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Query: UMLS:C0019693 (HIV)
 170,526 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Ribonucleases appear to have physiologic roles in host defense against cancer, viruses, and other parasites. Previously it was shown that select ribonucleases added to cells concurrently with virions blocked human immunodeficiency virus, type I (HIV-1) infection of H9 cells. We now report that a ribonuclease homologous to RNase A, named onconase, inhibits virus replication in chronically HIV-1-infected human cells without killing the virally infected cell. Examining the mechanism of this inhibition shows that onconase enters the infected cells and degrades HIV-1 RNA without degrading ribosomal RNA or the three different cellular messenger RNAs analyzed. The homologous human pancreatic RNase lacks anti-viral activity. Comparing recombinant forms of onconase and a onconase-human RNase chimera shows that the N-terminal 9 amino acids and the pyroglutamyl residue of onconase are required for full anti-viral activity. Thus extracellular ribonucleases can enter cells, metabolize select RNAs, and inhibit HIV virion production within viable replicating cells.
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PMID:Inhibition of HIV-1 production and selective degradation of viral RNA by an amphibian ribonuclease. 870 32

Unique transcriptional transactivation by the human immunodeficiency virus type 1 (HIV-1) Tat protein of long terminal repeat (LTR)-driven RNA expression, in the absence of the transactivator responsive element (TAR), was previously demonstrated in central nervous system (CNS)-derived astrocytic cell-lines, including U87MG. In the present study, RNase protection assays were utilized to reveal the molecular mechanism(s) underlying transactivation of the HIV-1-LTR in these cells. Short transcripts, which represent abortive HIV-1 transcription, could not be detected either in the absence or presence of Tat, and no differences in transcript levels were detected using 5' probes, as compared to 3' probes, in the experiments. Thus, the transactivational effects of Tat, in U87MG cells, were potentially based on the increase of transcriptional initiation, both in TAR-dependent and -independent states. Further, by using newly established stable cellular transformant, containing HIV-1-LTR-reporter gene constructs, TAR-independent transactivation was demonstrated to efficiently function primarily in transiently-transfected U87MG cells. U87MG cells, stably-transfected with the intact HIV-1 proviral genome, produced very low levels of virus after long-term culture, as previously reported in other astrocytic cells. These cells demonstrated profoundly restricted transcription of the HIV-1 genome, with no detectable levels of HIV-1-specific RNA by Northern blotting, indicating that the restriction of viral production in these cells is principally due to the low level of overall transcription from the 5' HIV-1-LTR. Transcription of HIV-1 RNA in this cell could not be significantly up-regulated by various stimulators, such as phorbol 12-myristate 13-acetate (PMA), tumor necrosis factor-alpha (TNF-alpha) and sodium butyrate. These data suggest that the restriction of HIV-1 transcription in these cells may be controlled by different mechanism(s) from those in lymphocytic or monocytic cells.
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PMID:Mechanisms of transcriptional transactivation and restriction of human immunodeficiency virus type I replication in an astrocytic glial cell. 871 Mar 70

We have analyzed by the RNase-A mismatch method 35 isolates from four WHO-sponsored vaccine evaluation sites as a secondary laboratory of the WHO Network for HIV Isolation and Characterization. The application of an estimator for the establishment of genetic distances based on the RNase-A digestion patterns in combination with the phylogenetic analysis has allowed us to construct a tree with five well defined groups of viruses. Because the clustering with known reference strains, samples from Brazil could be grouped as subtype B and the majority of those from Thailand were subtype E. Some of the samples from Uganda were classified as subtype D. Isolates from Rwanda and some from Uganda were identified as subtype A viruses. These results coincide with data obtained by heteroduplex mobility assay and nucleotide sequencing in env regions. The RNase-A mismatch method combined with phylogenetic analysis permitted the primary genetic classification of 33 of 35 samples from the WHO Network.
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PMID:Primary genetic characterization of HIV-1 isolates from WHO-sponsored vaccine evaluation sites by the RNase-A mismatch method. 883 88

The expression of antiviral genes in human hematopoietic stem or progenitor cells has been proposed as a strategy for gene therapy of AIDS. To be successful, this strategy requires safe and efficient transfer of the therapeutic gene into hematopoietic cells and gene expression has to be maintained in HIV susceptible cells following differentiation. We have used retroviral vectors to transfer the gene for a transdominant inhibitor of HIV replication (RevM10) into CD34+ stem/progenitor cells isolated from human umbilical cord blood (UCB). Following transduction, cells were allowed to differentiate either in vitro in clonogenic assays and long-term stromal cell cultures or in human thymus implanted in immunodeficient scid/scid mice in vivo (SCID-hu). Following differentiation and expansion, multiple lineages of cells were shown to carry the transgene. A higher percentage of gene-marked progenitor cells (10-30% in most cases) were detected in methylcellulose colony assays and in long-term stromal cell cultures (1-5%). In contrast, gene-marked T cells derived from transduced CD34+ cells in a SCID-hu model were detected at an even lower frequency (0.01-1%). RevM10 RNA expression was detected in CD34+ cells immediately after transduction and was maintained after in vitro differentiation of those cells into CD14+ myeloid cells. In T cells, the RevM10-specific RNA was detectable by RT-PCR and also by semiquantitative RNase protection. These findings demonstrate that LTR-driven gene expression is sustained in relevant cells derived from retrovirus-transduced hematopoietic progenitor cells after extensive differentiation in vitro and in vivo and suggest that stringent in vivo, rather than in vitro assays, may be a better preclinical system to improve gene marking and expression in hematopoietic cells.
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PMID:Sustained retroviral gene marking and expression in lymphoid and myeloid cells derived from transduced hematopoietic progenitor cells. 885 97

Rauscher murine leukemia virus induces an erythroleukemia in susceptible strains of mice that is associated with splenomegaly and viremia. This animal model has been used for evaluating the in vivo efficacy of potential anti-HIV agents. The in vivo antiviral activity of therapeutic agents has usually been determined by measuring a reduction in the spleen weights of compound-treated mice or by quantitating viremia with the UV-XC plaque assay. The UV-XC assay, however, is time-consuming and labor-intensive. Virions of Rauscher murine leukemia virus, like other retroviruses, contain the enzyme reverse transcriptase. Quantitating the level of this enzyme in infected mouse sera provides a more rapid measure of viremia in the animal. We have examined the effects of several reagents, including detergent, KCl, EGTA, dGMP, spermine, as well as protease and RNase inhibitors, on the reverse transcriptase assay. The optimized assay method was effective in evaluating the antiviral activity of AZT in the Rauscher murine leukemia virus in vivo model. The assay is also amenable to automation if large numbers of assays are required.
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PMID:Optimization of the reverse transcriptase assay for the detection of viral burden in mice infected with Rauscher murine leukemia virus. 891 Jun 49

Current research indicates that the nucleocapsid protein (NCp7) of human immunodeficiency virus type 1 (HIV-1) interacts with a variety of RNA substrates during the progression of the viral life cycle. The RNA features specifically recognized by the protein, however, have yet to be identified. SELEX was used to generate a set of RNAs whose affinities for nucleocapsid were on the order of 2 x 10(-9) M. Comparative analysis revealed that each RNA contains a highly conserved fourteen nucleotide sequence-block. Computer modeling and structure probing experiments indicate that the RNA ligands use the consensus sequence to fold into hairpins with an identical asymmetric bulge. The presence of the nucleocapsid protein protects the asymmetric bulge from ribonuclease attack, suggesting that it is the key element in protein recognition. A search for similar structural motifs within the HIV genome reveals several potential interaction sites for the nucleocapsid protein.
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PMID:A specific RNA structural motif mediates high affinity binding by the HIV-1 nucleocapsid protein (NCp7). 891 17

Despite extensive investigation, the pathogenesis of T cell depletions that characterize AIDS has not been elucidated. To study this process further, we evaluated T cell antigen receptor beta-chain variable gene (TCRBV) repertoires in peripheral blood lymphocytes (PBL) of 23 HIV-infected patients. Expression levels of 28 TCRBV were determined by multiprobe RNase protection assay after polymerase chain reaction (PCR) amplifications. Abnormal expansions (> 2 s.d. from mean normal values) were frequent in HIV CD4, accounting for 26% of total measured TCRBV in this population. The number and magnitude of abnormalities among individuals were inversely proportional to their CD4 counts (P < 0.012 and P < 0.01, respectively). While abnormalities were not randomly distributed among TCRBV subfamilies, no particular genes were expanded or contracted among all patients. Only 14% of CD8 TCRBV were proportionally expanded (P < 0.01 compared with CD4), and there were limited concordances between paired CD8 and CD4 repertoires among individuals. CDR3 length analyses and TCRBV sequencing showed that most CD4 expansions comprised clonal or oligoclonal populations. Thus, T cell responses in HIV patients are characterized by severe TCRBV biases and clonal expansions among CD4 subsets, and these processes are exaggerated with disease progression. The heterogeneity and oligoclonality of the TCRBV expansions are consistent with responses to HIV-encoded or other conventional antigens rather than superantigenic effects. The presence of CD4 clonal proliferations in these patients may be important in the pathogenesis of HIV, and the absence or reduction of many T cell specificities due to oligoclonal expansions may increase susceptibility to opportunistic infections.
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PMID:T cell receptor (TCR) BV gene repertoires and clonal expansions of CD4 cells in patients with HIV infections. 901 Feb 52

HIV-1 reverse transcriptase (RT) is multifunctional, with RNA-dependent DNA polymerase (RDDP), DNA-dependent DNA polymerase (DDDP), and ribonuclease H (RNase H) activities. N-(4-tert-Butylbenzoyl)-2-hydroxy-1-naphthaldehyde hydrazone (BBNH) inhibited both the polymerase and the RNase H activities of HIV-1 RT in vitro. IC50 values for inhibition of RDDP were 0.8-3.4 microM, depending on the template/primer (T/P) used in the assay. The IC50 for DDDP inhibition was about 12 microM, while that for inhibition of RNase H was 3.5 microM. EC50 for inhibition of HIV-1 replication in cord blood mononuclear cells was 1.5 microM. BBNH inhibition of RNase H in vitro was time-dependent, whereas inhibition of RT polymerase activities was immediate. BBNH was a linear mixed-type inhibitor of RT RDDP activity with respect to both T/P and to dNTP, whereas BBNH inhibition of RT RNase H activity was linear competitive. Protection experiments using an azidonevirapine photolabel showed that BBNH binds to the non-nucleoside RT inhibitor (NNRTI) binding pocket. Importantly, the compound inhibited recombinant RT containing mutations associated with high-level resistance to other NNRTI. While BBNH did not inhibit the DNA polymerase activities of other retroviral reverse transcriptases and DNA polymerases, the compound inhibited Escherichia coli RNase HI and the RNase H activity of murine leukemia virus RT. BBNH also inhibited HIV-1 RT RNase H in the presence of high concentrations of other non-nucleoside inhibitors with higher affinities for the NNRTI binding pocket, and of RT in which the NNRTI binding pocket had been irreversibly blocked by the azidonevirapine photolabel. We conclude that BBNH may therefore bind to two sites on HIV-1 RT. One site is the polymerase non-nucleoside inhibitor binding site and the second may be located in the RNase H domain. BBNH is therefore a promising lead compound for the development of multisite inhibitors of HIV-1 RT.
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PMID:Inhibition of the ribonuclease H and DNA polymerase activities of HIV-1 reverse transcriptase by N-(4-tert-butylbenzoyl)-2-hydroxy-1-naphthaldehyde hydrazone. 911 94

Monocyte chemotactic protein-1 (MCP-1) interacts with the chemokine receptor CCR2. Two CCR2 cDNAs have been described. Sequence analysis as well as Northern blotting and RNase protection with different probes revealed that the CCR2 gene is expressed in activated natural killer (NK) cells and mononuclear phagocytes as a predominant long transcript (3.4 kb) consisting of CCR2B followed by a novel sequence (X), corresponding to an intron in the genome, and by a CCR2A specific portion. The predominant long transcript is polyadenylated and present in the cytoplasm. We found that bacterial products and cytokines affect CCR2 expression. Interleukin-2 (IL-2) augmented CCR2 mRNA in monocytes and NK cells. The augmented migratory capacity of IL-2-activated versus resting NK cells was associated with increased CCR2 transcript levels. Lipopolysaccharide (LPS) and other microbial agents caused a rapid and drastic reduction of CCR2 mRNA levels. The rate of nuclear transcription of CCR2 was not affected by LPS, whereas the mRNA half life was reduced. These results suggest that regulation of receptor expression, in addition to agonist production, is probably a crucial point in the regulation of the chemokine system. Down-regulation of chemokine receptor expression may play a role in the modulation of HIV infection in macrophages by LPS. Levels of MCP-1 were markedly elevated in the cerebrospinal fluid (CSF) but not in blood of HIV-infected patients with cytomegalovirus (CMV) encephalitis. The CSF levels of MCP-1 in CMV encephalitis were markedly higher than those found in the CSF of HIV-infected patients with or without unrelated neurological diseases. IL-8, the prototype of C-X-C chemokines and RANTES and macrophage inflammatory protein-1 alpha (C-C chemokines) were not substantially increased in the liquor of CMV encephalitis patients. High levels of MCP-1 may underlie monocyte recruitment and tissue damage in CMV encephalitis and may represent a rapid and useful tool in the diagnostic armamentarium for neurological disorders associated with HIV.
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PMID:MCP-1 and CCR2 in HIV infection: regulation of agonist and receptor expression. 922 89

Histone mRNAs are naturally intronless and accumulate efficiently in the cytoplasm. To learn whether there are cis-acting sequences within histone genes that allow efficient cytoplasmic accumulation of RNAs, we made recombinant constructs in which sequences from the mouse H2a gene were cloned into a human beta-globin cDNA. By using transient transfection and RNase protection analysis, we demonstrate here that a 100-bp sequence within the H2a coding region permits efficient cytoplasmic accumulation of the globin cDNA transcripts. We also show that this sequence appears to suppress splicing and can functionally replace Rev and the Rev-responsive element in the cytoplasmic accumulation of unspliced HIV-1-related mRNAs. Like the Rev-responsive element, this sequence acts in an orientation-dependent manner. We thus propose that the sequence identified here may be a member of the cis-acting elements that facilitate the cytoplasmic accumulation of naturally intronless gene transcripts.
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PMID:The mouse histone H2a gene contains a small element that facilitates cytoplasmic accumulation of intronless gene transcripts and of unspliced HIV-1-related mRNAs. 929 70


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