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At least 811 records · Page 45Linked to original sources
PubMed2006

Vaccinia-virus-induced cellular contractility facilitates the subcellular localization of the viral replication sites.

Poxviruses, such as vaccinia virus (VV), replicate their DNA in endoplasmic-reticulum-enclosed cytoplasmic sites. Here, we compare the dynamics of the VV replication sites with those of the attenuated strain, modified VV Ankara (MVA). By live-cell imaging, small, early replication sites of both viruses undergo motility typical of microtubule (MT)-motor-mediated movement. Over time, growing replication sites of VV collect around the nucleus in a MT-dependent fashion, whereas those of MVA remain mostly scattered in the cytoplasm. Surprisingly, blocking the dynein function does not impair the perinuclear accumulation of large VV replication sites. Live-cell imaging demonstrates that in contrast to small replication sites, large sites do not display MT-motor-mediated motility. Instead, VV infection induces cellular contractility that facilitates the collection of growing replication sites around the nucleus. In a subset of cells (30-40%), this VV-induced contractility is alternated by phases of directed cell migration, suggesting that the two processes may be linked. The MVA-infected cells do not display contractility or cell migration, supporting the idea that these cellular activities facilitate the efficient accumulation of the VV replication sites around the nucleus. We propose that the recently described cytoskeletal rearrangements induced by VV are a prerequisite for the observed cell contractility and migration activities that apparently contribute to the organization of the complex cytoplasmic life cycle of VV.

Animals↗
PubMed1993

Activity of a novel quinoxaline derivative against human immunodeficiency virus type 1 reverse transcriptase and viral replication.

S-2720 [6-chloro-3,3-dimethyl-4-(isopropenyloxycarbonyl)-3,4- dihydroquinoxalin-2(1H)-thione], a quinoxaline derivative, was found to be a very potent inhibitor of both human immunodeficiency virus type 1 reverse transcriptase (HIV-1 RT) activity and HIV-1 replication in tissue culture. Like other nonnucleoside RT inhibitors, S-2720 does not affect the HIV-2 RT. A S-2720-resistant virus was selected and shown to possess a mutation within the RT-coding region that has not previously been described. Notably, this mutation gives rise to a dramatic decrease in enzyme activity. S-2720, therefore, belongs to a new class of RT inhibitors that bind differently to the RT than other known nonnucleoside RT inhibitors. As no toxic effects were observed with S-2720 in mice, these quinoxaline derivatives deserve further evaluation to prove their potency as possible therapeutic agents for HIV-1 infection.

Acquired Immunodeficiency Syndrome↗
PubMed1965

Sulfasymazine.

Explore the source record for details and available documents.

Escherichia coli Infections↗
PubMed2002

Viral replication and host gene expression in alveolar macrophages infected with Ebola virus (Zaire strain).

In order to characterize the cellular response to and identify potential diagnostic markers for the early detection of Ebola virus, an in vitro culture system involving nonhuman primate alveolar macrophages was developed. Ebola virus replication in the alveolar macrophages was characterized by plaque assay, immunohistochemical analysis, and in situ hybridization. Fluorogenic 5'-nuclease assays specific for nonhuman primate proinflammatory cytokines and chemokines were designed and used to evaluate mRNA transcription in macrophages infected with Ebola virus. Transient increases in cytokine and chemokine mRNA levels were observed immediately following exposure to Ebola virus. At 2 h postexposure, levels of cytokine and chemokine mRNAs were markedly reduced. Although Ebola virus infection of alveolar macrophages failed to induce a sustained increase in proinflammatory cytokine and chemokine mRNA transcription (potentially reducing the use of these markers as diagnostic tools), the fluorogenic 5'-nuclease assays developed may have prognostic value for individuals infected with Ebola virus. Recently published data have indicated that persons who remain asymptomatic after exposure to Ebola virus are capable of mounting an early proinflammatory cytokine response and that those who become clinically ill are not. If implemented immediately after exposure, these assays could be used to predict which individuals will be more likely to remain asymptomatic as opposed to those who will be more likely to develop clinical signs and eventually succumb to the virus.

Animals↗
PubMed1989

Inhibition of RNase H activity and viral replication by single mutations in the 3' region of Moloney murine leukemia virus reverse transcriptase.

Selected conserved amino acids in the putative RNase H domain of reverse transcriptase (RT) were modified in a molecularly cloned infectious provirus and in a Moloney murine leukemia virus RT expression vector by site-directed mutagenesis. Substitution of either of two conserved aspartic acid residues in proviral DNA prevented production of infectious particles in transfected NIH 3T3 cells, and the same modifications depressed RT-associated RNase H activity by more than 25-fold with little or no effect on polymerase activity.

Amino Acid Sequence↗
PubMed1996

Spontaneous mutations in the human immunodeficiency virus type 1 gag gene that affect viral replication in the presence of cyclosporins.

Human immunodeficiency virus type 1 mutants that are resistant to inhibition by cyclosporins arise spontaneously in vitro during propagation in a HeLa-CD4+ cell line in the presence of a nonimmunosuppressive analog of cyclosporin A. Interestingly, the phenotype of all of the mutants examined is drug resistant and drug dependent, with both cyclosporin A and its analog. Four independently isolated mutants have been analyzed genetically by construction of recombinant proviruses in the NL4-3 parental strain background and subsequent testing of the chimeric viruses in HeLa cells. The cyclosporin-resistant, cyclosporin-dependent phenotype consistently transfers with a 1.3-kb fragment of gag, within which the four mutants share one of two possible single amino acid exchanges in a proline-rich stretch in the capsid domain of Pr55gag. These mutants provide the first evidence that mutations in human immunodeficiency virus type 1 gag confer resistance to cyclosporins; however, replication is conditional on the presence of the drug. In the T-cell line CEM, replication of the recombinant mutant viruses is also cyclosporin dependent. The drug-dependent replication in HeLa cells is stringent, and in the absence of cyclosporin only revertant viruses with the parental phenotype grow out of cultures infected with cyclosporin-dependent virus. In at least one isolate examined, the revertant phenotype appears to be due to suppressor mutations near the proline-rich region.

Amino Acid Sequence↗
PubMed1996

Woodchuck hepatitis virus X protein is present in chronically infected woodchuck liver and woodchuck hepatocellular carcinomas which are permissive for viral replication.

The woodchuck hepatitis virus (WHV) X gene (WHx) is required for infectivity of WHV in woodchucks, and the gene encodes a broadly acting transcription factor. Several lines of evidence from cell culture and transgenic mice suggest that X proteins can promote hepatocarcinogenesis. To determine whether WHx-encoded proteins are present during persistent infection and hepatocellular carcinoma (HCC) in woodchucks, we surveyed livers and HCCs from a panel of WHV carrier woodchucks for the presence of WHx by utilizing an immunoprecipitation-Western blot (immunoblot) procedure. We detected a single 15.5-kDa WHx gene product in 100% of the persistently infected livers but not in livers from animals which had recovered from acute infection or in those of uninfected woodchucks. Analysis of HCCs revealed that all of the tumors which contained WHV replication intermediates were also positive for WHx. In contrast, WHx was undetectable in HCCs which did not contain replicative intermediates. Subcellular localization studies detected WHx in the cytoplasm but not in the nuclei of primary woodchuck hepatocytes. Comparative immunoprecipitation experiments revealed that there were 4 X 10(4) to 8 X 10(4) molecules of WHx per primary woodchuck hepatocyte. Four lines of WHx transgenic mice did not develop HCC spontaneously. However, when one line was treated with diethylnitrosamine, the occurrence of precancerous lesions was enhanced compared with that in diethylnitrosamine-treated nontransgenic controls. The apparent absence of WHx in some woodchuck HCCs indicates that WHx may not be required to maintain the tumor phenotype, whereas its presence in all persistently infected livers leaves open the possibility that it plays a role in hepatocarcinogenesis.

Animals↗
PubMed1997

Neuraminidase hemadsorption activity, conserved in avian influenza A viruses, does not influence viral replication in ducks.

The N1 and N9 neuraminidase (NA) subtypes of influenza A viruses exhibit significant hemadsorption activity that localizes to a site distinct from that of the enzymatic active site. To determine the conservation of hemadsorption activity among different NAs, we have examined most of the NA subtypes from avian, swine, equine, and human virus isolates. All subtypes of avian virus NAs examined and one equine virus N8 NA possessed high levels of hemadsorption activity. A swine virus N1 NA exhibited only weak hemadsorption activity, while in human virus N1 and N2 NAs, the activity was detected at a much lower level than in avian virus NAs. NAs which possessed hemadsorption activity for chicken erythrocytes (RBCs) were similarly able to adsorb human RBCs. However, none of the hemadsorption-positive NAs could bind equine, swine, or bovine RBCs, suggesting that RBCs from these species lack molecules, recognized by the NA hemadsorption site, present on human and chicken RBCs. Mutagenesis of the putative hemadsorption site of A/duck/Hong Kong/7/75 N2 NA abolished the high level of hemadsorption activity exhibited by the wild-type protein but also resulted in a 50% reduction of the NA enzymatic activity. A transfectant virus, generated by reverse genetics, containing this mutated NA replicated 10-fold less efficiently in chicken embryo fibroblast cultures than did a transfectant virus expressing the wild-type NA. However, both viruses replicated equally well in Peking ducks. Although conservation of NA hemadsorption activity among avian virus NAs suggests the maintenance of a required function of NA, loss of the activity does not preclude the replication of the virus in an avian host.

Animals↗
PubMed2002

Varicella-zoster virus open reading frame 2 encodes a membrane phosphoprotein that is dispensable for viral replication and for establishment of latency.

Varicella-zoster virus (VZV) encodes six genes that do not have homologs in herpes simplex virus. One of these genes, VZV open reading frame 2 (ORF2), was expressed as a 31-kDa phosphoprotein in the membranes of infected cells. Unlike equine and bovine herpesvirus type 1 ORF2 homologs that are associated with virions, VZV virions contained no detectable ORF2 protein. The ORF2 deletion mutant established a latent infection in cotton rats at a frequency and with a number of VZV genomes similar to that of the parental virus. ORF63 transcripts, a hallmark of latent infection, were present in ganglia latently infected with both the ORF2 deletion mutant and parental VZV. Thus, ORF2 is the first VZV gene shown to be dispensable for establishment of latent infection in an animal model.

Animals↗
PubMed2005

Activation of the N-Ras-PI3K-Akt-mTOR pathway by hepatitis C virus: control of cell survival and viral replication.

The hepatitis C virus (HCV) replication complex is localized within detergent-resistant membranes or lipid rafts. We analyzed the protein contents of detergent-resistant fractions isolated from Huh7 cells expressing a self-replicating full-length HCV-1b genome. Using two-dimensional gel electrophoresis followed by mass spectrometry, we identified N-Ras as one of the proteins in which expression was increased in the detergent-resistant fractions from HCV genomic replicon clones compared to control cells. N-Ras is an activator of the phosphatidylinositol-3-kinase (PI3K)-Akt pathway. We found that the activities of PI3K and Akt, as well as the activity of their downstream target, mTOR, in the HCV-replicating cells were increased. Both PI3K-Akt- and mTOR-dependent pathways have been shown to promote cell survival. In agreement with this, HCV replicon cells were resistant to serum starvation-induced apoptosis. We also characterized the role of this pathway in HCV replication. Reduction of N-Ras expression by transfection of N-Ras small interfering RNA (siRNA) resulted in increased replication of HCV. We observed a similar increase in HCV replication in cells treated with the PI3K inhibitor LY294002 and in cells transfected with mTOR siRNA. Taken together, these data suggest that increased N-Ras levels in subcellular sites of HCV replication and stimulation of the prosurvival PI3K-Akt pathway and mTOR by HCV not only protect cells against apoptosis but also contribute to the maintenance of steady-state levels of HCV replication. These effects may contribute to the establishment of persistent infection by HCV.

Apoptosis↗
PubMed2005

The immediate-early protein IE0 of the Autographa californica nucleopolyhedrovirus is not essential for viral replication.

The role of the Autographa californica multiple nucleopolyhedrovirus (AcMNPV) immediate-early protein IE0 in the baculoviral infection is not clear. In this study, we constructed the recombinant virus vAcDeltaie0 null for ie0 expression by targeted mutagenesis replacing exon0 with the cat gene. We found that vAcDeltaie0 replicated efficiently in Spodoptera littoralis SL2 cells, which are poorly permissive for AcMNPV. In contrast, in Spodoptera frugiperda SF9 cells, which are fully permissive for AcMNPV, vAcDeltaie0 DNA replication and budded virus production were delayed. These results and recently published data (X. Dai et al., J. Virol. 78:9633-9644, 2004) indicate that ie0 is not essential for AcMNPV replication but enhances it in permissive SF9 cells.

Animals↗
PubMed2005

Sialidase activity of influenza A virus in an endocytic pathway enhances viral replication.

N2 neuraminidase (NA) genes of the 1957 and 1968 pandemic influenza virus strains possessed avian-like low-pH stability of sialidase activity, unlike most epidemic strains. We generated four reverse-genetics viruses from a genetic background of A/WSN/33 (H1N1) that included parental N2 NAs of 1968 pandemic (H3N2) and epidemic (H2N2) strains or their counterpart N2 NAs in which the low-pH stability of the sialidase activity was changed by substitutions of one or two amino acid residues. We found that the transfectant viruses bearing low-pH-stable sialidase (WSN/Stable-NAs) showed 25- to 80-times-greater ability to replicate in Madin-Darby canine kidney (MDCK) cells than did the transfectant viruses bearing low-pH-unstable sialidase (WSN/Unstable-NAs). Enzymatic activities of WSN/Stable-NAs were detected in endosomes of MDCK cells after 90 min of virus internalization by in situ fluorescent detection with 5-bromo-4-chloro-indole-3-yl-alpha-N-acetylneuraminic acid and Fast Red Violet LB. Inhibition of sialidase activity of WSN/Stable-NAs on the endocytic pathway by pretreatment with 4-guanidino-2,4-dideoxy-N-acetylneuraminic acid (zanamivir) resulted in a significant decrease in progeny viruses. In contrast, the enzymatic activities of WSN/Unstable-NAs, the replication of which had no effect on pretreatment with zanamivir, were undetectable in cells under the same conditions. Hemadsorption assays of transfectant-virus-infected cells revealed that the low-pH stability of the sialidase had no effect on the process of removal of sialic acid from hemagglutinin in the Golgi regions. Moreover, high titers of viruses were recovered from the lungs of mice infected with WSN/Stable-NAs on day 3 after intranasal inoculation, but WSN/Unstable-NAs were cleared from the lungs of the mice. These results indicate that sialidase activity in late endosome/lysosome traffic enhances influenza A virus replication.

Animals↗
PubMed2006

Glioma virotherapy: effects of innate immune suppression and increased viral replication capacity.

Oncolytic viruses are genetically altered replication-competent viruses that infect, and reproduce in, cancer cells but do not harm normal cells. On lysis of the infected cells, the newly formed viruses burst out and infect other tumor cells. Experiments with injecting mutant herpes simplex virus 1 (hrR3) into glioma implanted in brains of rats show lack of efficacy in eradicating the cancer. This failure is attributed to interference by the immune system. Initial pretreatment with immunosuppressive agent cyclophosphamide reduces the percentage of immune cells. We introduce a mathematical model and use it to determine how different protocols of cyclophosphamide treatment and how increased burst size of the mutated virus will affect the growth of the cancer. One of our conclusions is that the diameter of the cancer will decrease from 4 mm to eventually 1 mm if the burst size of the virus is triple that which is currently available. The effect of repeated cyclophosphamide treatment is to maintain a low density of uninfected cells in the tumor, thus reducing the probability of migration of tumor cells to other locations in the brain.

Animals↗
PubMed1987

Discontinuation of immunosuppressive therapy in hepatitis B surface antigen-positive chronic hepatitis: effect on viral replication and on liver cell damage.

Immunosuppressive therapy was stopped in 12 individuals positive for hepatitis Be antigen (HBeAg) and hepatitis B surface antigen (HBsAg) and in 4 individuals positive for HBsAg but negative for HBeAg. Discontinuation of immunosuppressive therapy in HBeAg-positive patients was always associated with a bout of hepatitis and elimination of HBeAg in 8/12 patients. One patient died from liver failure and 2 patients experienced a decompensation of their liver disease indicating that this approach might be harmful if used therapeutically. A bout of hepatitis was not noted in any of the individuals negative for HBeAg when the immunosuppressive therapy was stopped, implying that this event is not potentially harmful to the patient.

Adult↗
PubMed1994

HIV-1 macrophage tropism is determined at multiple levels of the viral replication cycle.

The ability of HIV-1 to infect macrophages is thought to be essential in AIDS pathogenesis. We tested the ability of 19 primary virus isolates to infect monocyte-derived macrophages (MDM) from different donors. Two HIV-1 isolates were able to establish a productive infection in MDM from all donors tested, whereas eight completely lacked this capacity. Next to these isolates with extreme phenotypes, 50% of the primary isolates under study displayed an intermediate phenotype. These intermediate macrophage-tropic isolates established a productive infection in MDM from some but not all donors tested. PCR analysis demonstrated that the capacity to replicate in MDM could be determined at the previously described level of virus entry. However, for intermediate macrophage-tropic isolates replication was abrogated at the level of reverse transcription. Entry of highly macrophage-tropic isolates resulted in efficient completion of the reverse transcription process, whereas entry of intermediate macrophage-tropic isolates did not. Our experiments indicate that primary HIV-1 isolates may differ in their dependency on cellular factors required for reverse transcription in MDM. Differences in susceptibility of MDM for in vitro HIV-1 infection suggest variation in the availability of these cellular factors between MDM from different individuals.

Adult↗
PubMed1992

[RC-IAL: rabbit kidney continuous cell line--characteristics and substrate for viral replication].

A rabbit kidney cell line RC-IAL, isolated in 1976 and at present at 150a passage, has had its characteristics analysed. The cells presented morphology similar to fibroblasts throughout their culture. The cellular growth proportion remained unaltered from its isolation, with a cloning efficiency of around 9%. The line showed growth dependent on anchorage and chromosomic analysis presented the modal number of the species with small variations to about one chromosome, to a total of about 50%. The line's species of origin was confirmed through indirect immunofluorescence reaction and susceptibility to some viruses with cytopathic effect was verified with vaccinia, cowpox, herpes simplex types 1 and 2 and rubella viruses. This cellular substract is free from contaminating agents, thus satisfying the conditions for its use in scientific work, especially that relating to public health.

Animals↗