Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.7.7 (DNA polymerase)
 17,007 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In order to investigate the potential susceptibility of bone marrow stromal myoid cells to human immunodeficiency virus (HIV), a myoid cell population devoid of all other cellular components of marrow environment was isolated from 3 normal adult bone marrow samples. The bone marrow myoid cells were infected with 3 different strains of HIV-1, 2 strains isolated from patients with acquired immune deficiency syndrome (AIDS) and HTLV-IIIB strain. To demonstrate successful infection and HIV production, culture supernatants, harvested weekly until 2 months post-infection, were tested for the presence of p24 antigen and infectious virus. Myoid cell monolayers obtained from the 3 different bone marrow samples were shown to be susceptible to infection. In particular, infection led to the presence of p24 antigen and of infectious virus in culture supernatants up to day 49 post-infection. After day 49, it was not possible to demonstrate the presence of infectious virus in culture supernatants and HIV-DNA polymerase chain reaction (PCR) failed to show viral genome in any of the cultures assayed. Our results demonstrate the susceptibility of myoid stromal cells to HIV infection and may provide an in vitro model for studying the effects of HIV infection in disregulation of the haemopoietic function of bone marrow environment.
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PMID:Human immunodeficiency virus infection of human bone marrow stromal myoid cells. 889 94

Asymptomatic infection of macaques with macaques with simian retroviruses type D (SRV/D), the etiologic agents of one form of retrovirus-induced simian immunodeficiency disease, can confound experiments with the simian immunodeficiency virus (SIV), which also induces immunodeficiency disease in macaques. The SIV/macaque model is the preferred nonhuman primate model for AIDS-related research. Serological screening for SRV/D alone is insufficient because not all infected animals seroconvert, and virus isolation by cocultivation may require 4 to 6 weeks. We have established a DNA polymerase chain reaction (PCR) assay. One set of nested primers allows detection of SRV/D serotypes 1, 2, and 3 and distinguishes SRV-2 from the other two serotypes. The PCR assay is sensitive; a single proviral copy of SRV/D could be detected in 150,000 to 210,000 macaque peripheral blood mononuclear cells (PBMCs). When applied to a panel of virus isolation-positive macaque samples, the PCR assay was positive in 100% of the tests. No false-positive results were seen when known specific-pathogen-free (SPF) macaques were examined. We propose that macaques be screened with a combination of SRV/D serology and this DNA PCR assay prior to enrollment in experiments with SIV.
AIDS Res Hum Retroviruses 1997 Mar 20
PMID:Simultaneous detection of simian retrovirus type D serotypes 1, 2, and 3 by polymerase chain reaction. 907 85

Simian retroperitoneal fibromatosis (RF) is a vascular fibroproliferative neoplasm which has many morphological and histological similarities to human Kaposi's sarcoma (KS). Like epidemic KS in AIDS patients, RF is highly associated with an immunodeficiency syndrome (simian acquired immunodeficiency syndrome [SAIDS]) caused by a retrovirus infection. Recently, a new gammaherpesvirus, called Kaposi's sarcoma-associated herpesvirus (KSHV) or human herpesvirus 8 (HHV8), has been identified in KS tumors, suggesting that KS has a viral etiology. Our previous experimental transmission studies and epidemiological data suggest that RF also has an infectious etiology. In order to determine whether a similar virus is also associated with RF, we have assayed for the presence of an unknown herpesvirus using degenerate PCR primers targeting the highly conserved DNA polymerase genes of the herpesvirus family. Here we provide DNA sequence evidence for two new herpesviruses closely related to KSHV from RF tissues of two macaque species, Macaca nemestrina and Macaca mulatta. Our data suggest that KSHV and the putative macaque herpesviruses define a new group within the subfamily Gammaherpesvirinae whose members are implicated in the pathogenesis of KS and KS-like neoplasms in different primate species.
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PMID:Identification of two homologs of the Kaposi's sarcoma-associated herpesvirus (human herpesvirus 8) in retroperitoneal fibromatosis of different macaque species. 909 97

Zidovudine (AZT), didanosine (ddI) and zalcitabine (ddC) are the reference antiretroviral therapy in patients with AIDS. A toxic mitochondrial myopathy can be observed in patients treated with AZT, but not with ddI and ddC. All 3 compounds can inhibit mitochondrial (mt)DNA polymerase and cause termination of synthesis of growing mtDNA strands and mtDNA depletion. The propensity to injure particular target tissues is unexplained. In our work, cultured muscle cells prepared from human muscle biopsies, were exposed to various concentrations of AZT (4-5000 micromol/l), ddI (5-1000 micromol/l) and ddC (1-1000 micromol/l) for 10 days. We evaluated cell proliferation and differentiation and measured lipid droplet accumulation, lactate production and respiratory chain enzyme activities. All 3 compounds induced a dose-related decrease of cell proliferation and differentiation. AZT seemed to be the most potent inhibitor of cell proliferation. AZT, ddI and ddC induced cytoplasmic lipid droplet accumulations, increased lactate production and decreased activities of COX (complex IV) and SDH (part of complex II). NADHR (complex I) and citrate sinthase activities were unchanged. Zalcitabine (ddC) and, to a lesser extent, ddI, were the most potent inhibitors of mitochondrial function. In conclusion, AZT, ddI and ddC all exert cytotoxic effects on human muscle cells and induce functional alterations of mitochondria possibly due to mechanisms other than the sole mtDNA depletion. Our results provide only a partial explanation of the fact that AZT, but not ddI and ddC, can induce a myopathy in HIV-infected patients. AZT myopathy might not simply result from a direct mitochondrial toxic effect of crude AZT.
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PMID:Cellular and mitochondrial toxicity of zidovudine (AZT), didanosine (ddI) and zalcitabine (ddC) on cultured human muscle cells. 916 61

Conversion of the single-stranded RNA of an invading retrovirus into double-stranded proviral DNA is catalyzed in a multi-step process by a single virus-coded enzyme, reverse transcriptase (RT). Achieving this requires a combination of DNA polymerase abd ribonuclease H (RNase H) activities, which are located at the amino and carboxy terminus of the enzyme, respectively. Moreover, proviral DNA synthesis requires that three structurally-distinct nucleic acid duplexes are accommodated by this enzyme, namely (a) A-form RNA (initiation of minus strand synthesis), non-A, non-B RNA/DNA hybrid (minus strand synthesis and initiation of plus strand synthesis) and B-form duplex DNA (plus strand synthesis). This review summarizes our current understanding of the manner in which retroviral RT interacts with this diverse array of nucleic acid duplexes, exploiting in many cases mutants unable to catalyze a specific event. These studies illustrate that seemingly 'simple' events such as tRNA-primed initiation of minus strand synthesis are considerably more complex, involving intermolecular tRNA-viral RNA interactions outside the primer binding site. Moreover, RNase H activity, generally thought to catalyze non-specific degradation of the RNA-DNA replicative intermediate, is required for highly specialized events including DNA strand transfer and polypurine selection. Finally, a unique structure near the center of HIV proviral DNA, the central termination sequence, serves to halt the replication machinery in a manner analogous to termination of transcription. As these highly specialized events are better understood at the molecular level, they may open new avenues of therapeutic intervention in the continuing effort to stem the progression of HIV infection and AIDS.
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PMID:Interaction of retroviral reverse transcriptase with template-primer duplexes during replication. 930 71

Epidemiological studies strongly suggest that a newly discovered herpesvirus, Kaposi's sarcoma associated-herpesvirus (KSHV/HHV8), is the likely infectious cause of Kaposi's sarcoma (KS). Identification of this agent suggests the possibility that existing anti-herpesvirus chemotherapeutics have activity against KSHV. Using KSHV/Epstein-Barr virus (EBV)-coinfected cell line BC-1 and KSHV-infected/EBV-negative cell line BC-3, the effect of DNA polymerase inhibitors in the presence of virus-inducing agents was examined. The phorbol ester TPA induced 8.2-fold EBV replication in BC-1 cells with only minimal concurrent KSHV genome replication in BC-1, but induced fourfold KSHV in BC-3 cells. TPA, however, induced transcripts encoded by the lytic cycle major capsid protein gene that were inhibited by both phosphonoacetic acid and phosphonoformic acid either in the KSHV/EBV-infected cell line or in the EBV-negative/KSHV-infected cell line. Transcripts hybridizing to a KSHV-encoded cyclin gene were unaffected by either TPA or DNA polymerase inhibitors in both cell lines. These results show in vitro activity of DNA polymerase inhibitors on KSHV lytic transcript accumulation and may provide a simple assay for evaluating the efficacy of potential anti-KSHV chemotherapeutics.
AIDS Res Hum Retroviruses 1997 Sep 20
PMID:Effect of DNA synthesis inhibitors on Kaposi's sarcoma-associated herpesvirus cyclin and major capsid protein gene expression. 931 Feb 90

We have constructed a plasmid that induces in bacteria the synthesis of an enzymically active reverse transcriptase (RT) of mouse mammary tumour virus (MMTV), a retrovirus with a typical B-type morphology. The highest catalytic activity was detected only when 27 residues from the C-terminus of the protease were included in the N-terminus of the recombinant RT, after an extra deoxyadenosine was added between the pro and pol genes to overcome the -1 frameshift event (which occurs naturally in virus-infected cells). The recombinant protein with a six-histidine tag was purified to homogeneity by a two-column purification procedure, Ni2+ nitriloacetic acid/agarose followed by carboxymethyl-Sepharose chromatography. Unlike most RTs, the purified MMTV RT is enzymically active as a monomer even after binding a DNA substrate. Like all RTs studied, the recombinant MMTV RT possesses RNA-dependent and DNA-dependent DNA polymerase activities as well as RNase H activity, all of which show a preference for Mg2+ over Mn2+ ions. Other features of these enzymic activities, such as extension of DNA primers, processivity of DNA synthesis, pH dependence, steady-state kinetic constants, effects of Na+ or K+ ions and sensitivity to a thiol-specific reagent and to a zinc chelator, have been evaluated. The catalytic properties of MMTV RT were compared with those of the well-studied RT of HIV-1, the causative agent of AIDS. Interestingly, MMTV RT exhibits a high sensitivity to nucleoside triphosphate analogues (which are known to be potent inhibitors of HIV RTs and are being used as the major anti-AIDS drugs), as high as that of HIV-1 and HIV-2 RTs. Furthermore the recombinant MMTV RT shows a processivity of DNA synthesis higher than that of HIV-1 RT.
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PMID:Reverse transcriptase of mouse mammary tumour virus: expression in bacteria, purification and biochemical characterization. 944 85

Gene therapy is a promising treatment modality for acquired immunodeficiency syndrome (AIDS). Autologous transplantation with genetically altered pluripotent hematopoietic stem cells encoding anti-human immunodeficiency virus (HIV) genes could in theory completely and permanently reconstitute all blood lineages and immune functions with cells resistant to HIV. Recent studies showed that CD34+ stem cell can be mobilized in HIV-infected individuals after granulocyte colony-stimulating factor (G-CSF) administration without major side effects or increase of viral load. In this study, peripheral blood CD34+ cells of five HIV-infected individuals were mobilized with G-CSF and after leukapheresis and enrichment, subjected to retroviral transduction with genes encoding anti-HIV ribozyme-decoy fusion molecules. These cells were tested for the ability to give rise to progeny cells, for retroviral transduction efficiency, and for expression of the transgene. CD34+-derived macrophage-like cells were also challenged with HIV. Results showed that CD34+ cells from HIV-infected individuals gave rise to similar numbers of progeny colonies as cells from healthy donors. The transduction efficiency of these cells varied from 68.8 to 100% as assessed by DNA polymerase chain reaction (PCR) of the transgene in individual colonies. CD34+-derived macrophages expressed anti-HIV genes and displayed a substantial and sustained inhibition of HIV replication as compared to untransduced cells. Furthermore, we showed that after thawing, cryopreserved CD34+ cells from these individuals have survival, proliferation, and transduction parameters comparable to fresh cells. Thus, CD34+ cells from HIV-infected patients can be stored for further genetic manipulations with improved vectors or anti-HIV genes as they become available.
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PMID:Gene therapy targeting peripheral blood CD34+ hematopoietic stem cells of HIV-infected individuals. 944 70

Foscarnet, Phosphonoformate, has recently approved for the treatment of HCMV retinitis in AIDS patients in Japan. It inhibits the viral DNA polymerase and effective against ganciclovir-resistant HCMV. Cidofovir, (S)-1-[3-hydroxy-2-(phosphonylmethoxy)propyl]cytosine, a new acyclic nucleotide phosphonate analogues has potent activity against HCMV retinitis in AIDS at 3-7.5 mg/kg/week, once/week with concomitant oral probenecid and saline prehydration to prevent nephrotoxicity. The highest potency and selectivity against HHV-6 and HHV-7 was demonstrated S2242 (N7-isomer of 6-deoxy-ganciclovir). Ganciclovir, foscarnet, and cidofovir also exhibited selective inhibitory activity to these viruses, although the activities were not so remarkable compared with in case of HCMV. Thymidine kinase-dependent drugs (acyclovir, brivudin) showed little, if any, activity. These results suggest a structural homology of the DNA polymerase and a lack of TK gene among these three beta-herpesviruses.
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PMID:[Recent advances in antiviral drugs--antiviral agents to HCMV, HHV-6, and HHV-7]. 946 79

Two ganciclovir (GCV)-resistant human cytomegalovirus (HCMV) strains recovered from an AIDS patient (strain VR4990) and a heart transplant recipient (strain VR5474) showed a Cys607-->Tyr change in the UL97-encoded phosphotransferase. No amino acid substitutions were observed in the viral DNA polymerase. Marker transfer experiments showed marked reduction in GCV phosphorylation and drug susceptibility of the recombinant HCMV strain VR4990rec2-1-1. These results further extend the region of the carboxy-terminal domain of the UL97 phosphotransferase involved in GCV substrate recognition.
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PMID:The Cys607-->Tyr change in the UL97 phosphotransferase confers ganciclovir resistance to two human cytomegalovirus strains recovered from two immunocompromised patients. 952 4


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