Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.7.7.49 (reverse transcriptase)
31,746 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Antiviral effects were characterized for two oligodeoxyribonucleoside methylphosphonates synthesized in an antisense (3'-TCTTAACC-5') or a sense (5'-AGAATTGG-3') orientation, based on the RNA sequence of the first splice acceptor site of the tat-3 gene of human immunodeficiency virus (HIV) (5'...AGAAUUGG...3'). The development of syncytial cells and supernatant reverse transcriptase was inhibited by a single exposure to the antisense HIV, and HIV RNA synthesis was inhibited by both antisense and sense methylphosphonates but not by a control herpes simplex virus antisense sequence.
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PMID:Inhibition of human immunodeficiency virus by using an oligonucleoside methylphosphonate targeted to the tat-3 gene. 245 90

In central equatorial Africa the frequency of uninterpretable or atypical Western blots (WB)--ie. antibodies to gag proteins only--can represent up to 50% of enzyme-linked immunosorbent assay (ELISA)-positive samples. To date the significance of such serology remains unknown. Nevertheless, an unusual HIV-1 strain has been isolated from the blood of a healthy Gabonese individual who presented an atypical WB. This virus, identified as isolated HIV-1OYl, grew to low titres of reverse transcriptase activity (less than 50,000 cpm/ml) and was not obviously cytopathic. Radioimmunoprecipitation and peptide ELISA studies indicated that the lack of env-specific reactivity was probably due to the absence of antibodies to the viral glycoproteins, rather than the virus encoding a highly divergent envelope protein. Molecular cloning and sequencing of the provirus proved it to be a string of HIV-1 which was genetically closer to European and North American than to African strains. Furthermore the envelope protein sequence contained all the features of a typical HIV-1 env gene. However, the tat gene derived from the proviral clone was functionally defective. Site-directed mutagenesis of this gene showed that this was due to the substitution of an essential cysteine residue for a serine. Polymerase chain reaction amplification of the tat gene, as well as parts of the gag and env gene sequences of HIV-1OYl, showed that essentially all of the proviruses were defective. These data emphasize the need to view HIV isolates as populations of distinct genomes capable of complementing each other.
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PMID:A highly defective HIV-1 strain isolated from a healthy Gabonese individual presenting an atypical western blot. 255 49

We have used the polymerase chain reaction (PCR) to detect by co-amplification, multiple regions of the HIV-1 genome in infected cells. Genomic RNA and DNA from productively infected H9 cells were independently extracted and amplified in reactions with and without reverse transcriptase respectively using primer pairs to the gag, env, tat and nef regions of the viral genome in the same reaction mixture. PCR-products were analysed by liquid hybridization with end labelled oligonucleotide probes followed by gel-electrophoresis (oligomer hybridization). The primer pairs were capable of detecting as few as 10 copies of RNA and 10-20 copies of integrated proviral DNA. The ability to co-amplify multiple target regions in the same incubation mixture provides a method for detecting and confirming the presence of HIV-1 in samples for which limited nucleic acid is available. In addition, in reconstitution experiments, the same method was used to detect HIV-1 and HTLV-I simultaneously with comparable sensitivity (20-40 gene copies each). This offers the possibility of simultaneous diagnosis of multiple viral infections, such as those that occur in AIDS, on the same sample preparation.
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PMID:Co-amplification of multiple regions of the HIV-1 genome by the polymerase chain reaction: potential use in multiple diagnosis. 257 Nov 15

The human immunodeficiency virus is a member of the lentivirus subfamily of the retrovirus family. Retroviruses are RNA viruses which code for an RNA-dependent DNA polymerase (reverse transcriptase), which transcribes the RNA genome into a DNA provirus which, on integration with the host DNA, directs the synthesis of new virions. The RNA genome consists of a gag gene, which codes for the viral core proteins, a pol gene, which codes for the reverse transcriptase, an env gene, which codes for the glycoproteins of the viral envelope, and several genes (tat, rev, vif, vpr, and nef), that code for regulatory proteins. At each end of the genome are long terminal repeats, that contain regulatory elements for transcription. There are 3 subfamilies of Retroviridae (Oncovirinae, Spumavirinae, and Lentiverinae). The Lentiverinae ("slow viruses") include the bovine immunodeficiency virus (BIV), the feline immunodeficiency virus (FIV), the human immunodeficiency viruses (HIV), and the simian immunodeficiency viruses (SIV). SIV has been isolated from macaques (mac), African green monkeys (agm), sooty mangabeys (sm), and mandrills (mnd). Only SIVmac causes an AIDS-like disease in its natural host, but it is genetically closer to HIV-2 than to HIV-1. SIVsm causes an AIDS-like disease in macaques, but not in the sooty mangabey. Monkeys infected with SIV develop diarrhea, wasting, decrease in T4 lymphocytes, lymphadenopathy, development of giant cells, and encephalitis, as well as opportunistic infections. Kaposi's sarcoma, however, has not been found in SIV-infected primates. Virus is recovered from peripheral blood mononuclear cells and the brain. SIV models are useful for understanding the natural history of primate lentiviruses, for defining the pathogenesis of AIDS, and for developing vaccines. The ideal model would be one in which HIV causes AIDS, but so far only chimpanzees and gibbons have successfully been infected with HIV-1, and although virus, is recovered from peripheral blood mononuclear cells of chimpanzees within 2 weeks of infection, and 2 animals have lost antibodies to the p24 protein, none has so far developed clinical AIDS. Attempts to develop vaccines to immunize chimpanzees are continuing. Nonprimate lentiviruses include the visna virus, the feline immunodeficiency virus, and the bovine immunodeficiency virus. The visna virus infects fibroblasts by fusion of the viral envelope with the plasma membrane of the fibroblast; it infects macrophages by endocytosis. Infected macrophages regulate the production and dissemination of viral particles. The feline immunodeficiency virus infects T-lymphocytes of cats and produces oral, gastrointestinal and respiratory pathology as well as lymphadenopathy and opportunistic infections. Bovine immunodeficiency-like virus causes a generalized lymphadenopathy similar to that seen in AIDS-related complex.
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PMID:Animal models for HIV infection and AIDS: memorandum from a WHO meeting. 285 Jan 18

The human spumaretrovirus (HSRV) isolated from a nasopharynx carcinoma patient 17 years ago has a RNA genome 11 kb in size. It encodes besides the gag, pol, and env genes several novel genes (S1 and bel 1, 2, and 3) that are comparable to the regulatory genes R, X, tat, art, and 3'-orf of the human (HIVs) and simian immunodeficiency viruses (SIVs) with respect to genomic location and to sizes of the putative gene products. A comparison between the HIV protein sequences of the pol and the novel genes to the corresponding gene product sequences of HSRV revealed that HSRV is related to the lentiviruses but occupies a distinct phylogenetic placement of its own. A detailed analysis of the reverse transcriptase domain allows the construction of a phylogenetic tree for the known retroviral subfamilies and/or groups, including the oncoviruses, the lentiviruses, the spumaviruses, the HLTV-BLV group, and the D-type viruses. Regions of the putative novel HSRV gene products with segmental protein sequence homology to the regulatory protein of other human retroviruses are discussed. The results strengthen the view that HSRV and its novel genes should be studied in comparison to the new genes of acquired immunodeficiency syndrome (AIDS) viruses and human T cell leukemia viruses (HTLV).
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PMID:Genomic organization of the human spumaretrovirus and its relatedness to AIDS and other retroviruses. 285 7

Recent research suggests that the collapse of the immune system that accompanies acquired immunodeficiency syndrome (AIDS) stems largely from a single defect: a reduction in the number and a change in the function of the T4 lymphocytes. This knowledge may make it possible to lessen the effects of AIDS, and eventually to prevent and cure it. The loss of T4 lymphocytes from the blood, lymph nodes, spleen, and other tissues in which they are normally concentrated is a consistent finding in AIDS patients. Without the help of T4 cells, B cells are unable to produce adequate quantities of antibody to the AIDS virus or to any other infection. Infected T4 cells are also the source of soluble suppressor factor, which blocks T-cell-dependent immune responses. The drug suramin has been found to be capable of preventing the AIDS virus from infecting and damaging T cells in vitro. Other drugs that counteract reverse transcriptase or interfere with later stages in the viral life cycle may also prove useful. Under investigation are antiviral drugs that have the added effect of stimulating the host's immune system. These drugs seem to work by inhibiting enzymes that are crucial to the synthesis of viral RNA and DNA. The genetic variability of the AIDS virus complicates vaccine development, although it may be possible to identify invariant regions of the viral envelope to which antibody can bind effectively. The genome of the AIDS virus has been found to contain a sequence known as tat, which encodes a similar regulatory protein. If the AIDS virus could be modified genetically by deletion of tat or the sequence with which the tat protein interacts, it could serve as a safe vaccine.
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PMID:The immune system in AIDS. 299 72

The genome of the HTLV-III/LAV retrovirus, the etiologic agent of the acquired immunodeficiency syndrome (AIDS), encodes the viral structural proteins (envelope and core proteins), the reverse transcriptase, a transactivation protein (tat-III), as well as two other proteins (3'orf, sor) of unknown function. We studied the prevalence of natural antibodies against envelope, gag, 3'orf, sor, and tat-III in the sera of HTLV-III infected individuals in an attempt to correlate clinical status with seropositivity to specific HTLV-III antigens. We selected 101 sera; 16 were obtained from normal donors with no known risk factors, and 85 were from patients with full-fledged AIDS (28 cases), AIDS-related complex (ARC, 22 cases), and healthy people at risk (homosexuals, intravenous [IV] drug users, relatives of AIDS patients; 35 cases). Seropositivity for antibodies against the envelope (gp41) and gag antigens (p15, p24) was determined by Western blot using disrupted HTLV-III virions. Of the 101 sera, all 16 from nonrisk donors and 3/35 from healthy at-risk donors were negative for antibodies against either the gp41 or p15 and p24. The remaining 82 sera were seropositive for either the gp41 and/or the p15 and p24. All sera were then tested against the three known HTLV-III antigens (3'orf, sor, and tat-III) that have been synthesized in bacteria. Our data indicate that all the HTLV-III antigens tested are immunogenic in vivo. No significant difference in antibody prevalence to gp41 (close to 100%) and to the 3'orf, sor, and tat-III proteins (approximately 50%) was observed with regard to stage of the disease. In contrast, the prevalence of antibodies against the core antigens decreased from approximately 100% in infected people with no clinical signs of disease to 50% in ARC and AIDS patients. The percentage of patients seropositive for all five antigens tested was increased in the AIDS group. These results indicate that the greatest antibody prevalence was obtained using viral envelope antigen and further suggest that screening with the newly identified 3'orf, sor, and tat-III proteins as antigens would confer no further diagnostic advantage. The pattern of natural antibodies observed during disease progression did not suggest any pathogenetic mechanism.
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PMID:Spectrum of natural antibodies against five HTLV-III antigens in infected individuals: correlation of antibody prevalence with clinical status. 346 97

Human CD4+ T lymphoblastoid Jurkat cells were stably transfected with two different plasmid vectors containing the cDNA of human immunodeficiency virus-type 1 (HIV-1) tat gene under the control of either the promoter of simian virus 40 (pRPneo/tat) or the long terminal repeat region of SL3 murine leukaemia virus (pRPneo/SL3/tat). Both pRPneo/tat and pRPneo/SL3/tat Jurkat cell lines showed a constant and high production of bioactive Tat in transient co-transfection assays with an HIV-1 long terminal repeat (LTR)-chloramphenicol acetyltransferase (CAT) reporter plasmid. Tat-positive and mock-transfected Jurkat cells were cultured with various cytotoxic agents, which have been associated to the progressive loss of CD4 T-lymphocytes characteristic of HIV-1 disease. In the presence of recombinant tumour necrosis factor-alpha (TNF-alpha), anti-fas antibody, Leu3a anti-CD4 antibody, the percentage of apoptosis, evaluated in a 24-72 h short-term assay, was lower (P < 0.05) in tat-positive Jurkat cells than in mock-transfected controls. The low susceptibility to the cytotoxic activity of TNF-alpha and anti-fas antibody of tat-transfected cells was confirmed by counting viable cells up to 15 d of culture. Also, recombinant Tat protein was able to prevent the increase of apoptosis induced in mock-transfected Jurkat by TNF-alpha. Of note, tat-expressing cells showed a better survival with respect to mock-transfected control cells even when acutely infected with high doses (500,000 cpm of reverse transcriptase) of HIV-1 (strain IIIB) or treated with heat-inactivated HIV-1. These data demonstrate that the expression of the regulatory HIV-1 Tat protein is able to rescue Jurkat lymphoblastoid cells from apoptosis induced by a variety of cytotoxic agents. Since Tat protein expression is restricted to the initial phases of an active HIV-1 replication, the anti-apoptotic effect of Tat could have the physiological significance of selectively protecting HIV-1 producing cells from death, at least for the time necessary to allow virus production and spreading.
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PMID:Tat-expressing Jurkat cells show an increased resistance to different apoptotic stimuli, including acute human immunodeficiency virus-type 1 (HIV-1) infection. 753 Apr 79

We have investigated two regions of the viral RNA of human immunodeficiency virus type 1 (HIV-1) as potential targets for antisense oligonucleotides. An oligodeoxynucleotide targeted to the U5 region of the viral genome was shown to block the elongation of cDNA synthesized by HIV-1 reverse transcriptase in vitro. This arrest of reverse transcription was independent of the presence of RNase H activity associated with the reverse transcriptase enzyme. A second oligodeoxynucleotide targeted to a site adjacent to the primer binding site inhibited reverse transcription in an RNase H-dependent manner. These two oligonucleotides were covalently linked to a poly(L-lysine) carrier and tested for their ability to inhibit HIV-1 infection in cell cultures. Both oligonucleotides inhibited virus production in a sequence- and dose-dependent manner. PCR analysis showed that they inhibited proviral DNA synthesis in infected cells. In contrast, an antisense oligonucleotide targeted to the tat sequence did not inhibit proviral DNA synthesis but inhibited viral production at a later step of virus development. These experiments show that antisense oligonucleotides targeted to two regions of HIV-1 viral RNA can inhibit the first step of viral infection--i.e., reverse transcription--and prevent the synthesis of proviral DNA in cell cultures.
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PMID:Sequence-specific inhibition of human immunodeficiency virus (HIV) reverse transcription by antisense oligonucleotides: comparative study in cell-free assays and in HIV-infected cells. 756 37

The TAR element is a viral regulatory element extending from +1 to +60 in the human immunodeficiency virus type 1 (HIV-1) long terminal repeat, which is critical for activation by the transactivator protein Tat. Jurkat cell lines chronically infected with viruses containing HIV-1 TAR element mutations are extremely defective for both gene expression and replication. We previously demonstrated that viruses containing mutations of the TAR RNA stem, bulge, or loop structures have 200- to 5,000-fold-reduced levels of gene expression compared with lymphoid cells harboring wild-type virus. In this study, we characterized several Jurkat cell lines infected with TAR element mutant viruses which spontaneously produced culture supernatants with wild-type-like levels of reverse transcriptase activity. These viral supernatants were used to infect Jurkat cells, and following PCR amplification of the viral long terminal repeats, their DNA sequences were analyzed. This analysis demonstrated that revertant viruses isolated from these cell lines retained the original TAR mutations but also contained additional compensatory mutations within TAR. In gel retardation analysis, recombinant Tat protein bound to higher levels to in vitro-transcribed revertant TAR RNAs than the original TAR RNA mutants. Both the original and revertant TAR elements were inserted into both chloramphenicol acetyltransferase reporter and HIV-1 proviral constructs and assayed following transfection of Jurkat cells. Constructs containing revertant TAR element mutations were capable of strong activation by Tat in contrast to constructs containing the original TAR mutations. Analysis of the secondary structure of TAR RNA sequences suggested that TAR RNA structures which differed from that of wild-type TAR were still capable of strong activation in response to Tat. These results further define critical sequences in TAR RNA that are required for tat activation. In addition, since TAR structures with lower free energy that preserve the loop and bulge structures may be favored over fully formed TAR RNA with higher stable free energy, these results implicate nascent RNA rather than the fully formed TAR RNA structure as the target for tat activation.
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PMID:Human immunodeficiency virus type 1 TAR element revertant viruses define RNA structures required for efficient viral gene expression and replication. 760 59


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