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

Previous studies revealed that antisense oligodeoxynucleotides to specific regions of the human immunodeficiency virus-1 (HIV-1) are potent inhibitors of replication of HIV-1 in vitro (Zamecnik, P. C., Goodchild, J., Taguchi, Y., and Sarin, P. S. (1986) Proc. Natl. Acad. Sci. U. S. A. 83, 4143-4146). We now report that antisense RNA, synthesized in vitro using T7 and SP6 RNA polymerase, displayed an anti-HIV-1 effect in the HTLV-IIIB/H9 system in vitro. Treatment of HIV-1-infected H9 cells with viral env region antisense RNA encapsulated in liposomes targeted by antibodies specific for the T cell receptor molecule CD3 almost completely inhibited HIV-1 production. The viral env segment covered a part of exon II of HIV-1 tat gene. No anti-HIV activity could be detected with similarly targeted liposome-encapsulated sense env RNA or with pol RNA synthesized in either the sense or antisense orientations, or with env region antisense RNA free in solution, or encapsulated in liposomes in the absence of the targeting antibody. A semiquantitative evaluation revealed that 4000-7000 RNA molecules became cell-bound in targeted liposomes; the half-life of the intracellularly present hybridizable antisense env RNA was approximately 12 h. Western blots showed that antisense env RNA suppressed tat gene expression by approximately 90% and gp160 production by 100%. These data were confirmed by immunoprecipitation studies. Northern blots (using an env probe) demonstrated the existence of all major HIV RNA species (9.3-, 4.3-, and 2.0-kb mRNA) in HIV-infected cells treated with antisense env RNA although at a reduced level. We conclude that the antisense env RNA inhibited viral protein production at the translational level.
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PMID:Inhibition of expression of human immunodeficiency virus-1 in vitro by antibody-targeted liposomes containing antisense RNA to the env region. 169 56

An expression vector (pU3R-III/2-5AS) of human 2',5'-oligoadenylate (2-5A) synthetase was constructed in which a cDNA encoding an active form of the enzyme was located 3' to a 3'-long terminal repeat (LTR) of human immunodeficiency virus type 1 (HIV-1). The LTR-directed expression of this hybrid DNA could be activated in trans by the HIV tat gene product. This vector was used for transfection of HeLa-T4+ cells, which are permissive to HIV infection, as well as of normal HeLa cells. HIV replication after infection of the CD4-receptor-bearing HeLa-T4+ cells with HIV-1 was found to be strongly reduced when drug-selected cells cotransfected with pU3R-III/2-5AS and a hygromycin B resistance gene containing plasmid were used. In nontransfected cultures or after transfection with the selectable marker plasmid only, about 60% p17- and p24-positive cells were found 5 days after infection. However, after stable transfection with pU3R-III/2-5AS the number of positive cells was decreased to about 2%. The reverse transcriptase (RT) activity, as a measure for virus production, was markedly decreased in the culture fluids of pU3R-III/2-5AS transfected cells compared with the mock-transfected controls. In parallel experiments it was established that Tat-mediated trans-activation of HIV-1 LTR-directed 2-5A synthetase expression resulted in a great increase in both 2-5A synthetase mRNA level and activity as well as in cellular 2-5A content. Similar results were found in HeLa-T4+ cells and in HeLa cells (without CD4 receptor) cotransfected with pU3R-III/2-5AS and a tat gene containing plasmid or after introduction of purified Tat protein into the pU3R-III/2-5AS transfected cells by using a modified scrape loading procedure. These results indicate that HIV-trans-activated 2-5A synthetase can selectively inhibit HIV replication in vitro, and might be a promising gene therapeutical approach.
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PMID:Protection of HeLa-T4+ cells against human immunodeficiency virus (HIV) infection after stable transfection with HIV LTR-2',5'-oligoadenylate synthetase hybrid gene. 169 80

A frame-shift tat gene mutant of human immunodeficiency virus type 1 (HIV-1), which showed no detectable trans-activation potential, was constructed in vitro. Upon transfection, this clone directed the synthesis of virus mRNAs, gag proteins and virion-associated reverse transcriptase (RT) at a low level as was observed with the tat mutants of HIV-2 and simian immunodeficiency virus isolated from African green monkey (SIVAGM). Using these mutant viruses, trans-activation efficiency of viral gene expression by tat was compared among HIV-1, HIV-2, and SIVAGM. SIVAGM seemed to be less dependent on tat for RT production than HIV-1 and HIV-2.
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PMID:Comparative studies on tat mutants of three primate lentiviruses. 170 Jun 93

The time course of viral gene expression in H9 cells acutely infected with HIV-1 was analysed by the polymerase chain reaction (PCR). Virus-specific sequences were first detected in genomic DNA of H9 cells 1-2 h after infection. RNA for the regulatory genes such as the tat and nef appeared 2-3 h post-infection and RNA for the gag and env at 3 h. The results demonstrate that viral DNA synthesis occurs rapidly after infection of target cells followed by synthesis of viral RNA. Cell-associated reverse transcriptase activity increased after 24 h, while culture supernatant enzyme activity increased later, between 1-5 days. The delay in virus release after rapid integration and transcriptional activity suggests the involvement of additional factors, perhaps both cellular and viral, that control the formation and budding of mature virions.
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PMID:Kinetics of early HIV-1 gene expression in infected H9 cells assessed by PCR. 170 54

The herpes simplex virus type 1 (HSV-1)-mediated transactivation of human immunodeficiency virus type 1 (HIV-1) provirus was studied in cell lines containing either integrated tat-defective HIV-1 provirus (HNHIVdt4 cells) or the tat-defective HIV-1 provirus, and a plasmid in which the expression of human alpha 2 interferon (HuIFN-alpha 2) was under the control of the HIV-1 long terminal repeat (LTR) (HNHIV alpha 1 cells). In both cell lines, transcription of the HIV-1 provirus was below the limits of detection, but it could be induced effectively by transfection with a HIV-1 tat-expression plasmid. In HNHIV alpha 1 cells, HuIFN-alpha 2 was induced concomitantly with HIV-1 provirus, although these cells synthesized only low levels of IFN constitutively. In contrast, infections with HSV-1 activated transcription of HIV-1 provirus only in HNHIVdt4 cells but not in HNHIV alpha 1 cells. Similarly in a transient expression assay, HSV-1 up-regulated expression of a HIV LTR-CAT (chloramphenicol acetyltransferase gene) plasmid in HNHIVdt4 but not in HNHIV alpha 1 cells. No major differences could be detected in the expression of HSV-1 immediate-early (IE) genes IE175 and IE110 (which are essential for the activation of HIV-1 LTR) in HNHIVdt4 and HNHIV alpha 1 cells to account for the inability of HSV-1 to induce HIV-1 in HNHIV alpha 1 cells. However, major differences were observed in the binding pattern of NF-kappa B-specific nuclear proteins to the enhancer region of the HIV-1 LTR: whereas binding of the 45-kDa NF-kappa B-specific nuclear protein was detected in nuclear extracts from HNHIVdt4 cells, no protein binding was seen in extracts from HNHIV alpha 1 cells. These results suggest an alternate mechanism by which IFN may alter the expression of cellular and viral genes.
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PMID:Inhibition by interferon of herpes simplex virus type 1-activated transcription of tat-defective provirus. 171 35

The expression of Gag, Pol, Vif, Vpr, Vpu, and Env proteins from unspliced and partially spliced human immunodeficiency virus type 1 (HIV-1) mRNAs depends on the viral protein Rev, while the production of Tat, Rev, and Nef from multiply spliced mRNAs does not require Rev. To investigate the difference between gag and tat mRNAs, we generated plasmids expressing tat-gag hybrid mRNAs. Insertion of the gag gene downstream of the tat open reading frame in the tat cDNA resulted in the inhibition of Tat production. This inhibition was caused, at least in part, by a decrease in the stability of the produced mRNA. Deletions in gag defined a 218-nucleotide inhibitory sequence named INS-1 and located at the 5' end of the gag gene. Further experiments indicated the presence of more than one inhibitory sequence in the gag-protease gene region of the viral genome. The inhibitory effect of INS-1 was counteracted by the positive effect mediated by the Rev-Rev-responsive element interaction, indicating that this sequence is important for Rev-regulated gag expression. The INS-1 sequence did not contain any known HIV-1 splice sites and acted independently of splicing. It was found to have an unusually high AU content (61.5% AU), a common feature among cellular mRNAs with short half-lives. These results suggest that HIV-1 and possibly other lentiviruses have evolved to express unstable mRNAs which require additional regulatory factors for their expression. This strategy may offer the virus several advantages, including the ability to enter a state of low or latent expression in the host.
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PMID:Distinct RNA sequences in the gag region of human immunodeficiency virus type 1 decrease RNA stability and inhibit expression in the absence of Rev protein. 172 77

Transcriptional regulation of the proviral form of the human immunodeficiency virus type 1 (HIV-1) is exerted by its 5' long terminal repeat (LTR), which contains recognition sites for several cell factors. By gel retardation and DNase I footprinting experiments we have identified a binding site for a human nuclear protein between nucleotides -152 to -174 upstream of transcription start site, in a region previously recognized as a negative regulator of transcription (negative regulatory element, NRE). The recognized sequence contains the dyad symmetry element CACGTG, which represents a binding motif, very conserved through evolution, present in a putative human DNA replication origin (pB48), in the upstream element of the major late promoter (MLP-UE) of adenovirus, and, as transcriptional element, upstream of many eukaryotic genes. Common binding activities exist in human nuclear extracts for pB48, MLP-UE and the HIV-1 LTR; at least three protein species recognize the LTR sequence, of 44 (corresponding to transcription factor USF/MLTF), 70, and 110 kDa, respectively. Chloramphenicol acetyltransferase assays suggest that the USF/MLTF binding site located in the HIV-1 LTR acts as a negative regulator of transcription, and that it contributes to the overall negative function exerted by the NRE. An oligonucleotide corresponding to another characterized human USF/MLTF binding site can functionally replace part of the activity of the NRE. This negative function is exerted both in presence or absence of tat transactivation, in different cell lines, and after PMA stimulation.
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PMID:A human binding site for transcription factor USF/MLTF mimics the negative regulatory element of human immunodeficiency virus type 1. 172 95

We have used a panel of cDNA clones expressing wild-type and mutant human immunodeficiency virus type 1 (HIV-1) mRNAs to study translation of these mRNAs in eucaryotic cells. The tat open reading frame (ORF) has a strong signal for translation initiation, while rev and vpu ORFs have weaker signals. The expression of downstream ORFs is inhibited in mRNAs that contain the tat ORF as the first ORF. In contrast, downstream ORFs are expressed efficiently from mRNAs that have rev or vpu as the first ORF. All env mRNAs contain the upstream vpu ORF. Expression of HIV-1 Env protein requires a weak vpu AUG, which allows leaky scanning to occur, thereby allowing ribosomes access to the downstream env ORF. We concluded that HIV-1 mRNAs are translated by the scanning mechanism and that expression of more than one protein from each mRNA was caused by leaky scanning at the first AUG of the mRNA.
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PMID:Mechanism of translation of monocistronic and multicistronic human immunodeficiency virus type 1 mRNAs. 172 99

Sera were obtained from 50 individuals infected with human immunodeficiency virus type 1 or from HIV-1-uninfected individuals before or after vaccination with recombinant gp160. These sera were evaluated for activity antagonistic to the cell-killing activity of the chimeric Pseudomonas exotoxin hybrid protein, sCD4-PE40. For these studies, Chinese hamster ovary (CHO) cells were transfected with a chimeric plasmid encoding the tat, rev, and envelope genes of HIV-1 and a cell line was selected for stable expression of the envelope glycoproteins at the cell surface (CHO-env). Cytotoxicity of sCD4-PE40 for CHO-env in the presence or absence of added human serum was quantitated spectrophometrically following enzymatic reduction of a tetrazolium bromide within the mitochondria of viable cells (MTT assay). Several HIV+ sera inhibited the cytotoxic activity of sCD4-PE40; the antagonist had properties consistent with those of immunoglobulins in that it was heat stable, absorbed by protein A, and reversible by increasing the concentration of sCD4-PE40. Of 15 HIV+ sera which strongly reacted with gp120, 11 (73%) also potently inhibited sCD4-PE40 cytotoxicity, and cytotoxicity was inhibited by sera from some HIV- individuals after, but not before, immunization with gp160. These data suggested a role for antibody to gp120 in the antagonistic activity. However, not all sera with antibody to gp120 antagonized sCD4-PE40 cytotoxicity and high levels of antagonist activity were frequently (40%) found in HIV+ sera lacking immunoblot-detectable antibody to gp120, or antibody to either CD4 or PE40. Grouping of the HIV+ sera according to the patients' absolute number of CD4+ cells revealed that the degree of inhibition of sCD4-PE40 cytotoxicity approached a Gaussian distribution, suggesting that persons with CD4+ cell counts between 200 and 700/mm3 may be more likely to possess significant levels of serum antagonist. This data have implications for the clinical development of sCD4-PE40 or other sCD4-based therapeutics in the management of HIV-1 infection.
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PMID:Soluble CD4-PE40 is cytotoxic for a transfected mammalian cell line stably expressing the envelope protein of human immunodeficiency virus (HIV-1), and cytotoxicity is variably inhibited by the sera of HIV-1-infected patients. 174 81

The relation of the initial products of the HIV-1 gag gene to the final products was determined in virus samples and cell fractions of infected H9 and Jurkat-tat cell cultures. The proteins were identified by immunoblotting with pooled sera from AIDS patients or monoclonal antibodies. The proportion in the virions of gag precursor proteins and the products of their proteolytic cleavage varied according to the maturity of the virus particles as determined by electron microscopy. The distribution of viral gag proteins in the cell fractions was determined 2, 4, and 24 h after infection. Treatment of cells with cycloheximide to block de novo protein synthesis did not significantly affect the results. Gag proteins containing the N terminus of the precursor p55 (including p55, the intermediate precursors p41(45) and p39, and mature protein p17) were found in the cell nuclei up to 24 h after infection. The major core protein p24 was located in the cytoplasmic fraction. These data strongly suggest that gag precursors from the p55 N terminus and the matrix protein p17 enter the infected cell separately from the major core protein p24, or become separated from it in the cytoplasm.
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PMID:HIV-1 gag proteins in virions and in infected cell fractions. 175 61


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