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At least 325 records · Page 18Linked to original sources
PubMed1995

The tempo and mode of SIV quasispecies development in vivo calls for massive viral replication and clearance.

Simian immunodeficiency virus (SIV) quasispecies development was followed in four monkeys (Macacca fascicularis) infected by intramuscular inoculation of phage lambda-SIVmac239 DNA. Rooted phylogenetic trees were reconstructed and used to interpret the data. The rate of fixation of base substitutions varied within and between animals reaching 3.3 x 10(-2) per site per year. These data suggest that the tempo of quasispecies development requires both massive viral replication and efficient clearance of SIV. Despite this, no significant difference was found between the observed and expected ratio of synonymous/nonsynonymous substitutions, suggesting that there was little or no selection of antigenic variants in the V1 and V2 hypervariable regions of envelope.

Amino Acid Sequence↗
PubMed2003

The dimer initiation sequence stem-loop of human immunodeficiency virus type 1 is dispensable for viral replication in peripheral blood mononuclear cells.

Human immunodeficiency virus type 1 (HIV-1) contains two copies of genomic RNA that are noncovalently linked via a palindrome sequence within the dimer initiation site (DIS) stem-loop. In contrast to the current paradigm that the DIS stem or stem-loop is critical for HIV-1 infectivity, which arose from studies using T-cell lines, we demonstrate here that HIV-1 mutants with deletions in the DIS stem-loop are replication competent in peripheral blood mononuclear cells (PBMCs). The DIS mutants contained either the wild-type (5'GCGCGC3') or an arbitrary (5'ACGCGT3') palindrome sequence in place of the 39-nucleotide DIS stem-loop (NL(CGCGCG) and NL(ACGCGT)). These DIS mutants were replication defective in SupT1 cells, concurring with the current model in which DIS mutants are replication defective in T-cell lines. All of the HIV-1 DIS mutants were replication competent in PBMCs over a 40-day infection period and had retained their respective DIS mutations at 40 days postinfection. Although the stability of the virion RNA dimer was not affected by our DIS mutations, the RNA dimers exhibited a diffuse migration profile when compared to the wild type. No defect in protein processing of the Gag and GagProPol precursor proteins was found in the DIS mutants. Our data provide direct evidence that the DIS stem-loop is dispensable for viral replication in PBMCs and that the requirement of the DIS stem-loop in HIV-1 replication is cell type dependent.

Base Sequence↗
PubMed1996

Nitric oxide inhibits viral replication in murine myocarditis.

Nitric oxide (NO) is a radical molecule that not only serves as a vasodilator and neurotransmitter but also acts as a cytotoxic effector molecule of the immune system. The inducible enzyme making NO, inducible NO synthase (iNOS), is transcriptionally activated by IFN-gamma and TNF-alpha, cytokines which are produced during viral infection. We show that iNOS is induced in mice infected with the Coxsackie B3 virus. Macrophages expressing iNOS are identified in the hearts and spleens of infected animals with an antibody raised against iNOS. Infected mice have increased titers of virus and a higher mortality when fed NOS inhibitors. Thus, viral infection induces iNOS in vivo, and NO inhibits viral replication. NO is a novel, nonspecific immune defense against viruses in vivo.

Amino Acid Sequence↗
PubMed2004

Enhanced surfactant protein and defensin mRNA levels and reduced viral replication during parainfluenza virus type 3 pneumonia in neonatal lambs.

Defensins and surfactant protein A (SP-A) and SP-D are antimicrobial components of the pulmonary innate immune system. The purpose of this study was to determine the extent to which parainfluenza type 3 virus infection in neonatal lambs alters expression of sheep beta-defensin 1 (SBD-1), SP-A, and SP-D, all of which are constitutively transcribed by respiratory epithelia. Parainfluenza type 3 viral antigen was detected by immunohistochemistry (IHC) in the bronchioles of all infected lambs 3 days postinoculation and at diminished levels 6 days postinoculation, but it was absent 17 days postinoculation. At all times postinoculation, lung homogenates from parainfluenza type 3 virus-inoculated animals had increased SBD-1, SP-A, and SP-D mRNA levels as detected by fluorogenic real-time reverse transcriptase PCR. Protein levels of SP-A in lung homogenates detected by quantitative-competitive enzyme-linked immunosorbent assay and protein antigen of SP-A detected by IHC were not altered. These studies demonstrate that parainfluenza type 3 virus infection results in enhanced expression of constitutively transcribed innate immune factors expressed by respiratory epithelia and that this increased expression occurs concurrently with decreased viral replication.

Animals↗
PubMed1997

A lipidated anti-Tat antibody enters living cells and blocks HIV-1 viral replication.

We have developed a chemical modification of antibodies, lipidation, which enables their intracellular delivery into living cells. Intracellular localization of lipidated antibodies was demonstrated by confocal microscopy and by measuring cellular uptake of 125I-labeled lipidated antibodies. Functionally, a lipidated monoclonal antibody directed against the Tat protein from human immunodeficiency virus type 1 (HIV-1) inhibited viral replication of several HIV-1 isolates by approximately 85% as shown by increased viability of infected cells and decreased reverse transcriptase activity. The antibody in its native form had no such effect. These data show that lipidated antibodies can reach and functionally inhibit intracellular targets. Lipidation may help to facilitate the development of intracellular immunotherapy for AIDS.

Antibodies, Monoclonal↗
PubMed2005

Inhibition of viral replication by ribozyme: mutational analysis of the site and mechanism of antiviral activity.

A controlled mutational study was used to determine the site and mechanism of the antiviral action of ribozymes that inhibit Sindbis virus replication. A hairpin ribozyme targeting G575 of the Sindbis virus genomic RNA was designed and cloned into a minimized alphavirus amplicon vector. Cells that were stably transfected with this construct expressed low levels of a constitutive transcript containing the ribozyme plus recognition sequences for Sindbis RNA replicase. Upon infection, the ribozyme transcript was amplified to high levels by the viral replicase, resulting in decreased viral production from infected ribozyme-expressing cells. Mutations were then introduced into the viral RNA target sequence to interfere with ribozyme binding, and compensatory changes were generated in the ribozyme recognition sequence. Single mutations in the virus or ribozyme decreased the efficacy of the ribozyme's inhibition of viral replication, and compensatory mutations restored it. To confirm that ribozyme-catalyzed RNA cleavage was actually needed for inhibition, we performed tests with a cell line expressing an inactivated ribozyme and with a virus containing a single nucleotide target mutation that allowed the ribozyme to bind but blocked cleavage at the recognition site. The results show that most of the antiviral activity of ribozymes is due to ribozyme-catalyzed cleavage at the targeted RNA sequence, but some additional inhibition seems to occur through an antisense mechanism.

Animals↗
PubMed1987

Novobiocin and coumermycin A1 inhibit viral replication and the reactivation of herpes simplex virus type 1 from the trigeminal ganglia of latently infected mice.

Herpes simplex virus type 1 was reactivated from the trigeminal ganglia of latently infected mice in a quantitative and time-dependent manner. Novobiocin and coumermycin A1 reversibly inhibited the reactivation of herpes simplex virus type 1. They did not inhibit viral replication in permissive cells (CV-1) but did inhibit replication in cells of neuronal origin (C1300) and acutely infected trigeminal ganglia.

Aminocoumarins↗
PubMed2000

Variation in the nucleotide sequence of cottontail rabbit papillomavirus a and b subtypes affects wart regression and malignant transformation and level of viral replication in domestic rabbits.

We previously reported the partial characterization of two cottontail rabbit papillomavirus (CRPV) subtypes with strikingly divergent E6 and E7 oncoproteins. We report now the complete nucleotide sequences of these subtypes, referred to as CRPVa4 (7,868 nucleotides) and CRPVb (7,867 nucleotides). The CRPVa4 and CRPVb genomes differed at 238 (3%) nucleotide positions, whereas CRPVa4 and the prototype CRPV differed by only 5 nucleotides. The most variable region (7% nucleotide divergence) included the long regulatory region (LRR) and the E6 and E7 genes. A mutation in the stop codon resulted in an 8-amino-acid-longer CRPVb E4 protein, and a nucleotide deletion reduced the coding capacity of the E5 gene from 101 to 25 amino acids. In domestic rabbits homozygous for a specific haplotype of the DRA and DQA genes of the major histocompatibility complex, warts induced by CRPVb DNA or a chimeric genome containing the CRPVb LRR/E6/E7 region showed an early regression, whereas warts induced by CRPVa4 or a chimeric genome containing the CRPVa4 LRR/E6/E7 region persisted and evolved into carcinomas. In contrast, most CRPVa, CRPVb, and chimeric CRPV DNA-induced warts showed no early regression in rabbits homozygous for another DRA-DQA haplotype. Little, if any, viral replication is usually observed in domestic rabbit warts. When warts induced by CRPVa and CRPVb virions and DNA were compared, the number of cells positive for viral DNA or capsid antigens was found to be greater by 1 order of magnitude for specimens induced by CRPVb. Thus, both sequence variation in the LRR/E6/E7 region and the genetic constitution of the host influence the expression of the oncogenic potential of CRPV. Furthermore, intratype variation may overcome to some extent the host restriction of CRPV replication in domestic rabbits.

Animals↗
PubMed1985

Clinical and serological events accompanying changes in hepatitis B viral replication: case reports.

We measured serum markers of hepatitis B virus replication in two HBsAg-, HBeAg-positive hepatitis B carriers with chronic active hepatitis and cirrhosis. The first of these patients was HBsAg-, HBeAg-, HBV DNA- and HBV DNA polymerase-positive initially and spontaneously lost HBV DNA polymerase and HBV DNA. During the HBeAg-positive, DNA polymerase-negative "window phase", an increase in viral replication, characterized by the reappearance of HBV DNA and HBV DNA polymerase occurred, together with an aggravation of the underlying chronic hepatitis. In the second HBsAg-, HBeAg-positive carrier, spontaneous fluctuations in HBV replication were associated with clinical deterioration. Delta agent and hepatitis A virus superinfection were excluded. These observations suggest that spontaneous low-grade fluctuations of HBV replication accompanied by an increase in the biochemical activity of the underlying chronic hepatitis can be observed in certain HBV carriers.

Alanine Transaminase↗
PubMed1983

Acyclovir in hepatitis B antigen-positive chronic liver disease: inhibition of viral replication and transient renal impairment with iv bolus administration.

Six patients with hepatitis B virus (HBV) related chronic liver disease were treated with acyclovir, 5-15 mg/kg 8 hourly, given as an iv bolus or iv infusion over 1 h for up to 7 days. Two patients treated with 10 and 15 mg/kg 8 hourly showed a decrease in HBV-DNA polymerase and HBV-DNA when mean trough acyclovir plasma concentrations of 5.0 +/- 0.6 and 13.2 +/- 3.0 microM were attained. Inhibition of viral replication was not seen in patients treated with lower doses. Transient renal impairment was seen in two patients who received high dosage by the iv bolus mode of administration. This complication may be prevented by a high oral fluid intake or iv infusion of the drug over 1 h. Further study with acyclovir 15 mg/kg 8-hourly given as an iv infusion for longer periods is warranted.

Acyclovir↗
PubMed1991

Dispensable role of the human immunodeficiency virus type 2 Vpx protein in viral replication.

Human immunodeficiency virus type 2 (HIV-2) is similar in genetic organization to HIV-1 but contains a unique gene (vpx) that encodes a 16-kDa protein. A replication-competent molecular clone of HIV-2 (HIV-2sbl/isy) that infects human primary cells in vitro and rhesus monkeys was used to generate three mutations in the vpx gene. In the first mutant, the vpx open reading frame was truncated at amino acid 20; the second mutant was tailored to eliminate the proline-rich carboxyl terminus of the protein; and the third mutant was obtained by addition of four amino acids (KDEL) to the carboxyl terminus of the protein to provide a retention signal in the endoplasmic reticulum. The viral infection kinetics of the three mutant viruses and isogeneic HIV-2sbl/isy in the SupT1 cell line were similar. Slight impairment in the early phases of viral replication was observed during infection of primary human peripheral blood mononuclear cells with the vpx mutant viruses. All of the vpx mutant viruses readily infected macrophages, indicating that vpx expression is dispensable for HIV-2 infection and replication in human macrophages.

Amino Acid Sequence↗
PubMedDate not supplied

Interferon-regulated viral replication in chronically HIV1-infected promonocytic U937 cells.

To further document the role of interferons in the restriction of HIV1 replication in cells of the monocyte/macrophage lineage, the antiviral effects of alpha-interferon (IFN alpha) were studied in chronically HIV 1-infected promonocytic cells U937, in which IFN alpha was endogenously produced or to which recombinant IFN alpha was added. Protein analysis performed after immunoprecipitation of culture media revealed that the addition of anti-IFN alpha antibody led to an increase in the production of viral particles, whereas addition of IFN alpha caused its decrease in a dose-dependent manner, indicating that IFN alpha mainly affects viral assembly and virion release. However, the treatment of HIV 1-infected cells with IFN alpha did not cause an increase in the amount of intracellular viral proteins, suggesting that this cytokine might act on other steps in the viral life cycle. No decrease in the level of the 3 main types of viral RNA was observed by Northern blot analysis, indicating that proviral transcription was not restricted by IFN alpha. Furthermore, cotransfection experiments with TAR(trans-activation responsive)-element-containing expression plasmids demonstrated that viral replication appears to be restricted at either a post-transcriptional and/or a translational level. These experiments suggest that the double-stranded (ds) inverted repeat TAR sequence present in HIV1 leader RNA may inhibit viral protein synthesis by phosphorylating the dsRNA-activated protein kinase induced by IFN. These results provide an impetus for achieving antiretroviral therapy based on the constitutive expression of IFN in monocytes and macrophages.

Cell Line↗
PubMed1990

[Correlation of viral replication activity, the clinical phase of the disease and immunologic deterioration in HIV infection].

The authors have studied the relationship among HIV replicative activity, expressed by HIV isolation from peripheral blood lymphocytes and p24 antigenemia detection, and the evolution of infection, in terms of stage of disease and T4 cells absolute number, in a group of 120 infected subjects. Data presented indicate that the expression of viral gene products is related both to clinical and immunological deterioration and suggest that viral replication plays a central role in the progression of the disease.

AIDS-Related Complex↗
PubMed2000

The nuclear receptor chicken ovalbumin upstream promoter transcription factor interacts with HIV-1 Tat and stimulates viral replication in human microglial cells.

Human immunodeficiency virus type 1 (HIV-1) infects the central nervous system and plays a direct role in the pathogenesis of AIDS dementia. However, the molecular mechanisms underlying HIV-1 expression in the central nervous system are poorly understood. We have recently reported that the nuclear receptor chicken ovalbumin upstream promoter transcription factor (COUP-TF), an orphan member of the nuclear receptor superfamily, is an activator of HIV-1 gene transcription. Here, our results show that COUP-TF stimulates HIV-1 transcription in primary cultured human microglial cells, the primary target for HIV-1 infection in brain. Run-on assays indicated that COUP-TF acts on the initiation step of transcription. Results from reverse transcription-polymerase chain reaction and immunocytochemistry analysis further revealed the importance of this factor by demonstrating that overexpression of COUP-TF leads to initiation of viral replication in primary HIV-infected human microglia. In addition, COUP-TF is able to physically interact and cooperate with the viral transactivator Tat. The combination of COUP-TF and Tat leads to NF-kappaB- and Sp1-independent enhanced transcriptional stimulation. In vitro binding studies showed that COUP-TF interacts with Tat through amino acids within the N-terminal DNA-binding domain of COUP-TF. Amino acids 48-72 in the basic and C-terminal regions of Tat are required for the binding of Tat to COUP-TF. These results suggest that COUP-TF is an essential transcription factor involved in HIV-1 expression in microglia and reveal a novel interplay of Tat and COUP-TF during regulation of viral expression.

Animals↗
PubMed1994

Mutagenesis of the yellow fever virus NS2A/2B cleavage site: effects on proteolytic processing, viral replication, and evidence for alternative processing of the NS2A protein.

The yellow fever virus NS2B-3 proteinase mediates cleavages within the nonstructural region at a consensus sequence defined by G/ARR decreases S/G and also at an alternative site within the NS4A region. To determine the importance of specific residues within the consensus sequence for cleavage at the 2A/2B site, amino acid substitutions were introduced at each of the P4, P3, P2, P1, and P1' positions and the effects on proteolytic processing of a sig2A-5(356) polyprotein were examined using a vaccinia virus-T7 transient expression system. At the P1 and P1' positions, only the conservative substitutions P1:R-->K and P1':S-->G allowed efficient cleavage, suggesting that basic and small aliphatic amino acids are preferred at the P1 and P1' positions, respectively. At the P2 position, a preference for a basic amino acid was observed. In contrast, the P3 and P4 positions tolerated nonconservative substitutions and at P4 both enhancement and reduction in cleavage efficiency was observed. Evidence for cleavage at an alternative site within NS2A, defined by the sequence QK decreases T (NS2A residues 189-191) was obtained. Cleavage at this site, designated at NS2A alpha, is dependent upon an active NS2B-3 proteinase. To examine the effects of reduced cleavage efficiency at the 2A/2B and NS2A alpha cleavage sites on viral replication, mutations at each or both of these sites were incorporated into a full-length YF-17D cDNA template. RNA transcripts containing mutations which abolish cleavage were noninfectious whereas virus was recovered from several clones with mutations allowing partial cleavage at 2A/2B. However, some of these mutants exhibited a small plaque phenotype as well as reductions in RNA-specific infectivity and virus yield.

Amino Acid Sequence↗
PubMed2025

Marek's disease virus-1 unique gene LORF1 is involved in viral replication and MDV-1/Md5-induced atrophy of the bursa of Fabricius.

Marek's disease virus (MDV), an alphaherpesvirus, causes severe immunosuppression and T cell lymphomas in chickens, known as Marek's disease (MD), an economically important poultry disease primarily controlled by vaccination. Importantly, it also serves as a comparative model for studying herpesvirus-induced tumor formation in humans. MDV encodes more than 100 genes, most of which have unknown functions. MDV LORF1 is unique to serotype I MDV (MDV-1), lacking homologs in other herpesviruses, and has not been explored yet. To this end, an infectious bacterial artificial chromosome (BAC) harboring the complete genome of the MDV-1 very virulent strain Md5 was generated, and the rescued rMd5 maintained biological properties similar to the parental virus both in vitro and in vivo. Subsequently, rMd5ΔLORF1, a recombinant Md5 virus deficient in pLORF1 expression, was generated by a frameshift mutation in the LORF1 gene. Chickens infected with rMd5ΔLORF1 exhibited a lower mortality rate and delayed bursal atrophy than those infected with the parental rMd5 and the revertant virus (rMd5-reLORF1). Consistently, viral loads of rMd5ΔLORF1 were obviously lower than those of rMd5 or rMd5-reLORF1 in the bursa, but not in the spleen. Importantly, we found that pLORF1 deficiency impairs viral replication in bursal B cells. Furthermore, we showed that pLORF1 associated with the cellular membrane, interacted with MDV structural proteins, and exhibited punctate colocalization with tegument or capsid proteins in the cytoplasm. Taken together, this study demonstrates for the first time that the MDV-1 unique gene LORF1 is involved in MDV-induced bursal atrophy but not in tumor formation.

Animals↗
PubMed1994

Human immunodeficiency virus type 1 integrase: effect on viral replication of mutations at highly conserved residues.

Sequence comparisons of the integrase (IN) proteins from different retroviruses have identified several highly conserved residues. We have introduced mutations at 16 of these sites into the integrase gene of human immunodeficiency virus type 1 and analyzed the phenotypes of the resulting viruses. The viruses were all normal for p24 content and reverse transcriptase activity. In addition, all of the mutants could infect T-cell lines and undergo reverse transcription, as assessed by PCR analysis. Most of the mutant viruses also had normal Western blot (immunoblot) profiles, although three of the mutations resulted in reduced signals for IN relative to the wild type on the immunoblots and mutation of residue W235 completely abolished recognition of the protein by pooled sera from human immunodeficiency virus type 1-positive patients. Mutations that have previously been shown to abolish activity in in vitro studies produced noninfectious viruses. The substitution of W235 was notable in producing a noninfectious virus, despite previous reports of this residue being nonessential for IN activity in vitro (A.D. Leavitt, L. Shiue, and H.E. Varmus, J. Biol. Chem. 268:2113-2119, 1993). In addition, we have identified four highly conserved residues that can be mutated without any affect on viral replication in T-cell lines.

Amino Acid Sequence↗