisomerase inhibitors on the circular form of EBV DNA during virus replication are discussed.

Camptothecin↗

PubMed1999

Enhancement of UV-induced cytotoxicity by the adeno-associated virus replication proteins.

Adeno-associated virus (AAV) normally requires co-infection of a helper virus to complete its life cycle. However, under conditions of cellular stress, such as treatment with carcinogens or ultraviolet (UV) light, a permissive intracellular environment is established and AAV completes its replicative cycle producing low levels of progeny virus. AAV DNA replication is dependent upon viral replication proteins, Rep78 and Rep68. The detailed mechanism by which these proteins interact with host cell factors is unknown. We have used a cell line (Neo6) that inducibly expresses the AAV Rep proteins to study their effects on cells that have undergone UV-induced DNA damage. Induction of Rep protein expression immediately after a sub-lethal dose of UV irradiation resulted in rapid cell killing. Those cells that die had chromatin condensation while cellular membranes remained intact, suggesting that concurrent Rep expression and UV damage induces an apoptosis-like response. However, we did not observe any DNA degradation. Thus we believe that the combination of Rep expression and UV irradiation induces cell death that shares some of the characteristics of apoptosis. UV irradiation and Rep expression induced an increase in the level of the CDK inhibitor, p21Cip, and the appearance of modified forms of both p21Cip and Bcl-2. Alteration of normal expression of these cytostatic/apoptotic proteins provides insight into the intracellular targets of the AAV replication proteins.

Apoptosis↗

PubMed1977

Localization of the restrictive event of EMC virus replication in semi-permissive monkey and monkey-mouse hybrid cells.

Encephalomyocarditis (EMC) virus replication was investigated in permissive mouse MKS cells, semi-permissive monkey CV1 cells, and in somatic monkey-mouse MKCVIII hybrid cells whose permissiveness is under the negative control of the simian genome. We found that in CV1 cells the synthesis of both single- and double-stranded virus RNAs was restricted. In contrast, in semi-permissive hybrid Cl4/3 cells only the single-stranded virus RNA was synthesized in small amounts, whereas the double-stranded virus RNA accumulated late after infection. The synthesis of virus polyribosomes and virus polypeptides was lowered in semipermissive conditions. In the presence of quaternary ammonium ions, the synthesis of EMC virus was partially relieved in CV1 cells. Thus, it can be postulated that a defective function in the replication complex is involved in the restrictive event.

Animals↗

PubMed2000

Effects of deletion or stringent repression of the H3L envelope gene on vaccinia virus replication.

The C-terminal membrane anchor protein encoded by the H3L open reading frame of vaccinia virus is located on the surfaces of intracellular mature virions. To investigate the role of the H3L protein, we constructed deletion (vH3Delta) and inducible (vH3i) null mutants. The H3L protein was not detected in lysates of cells infected with vH3Delta or vH3i in the absence of inducer. Under these conditions, plaques were small and round instead of large and comet shaped, indicative of decreased virus replication or cell-to-cell spread. The mutant phenotype was correlated with reduced yields of infectious intra- and extracellular virus in one-step growth experiments. The defect in vH3i replication could not be attributed to a role of the H3L protein in virus binding, internalization, or any event prior to late gene expression. Electron microscopic examination of cells infected with vH3Delta or vH3i in the absence of inducer revealed that virion assembly was impaired, resulting in a high ratio of immature to mature virus forms with an accumulation of crescent membranes adjacent to granular material and DNA crystalloids. The absence of the H3L protein did not impair the membrane localization of virion surface proteins encoded by the A27L, D8L, and L1R genes. The wrapping of virions and actin tail formation were not specifically blocked, but there was an apparent defect in low-pH-mediated syncytium formation that could be attributed to decreased virus particle production. The phenotypes of the H3L deletion and repression mutants were identical to each other but differed from those produced by null mutations of genes encoding other vaccinia virus membrane components.

Animals↗

PubMed2005

G-protein signaling triggered by R5 human immunodeficiency virus type 1 increases virus replication efficiency in primary T lymphocytes.

The binding of R5 envelope to CCR5 during human immunodeficiency virus type 1 (HIV-1) entry provokes cell activation, which has so far been considered to have no effect on virus replication, since signaling-defective CCR5 molecules have been shown to function normally as HIV-1 coreceptors on transformed cells or mitogen-stimulated T lymphocytes. As the background state of activation of these cells might have biased the results, we performed experiments using the same approach but with nonactivated primary T lymphocytes. We now report that the single R126N mutation in the DRY motif, involved in G-protein coupling, results in a signaling-defective CCR5 coreceptor with a drastically impaired capacity to support HIV-1 infection.

Cells, Cultured↗

PubMed1989

Rev-induced modulation of Nef protein underlies temporal regulation of human immunodeficiency virus replication.

The replication of human immunodeficiency virus type 1 (HIV-1) requires the concerted action of two virus-encoded transactivator proteins, Tat and Rev, and is in turn moderated by the viral transcriptional repressor Nef. We show here that the phenotype of a Rev- HIV-1 provirus was nonreplicating and was distinguished by accumulation of Nef protein and reduced Tat function. Provirus defective in both the rev and nef genes (Rev-Nef-) was also nonreplicating but had normal Tat function. Trans-complementation of the Rev- mutant with Rev caused a decrease of both the steady-state level and the rate of synthesis of Nef. This was accompanied by enhanced synthesis of viral structural proteins. Rev induced even greater levels of virus production from the Rev-Nef- double mutant. In contrast, exogenous Rev did not augment virus production from wild-type provirus. Virus production from Rev- and Rev-Nef- mutants induced by Rev was repressed by exogenous Nef. The repression induced by Nef could not be reversed by exogenous Rev. The ability of Rev to modulate Nef expression solely from the provirus, and thereby relieve the Nef-mediated inhibition of transcription from the viral long terminal repeat, reveals a delicate balance of the functions of these two proteins that might underlie the switch between latency and reactivation.

Base Sequence↗

PubMed2002

Grapevine fanleaf virus replication occurs on endoplasmic reticulum-derived membranes.

Infection by Grapevine fanleaf nepovirus (GFLV), a bipartite RNA virus of positive polarity belonging to the Comoviridae family, causes extensive cytopathic modifications of the host endomembrane system that eventually culminate in the formation of a perinuclear "viral compartment." We identified by immunoconfocal microscopy this compartment as the site of virus replication since it contained the RNA1-encoded proteins necessary for replication, newly synthesized viral RNA, and double-stranded replicative forms. In addition, by using transgenic T-BY2 protoplasts expressing green fluorescent protein in the endoplasmic reticulum (ER) or in the Golgi apparatus (GA), we could directly show that GFLV replication induced a depletion of the cortical ER, together with a condensation and redistribution of ER-derived membranes, to generate the viral compartment. Brefeldin A, a drug known to inhibit vesicle trafficking between the GA and the ER, was found to inhibit GFLV replication. Cerulenin, a drug inhibiting de novo synthesis of phospholipids, also inhibited GFLV replication. These observations imply that GFLV replication depends both on ER-derived membrane recruitment and on de novo lipid synthesis. In contrast to proteins involved in viral replication, the 2B movement protein and, to a lesser extent, the 2C coat protein were not confined to the viral compartment but were transported toward the cell periphery, a finding consistent with their role in cell-to-cell movement of virus particles.

Cell Line↗

PubMed1997

Respiratory syncytial virus triggers synthesis of IL-6 in BALB/c mouse alveolar macrophages in the absence of virus replication.

Cytokines produced by alveolar macrophages are likely involved in the regulation of the immune response arising from respiratory syncytial virus (RSV) infection. Both infectious and UV-inactivated RSV were effective in inducing BALB/c mouse alveolar macrophages to synthesize increased levels of IL-6 mRNA and secreted IL-6 protein. No increase in IL-1beta (either mRNA or secreted protein) was observed. The augmented production of IL-6 was activated by purified virus and was reduced by pretreating virus with virus-neutralizing antiserum, demonstrating a requirement for virus in the enhanced IL-6 response. The results suggest that the exposure of BALB/c alveolar macrophages to small quantities of RSV (in the absence of detectable virus replication) is sufficient to trigger IL-6 production. The finding that UV-inactivated virus was effective in triggering IL-6 production by mouse alveolar macrophages is similar to that reported in human alveolar macrophages, providing further validation of the BALB/c mouse as a useful animal model for human RSV infection.

Animals↗

PubMed1997

Formation of plant RNA virus replication complexes on membranes: role of an endoplasmic reticulum-targeted viral protein.

The mechanisms that direct positive-stranded RNA virus replication complexes to plant and animal cellular membranes are poorly understood. We describe a specific interaction between a replication protein of an RNA plant virus and membranes in vitro and in live cells. The tobacco etch virus (TEV) 6 kDa protein associated with membranes as an integral protein via a central 19 amino acid hydrophobic domain. In the presence or absence of other viral proteins, fluorescent fusion proteins containing the 6 kDa protein associated with large vesicular compartments derived from the endoplasmic reticulum (ER). Infection by TEV was associated with a collapse of the ER network into a series of discrete aggregated structures. Viral RNA replication complexes from infected cells were also associated with ER-like membranes. Targeting of TEV RNA replication complexes to membranous sites of replication is proposed to involve post-translational interactions between the 6 kDa protein and the ER.

Amino Acid Sequence↗

PubMed1996

Reactivated fulminant hepatitis B virus replication after bone marrow transplantation: clinical course and possible treatment with ganciclovir.

A female chronic hepatitis B virus carrier (HBV-DNA negative) suffered from simultaneous hepatitis B virus and cytomegalovirus reactivation after in vivo T cell depletion preceding transplantation of an in vitro T cell depleted marrow graft for treatment of acute leukaemia. Interstitial pneumonia developing after bone marrow transplantation was successfully treated with ganciclovir (day 13 until day 46). The initially unnoticed extensive hepatitis B virus replication finally led to clinical hepatitis (day 85) and liver failure (day 96). Liver transplantation was performed, but the patient died from septicaemia. Retrospective analysis of hepatitis B virus DNA revealed that the HBV replication started immediately after T cell depletion and was completely suppressed during ganciclovir administration. Screening for HBV-DNA seems to be mandatory in comparable cases, and antiviral chemotherapy should be seriously considered.

Adult↗

PubMed2005

An experiment on standardized cell culture assay in assessing the activities of Composite Artemisia Capillaris Tablets against hepatitis B virus replication in vitro.

OBJECTIVE: To explore the activities of Composite Artemisia Capillaris Tablet (CACT) against hepatitis B virus replication in vitro. METHODS: By means of radioimmunoassay (RIA), Dot blot and Southern blot, the surface and e antigen production of 2.2.15 cells, HBV DNA in 2.2.15 cell culture medium and that in 2.2.15 cells were examined respectively. RESULTS: HBsAg, HBeAg values of 2.2.15 cells treated by CACT were lower than those of the control, the HBV DNA quantities in culture medium and in 2.2.15 cells decreased as compared with those cells with no treatment by CACT given to them. CONCLUSION: CACT could inhibit HBV DNA replication, showing its potential antiviral activity in hepatitis B treatment.

Cell Line, Tumor↗

PubMed2004

[The inhibitory effects on hepatitis B virus replication by stable expression of DN mutants of hepatitis B virus X gene pRev X-GFP].

OBJECTIVE: Persistent replication of hepatitis B virus (HBV) is one of the major obstacles in HBV infection treatment. Reduction or clearance of HBV propagation would be one of the aims of HBV therapy. The drugs approved in clinical used such as nucleotide analogs or interferon, were limited effects on HBV replication. The newly developing gene therapy method, dominant negative mutants, were be used as new promising HBV therapy strategy, and a dominant negative mutant of HBVX gene pRev X-GFP which we have reported in our previous study has some effects both on HBV replication and expression in transient expression, but the effects were interfered by persistent secretion of HBV in HepG2 2.2.15 cells without transfection pRev X-GFP in the experiment. To make sure the effects of dominant negative mutant of pRev X-GFP, we established a HBX DN stable express cell clone, and evaluated the effects of HBX dominant negative mutant on HBV replication. METHODS: The X gene mutant, in which a specific point mutation of 3'-end ATG to AAG and fused with human green fluorescence protein (GFP) were cloned into pRev TRE vector, assigned to pRev HBX-GFP dominant mutant (pRev X-GFP). And the plasmid contains the wild type X gene or GFP gene was cloned into the same vector to construct pRev Xwt, pRev GFP constructs. All the constructs then transfected into HepG2 2.2.15 cells by liposome. After 7 days resistance selection of hygromycin (300 microg/ml), and cell clones which stable expression HBX-GFP, HBXwt, GFP were obtained. After reseeding of 106 cells of each clones in 12 wells with a 12 well cell plate and another 12 wells 2.2.15 cell were serve as blank control. The cells and media were harvested after cultured in DMEM with 10% FBS for 3 days. HBV-related DNA was assayed by dot blot and Southern blot. RESULTS: The 100% expression of pRev HBX-GFP, GFP and wild type X constructs were obtained. The stable expressed HBX-GFP can significantly reduce HBV DNA level both in cell media and cells by dot blot and Southern blot analysis, but not for pRev Xwt and pRev GFP. CONCLUSION: The dominant negative mutant pRev HBX-GFP can significantly inhibit the HBV gene expression. It also suggested that X gene might be one of promising target for HBV gene therapy.

Carcinoma, Hepatocellular↗

PubMed2004

The Tobacco mosaic virus 126-kDa protein associated with virus replication and movement suppresses RNA silencing.

Systemic symptoms induced on Nicotiana tabacum cv. Xanthi by Tobacco mosaic virus (TMV) are modulated by one or both amino-coterminal viral 126- and 183-kDa proteins: proteins involved in virus replication and cell-to-cell movement. Here we compare the systemic accumulation and gene silencing characteristics of TMV strains and mutants that express altered 126- and 183-kDa proteins and induce varying intensities of systemic symptoms on N. tabacum. Through grafting experiments, it was determined that M(IC)1,3, a mutant of the masked strain of TMV that accumulated locally and induced no systemic symptoms, moved through vascular tissue but failed to accumulate to high levels in systemic leaves. The lack of M(IC)1,3 accumulation in systemic leaves was correlated with RNA silencing activity in this tissue through the appearance of virus-specific, approximately 25-nucleotide RNAs and the loss of fluorescence from leaves of transgenic plants expressing the 126-kDa protein fused with green fluorescent protein (GFP). The ability of TMV strains and mutants altered in the 126-kDa protein open reading frame to cause systemic symptoms was positively correlated with their ability to transiently extend expression of the 126-kDa protein:GFP fusion and transiently suppress the silencing of free GFP in transgenic N. tabacum and transgenic N. benthamiana, respectively. Suppression of GFP silencing in N. benthamiana occurred only where virus accumulated to high levels. Using agroinfiltration assays, it was determined that the 126-kDa protein alone could delay GFP silencing. Based on these results and the known synergies between TMV and other viruses, the mechanism of suppression by the 126-kDa protein is compared with those utilized by other originally characterized suppressors of RNA silencing.

Biological Transport↗

PubMed1992

Untranslatable transcripts of the tobacco etch virus coat protein gene sequence can interfere with tobacco etch virus replication in transgenic plants and protoplasts.

Transgenic tobacco plants which express untranslatable sense or antisense forms of the tobacco etch virus potyvirus (TEV) coat protein (CP) gene sequence have been generated. One of seven transgenic plant lines expressing a CP gene antisense transcript showed an attenuation of symptoms when inoculated with TEV. Three of ten transgenic plant lines expressing untranslatable sense transcripts did not develop symptoms when inoculated with TEV. These lines were resistant to either aphid or mechanically transmitted TEV. In contrast to CP-mediated resistance reported for other viruses, resistance was (1) mediated by an RNA molecule; (2) TEV-specific (i.e., "broad-spectrum resistance" was not observed); (3) independent of inoculum levels; (4) not dependent on plant size and; (5) due to decreased levels of virus replication. Protoplast experiments were used to demonstrate that resistant plant lines did not support the production of virus protein and progeny virus at wild-type levels.

Animals↗

PubMed1987

Mutational analysis of the human immunodeficiency virus: the orf-B region down-regulates virus replication.

Mutations were made by recombinant DNA techniques in an infectious molecular clone of the human immunodeficiency virus San Francisco isolate 2 (HIVSF2) [formerly the prototype isolate of the acquired immunodeficiency syndrome-associated retrovirus (ARV-2)]. The effect of these changes on the replicative and cytopathologic properties of the virus was studied by transfecting modified virus clones into cultured human cells. Mutations in the gag, pol, env, and tat regions precluded virus replication and cytopathology in lymphoid cells. A mutation in orf-A dramatically reduced but did not abolish virus replication. Mutant viruses with deletions in the orf-B region were highly cytopathic and replicated to approximately 5-fold higher levels than wild-type virus. They also produced approximately 5-fold more viral DNA in infected lymphoid cells than did wild-type virus. Thus, the orf-B region may function to down-regulate virus replication. This mutational analysis of the HIVSF2 genome is a means of assessing genes regulating viral replication and cytopathology.

Cloning, Molecular↗

PubMed1987

Novel acyclic adenosine analogs inhibit Epstein-Barr virus replication.

The effect of three new acyclic adenosine analogs, (S)-9-(3-hydroxy-2-phosphonylmethoxypropyl)adenine [(S)-HPMPA], 9-(2-phosphonylmethoxyethyl)adenine (PMEA), and (S)-9-(2,3-dihydroxypropyl)adenine [(S)-DHPA], on Epstein-Barr virus (EBV) replication was studied. Both (S)-HPMPA and PMEA but not (S)-DHPA effectively inhibited EBV DNA replication in virus-producer P3HR-1 cells and in latently infected Raji cells superinfected with P3HR-1 virus, as determined by cRNA-DNA hybridization and density gradient centrifugation. The 50% effective doses for inhibiting virus replication were 0.08 and 1.1 microM for (S)-HPMPA and PMEA, respectively. Both drugs were cytostatic but not cytotoxic to the cells at a concentration as high as 100 microM. These results indicate that (S)-HPMPA and PMEA are potent and selective anti-EBV agents in vitro.

Adenine↗

PubMed1975

Irreversible inhibition of herpes simplex virus replication in BSC-1 cells by zinc ions.

Zinc sulfate added to the medium of herpes simplex virus-infected BSC-1 cells, at a concentration of 0.1 mM, inhibited the synthesis of infectious virus progeny by 95 to 96%. A concentration of 0.2 mM zinc sulfate inhibited herpes simplex virus synthesis by 99.8% as determined by centrifugation in sucrose gradients and by plaque assay on BSC-1 monolayers. The inhibition of herpes simplex virus replication in BSC-1 cells was found to be irreversible.

Cell Line↗

PubMed1994

Hepatitis C virus replication and antibodies to structural and nonstructural viral proteins in chronic hepatitis C.

The correlation between hepatitis C virus replication and antibodies to both structural (core) and nonstructural (C100-3) hepatitis C virus proteins (anti-HCVcore and anti-C100-3, respectively) was assessed. The concentration of serum hepatitis C virus RNA was determined by a competitive reverse transcription-polymerase chain reaction assay, and antibody titers were determined by endpoint dilution. No correlation was found between viremic levels and antibody titers in 42 chronic hepatitis C patients. At the end of a 6-month course of interferon-alpha therapy, 18 patients became negative for hepatitis C virus RNA. In the other 24 patients, post-treatment viremic levels ranged from 10(-6.5)-10(0.5) of pretreatment levels. Both anti-C100-3 and anti-HCVcore frequently decreased in patients whose viremic levels dropped to the negative range or to < 10(-2) of pretreatment levels. Anti-C100-3 decreased in all such cases (25/25), while anti-HCVcore decreased in 18/25 (72%) (p < 0.01), indicating that anti-C100-3 is more likely to decrease following suppression of viral replication than anti-HCVcore. These data suggest that hepatitis C virus antibodies may serve as a marker of suppression of viremia following interferon therapy even in patients who do not clear the virus.

Adult↗