Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: UMLS:C0019693 (HIV)
 170,526 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A library of three synthetic ribozymes with randomized arms, targeting NUX, GUX and NXG triplets, respectively, were used to identify ribozyme-accessible sites on the HIV-1 LTR transcript comprising positions -533 to 386. Three cleavable sites were identified at positions 109, 115 and 161. Ribozymes were designed against these sites, either unmodified or with 2'-modifications and phosphorothioate groups, and their cleavage activities of the transcript were determined. Their biological activities were assessed in cell culture, using a HIV-1 model assay system where the LTR is a promoter for the expression of the reporter gene luciferase in a transient expression system. Intracellular efficiency of the ribozymes were determined by cotransfection of ribozyme and plasmid DNA, expressing the target RNA. Modified ribozymes, directed against positions 115 and 161, lowered the level of LTR mRNA in the cell resulting in inhibition of expression of the LTR-driven reporter gene luciferase of 87 and 61%, respectively. In the presence of Tat the inhibitions were 43 and 25%. The inactive variants of these ribozymes exhibited a similar inhibitory effect. RNase protection revealed a reduction of RNA which was somewhat stronger for the active than the inactive ribozymes, particularly for ribozyme 115. Unmodified ribozymes showed no inhibition in the cell. The third ribozyme, targeting a GUG-triplet at position 109, possessed only low cleavage activity in vitro and no inhibitory effect in cell culture.
 ...
PMID:HIV-1 LTR as a target for synthetic ribozyme-mediated inhibition of gene expression: site selection and inhibition in cell culture. 1105

The retroviral primary transcription product is a multifunctional RNA that is utilized as pre-mRNA, mRNA, and genomic RNA. The relationship between human immunodeficiency virus type 1 (HIV-1) unspliced transcripts used as mRNA for viral protein synthesis and as virion precursor RNA (vpRNA) for encapsidation remains an important question. We developed a biochemical assay to evaluate the hypothesis that prior utilization as mRNA template for protein synthesis is necessary to generate vpRNA. HIV-1-infected T cells were treated with translation inhibitors under conditions that maintain virus production. Immunoprecipitation of newly synthesized HIV-1 Gag protein revealed that de novo translation is not necessary to sustain assembly, release, or processing of Gag structural protein. Both newly synthesized protein and steady-state Gag are competent for assembly, and the extracellular accumulation of Gag is proportional to the intracellular abundance of Gag. As early as 2 h after transcription, newly synthesized RNA is detectable in cell-free virions and encapsidation is sustained upon inhibition of host cell translation. Results of both [(3)H]uridine incorporation assays and HIV-1-specific RNase protection assays (RPAs) indicate that translation inhibition reduces the absolute amounts of both cytoplasmic and virion-associated RNA. Evaluation of encapsidation efficiency by RPA revealed that the cytoplasmic availability of vpRNA is increased, indicating that HIV-1 unspliced mRNA can be rerouted to function as vpRNA. Our data contrast with results from the HIV-2 and murine leukemia virus systems and indicate that HIV-1 unspliced RNA constitutes a single functional pool that can function interchangeably as mRNA and as vpRNA.
 ...
PMID:Translation is not required To generate virion precursor RNA in human immunodeficiency virus type 1-infected T cells. 1109 Jan 50

Extracellular Nef which has been implicated in disease progression and development of AIDS induces IL-10, a potent immunosuppressive cytokine, in vitro. The present study was designed to examine whether the action of extracellular Nef is a protein tyrosine kinase (PTK)-dependent event. Anti-phosphotyrosine immunoblotting reveals that recombinant HIV-1 Nef induces rapid tyrosyl phosphorylation of several cellular proteins in human peripheral blood mononuclear cells. Pre-treatment of cells with herbimycin A, but not with genistein, significantly abolishes the Nef-induced tyrosine phosphorylation of cellular proteins. Furthermore, ELISA and RNase protection assays show that herbimycin A significantly blocks Nef-induced production of IL-10 at both the protein and the mRNA level. Genistein and aminogenistein have a much less blocking effect on the ability of Nef to induce IL-10. These results provide evidence for the involvement of a herbimycin A-sensitive PTK in the signal transduction pathway for exogenous HIV-1 Nef.
 ...
PMID:Involvement of a herbimycin A-sensitive protein tyrosine kinase in extracellular action of HIV-1 Nef. 1113 32

The human immunodeficiency virus type 1 (HIV-1) Tat protein has been reported to transactivate several cellular genes, including the potent chemotactic factor interleukin-8 (IL-8). Consistent with these in vitro assays, elevated levels of IL-8 protein are found in the serum of HIV-infected individuals. We now extend these observations by demonstrating that Tat induction of IL-8 is linked to the cell cycle. Cells that constitutively express the Tat(1-86) protein (eTat) and control cells (pCEP) were reversibly blocked at the G(1)/S border with hydroxyurea or thymidine. The cells were subsequently released, and IL-8 expression was monitored by RNase protection assays and enzyme-linked immunosorbent assay (ELISA). RNase protection assays demonstrated that IL-8 mRNA expression is transiently induced, approximately fourfold, as the Tat-expressing cells enter S phase. Consistent with the RNase protection assay, an increase in IL-8 protein was observed in the cell supernatant using an IL-8 ELISA. Similar experiments were performed following a reversible block at the G(2)/M border with nocodazole and release into G(1). Using the RNase protection assay and ELISA, little or no increase in IL-8 expression was observed during G(1). Using gel shift as well as an immobilized DNA binding assay, we demonstrate that the increase in IL-8 gene expression correlates with a specific increase in p65 NF-kappa B binding activity only in the nucleus of the Tat-expressing cells. Moreover, the CREB-binding protein coactivator is present in the complex in the Tat cell line. Finally, we demonstrate that the presence of the proteasome inhibitor MG-132 inhibits the induction of NF-kappa B binding, as well as IL-8 expression, supporting the role of NF-kappa B.
 ...
PMID:Cell cycle regulation of human interleukin-8 gene expression by the human immunodeficiency virus type 1 Tat protein. 1116 Jun 71

The methodological and logistic benefits gained from assessing in vitro antiretroviral replication using one-tube real-time RT-PCR procedures are currently diminished by a continuing need for prior RNA isolation. We now report a simple and inexpensive modification of a commercially available one-tube RT-PCR assay, consisting of detergent-based virus lysis in the presence of a ribonuclease inhibitor, which can be used to directly quantify retroviral RNA levels in culture supernatant. This approach circumvents the potential loss of RNA inherent to RNA-isolation procedures based on prior extraction and demonstrates a dynamic range of at least 4 logs. Using in vitro culture systems incorporating either HIV-1 or FIV, we show that this ability to isolate retroviral RNA directly during the RT-PCR process can provide an equivalent alternative to one of the more time and resource-consuming steps in quantifying in vitro retroviral RNA levels.
 ...
PMID:Quantification of in vitro retroviral replication using a one-tube real-time RT-PCR system incorporating direct RNA preparation. 1116 96

A human immunodeficiency virus type 1 (HIV-1)-based retroviral vector pseudotyped with HIV envelope containing the herpes simplex virus-thymidine kinase (HSV-TK) gene under the control of the HIV LTR promoter (pHXTKN) was constructed and stably transferred into human CD4(+) H9, CEM, and U937 cells. RNase protection assays did not initially detect expression of the HSV-TK gene in HXTKN-transduced CD4(+) cells (HXTKN/CD4), but expression was then efficiently induced by infection with HIV-1. MTT assays showed that after HIV-1 infection, the susceptibility of HXTKN/CD4 cells to ganciclovir (GCV) was 1000-fold higher than prior to infection. This enabled HIV-1-infected cells to be selectively killed by transduction with HXTKN followed by exposure to GCV. Because the HSV-TK gene is specifically transferred into HIV-1-permissive cells and expressed only after HIV-1 infection, the frequency of unwanted cell death should be low. Elimination of the HIV-1-infected cells effectively inhibited further spread of infectious virus. In addition, the integrated HIV vector sequences were repackaged on infection with HIV-1 and transferred to surrounding untransduced cells. These results are indicative of the potential benefits of using HIV vectors in gene therapies for the treatment of HIV-1 infection.
 ...
PMID:Selective killing of human immunodeficiency virus-infected cells by targeted gene transfer and inducible gene expression using a recombinant human immunodeficiency virus vector. 1117 60

Peptide T (ASTTTNYT), a fragment corresponding to residues 185-192 of gp120, the coat protein of HIV, is endowed with several biological properties in vitro, notably inhibition of the binding of both isolated gp120 and HIV-1 to the CD4 receptor, and chemotactic activity. Based on previous nuclear magnetic resonance (NMR) studies performed in our laboratory, which were consistent with a regular conformation of the C-terminal pentapeptide, and SAR studies showing that the C-terminal pentapeptide retains most of the biological properties, we designed eight hexapeptides containing in the central part either the TNYT or the TTNY sequence, and charged residues (D/E/R) at the two ends. Conformational analysis based on NMR and torsion angle dynamics showed that all peptides assume folded conformations. albeit with different geometries and stabilities. In particular, peptides carrying an acidic residue at the N-terminus and a basic residue at the C-terminus are characterized by stable helical structures and retain full chemotactic activity. The solution conformation of peptide ETNYTR displays strong structural similarity to the region 19-26 of both bovine pancreatic and bovine seminal ribonuclease, which are endowed with anti-HIV activity. Moreover, the frequent occurrence, in many viral proteins, of TNYT and TTNY, the two core sequences employed in the design of the hexapeptides studied in the present work, hints that the sequence of the C-terminal pentapeptide TTNYT is probably representative of a widespread viral recognition motif.
 ...
PMID:Peptide T revisited: conformational mimicry of epitopes of anti-HIV proteins. 1135 63

Human immunodeficiency virus type 1 (HIV-1) is able to establish a persistent latent infection during which the integrated provirus remains transcriptionally silent. Viral transcription is stimulated by NF-kappaB, which is activated following the exposure of infected T cells to antigens or mitogens. Although it is commonly assumed that NF-kappaB stimulates transcriptional initiation alone, we have found using RNase protection assays that, in addition to stimulating initiation, it can also stimulate elongation from the HIV-1 long terminal repeat. When either Jurkat or CCRF/CEM cells were activated by the mitogens phorbol myristate acetate and phytohemagglutinin, elongation, as measured by the proportion of full-length transcripts, increased two- to fourfold, even in the absence of Tat. Transfection of T cells with plasmids carrying the different subunits of NF-kappaB demonstrated that the activation of transcriptional elongation is mediated specifically by the p65 subunit. It seems likely that initiation is activated because of NF-kappaB's ability to disrupt chromatin structures through the recruitment of histone acetyltransferases. To test whether p65 could stimulate elongation under conditions where it did not affect histone acetylation, cells were treated with the histone deacetylase inhibitor trichostatin A. Remarkably, addition of p65 to the trichostatin A-treated cell lines resulted in a dramatic increase in transcription elongation, reaching levels equivalent to those observed in the presence of Tat. We suggest that the activation of elongation by NF-kappaB p65 involves a distinct biochemical mechanism, probably the activation of carboxyl-terminal domain kinases at the promoter.
 ...
PMID:Activation of human immunodeficiency virus transcription in T cells revisited: NF-kappaB p65 stimulates transcriptional elongation. 1150 98

From the fruiting bodies of the mushroom Lyophyllum shimeji, a novel ribosome inactivating protein with a molecular weight of 20 kDa and exhibiting antifungal activity against Physalospora piricola (IC(50) = 2.5 microM) and Coprinus comatus was isolated. The protein, designated lyophyllin, was purified by ion exchange chromatography on CM-cellulose, affinity chromatography on Affi-gel Blue Gel, and then ion exchange chromatography on Mono S. Lyophyllin possessed an N-terminal sequence with some similarity to those of plant ribosome-inactivating proteins. It inhibited translation in rabbit reticulocyte lysate with an IC(50) of 1 nM, thymidine uptake by murine splenocytes with an IC(50) of 1 microM and HIV-1 reverse transcriptase activity with an IC(50) of 7.9 nM. Lyophyllin did not manifest ribonuclease or hemagglutinating activity. An antifungal protein, designated Lyophyllum antifungal protein (LAP), with a molecular weight of 14 kDa, and an N-terminal sequence somewhat analogous to those of angiosperm thaumatin-like proteins and thaumatins and an inactive variant of the ubiquitin-conjugating enzyme, was first isolated from Lyophyllum shimeji. LAP was adsorbed on CM-cellulose, Affi-gel blue gel, and Mono S. LAP exerted antifungal activity against P. piricola (IC(50) = 70 nM) and Mycosphaerella arachidicola but not against Rhizoctonia solani, Colletotrichum gossypii, and Coprinus comatus. It exerted very low translation inhibitory activity in a rabbit reticulocyte lysate system (IC(50) = 70 microM) and negligible ribonuclease activity toward yeast transfer RNA and hemagglutinating activity toward rabbit erythrocytes. It inhibited HIV-1 reverse transcriptase with an IC(50) of about 5.2 nM. A synergism in antifungal activities of LAP and lyophyllin against P. piricola was demonstrable.
 ...
PMID:First simultaneous isolation of a ribosome inactivating protein and an antifungal protein from a mushroom (Lyophyllum shimeji) together with evidence for synergism of their antifungal effects. 1155 14

A deoxyribonuclease distinct from the previously isolated asparagus ribosome-inactivating proteins, possessing a molecular weight of 30 kDa and requiring a pH of 7.5 for optimum hydrolytic activity toward herring sperm DNA, was isolated from Asparagus officinalis seeds. The isolation procedure involved extraction with saline, (NH(4))(2)SO(4) precipitation, ion-exchange chromatography on DEAE-cellulose, affinity chromatography on Affi-gel blue gel, ion-exchange chromatography on CM-Sepharose, and FPLC gel filtration on Superdex 75. The doxyribonuclease was unadsorbed onto DEAE-cellulose and Affi-gel blue gel and adsorbed onto CM-Sepharose. It exhibited the novel N-terminal sequence, GIEVIKIREL. The deoxyribonuclease was purified to a specific activity of 1584 units/mg. It was devoid of ribonuclease, protease, and HIV-1 reverse transcriptase-inhibitory activities. However, it inhibited cell-free translation in a rabbit reticulocyte lysate system with an IC(50) of 20 microM. It exhibited antifungal activity toward Botrytis cinerea but not toward Fusarium oxysporum and Mycosphaerella arachidicola.
 ...
PMID:Isolation of a novel deoxyribonuclease with antifungal activity from Asparagus officinalis seeds. 1170 87


<< Previous 1 2 3 4 5 6 7 8 9 10 Next >>