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

Human immunodeficiency virus type 1 (HIV-1) Tat is a potent transcriptional activator of the HIV-1 promoter and also has the ability to modulate a number of cellular regulatory circuits including apoptosis. Tat exerts its effects through interaction with viral as well as cellular proteins. Here, we studied the influence of p73, a protein that is implicated in apoptosis and cell cycle control, on Tat functions in the central nervous system. Protein interaction studies using immunoprecipitation followed by Western blot and glutathione S-transferase pull-down assays demonstrated the association of Tat with p73. Tat bound to the N-terminal region of p73 spanning amino acids 1 to 120, and this interaction required the cysteine-rich domain (amino acids 30 to 40) of Tat. Association of p73 with Tat prevented the acetylation of Tat on lysine 28 by PCAF. Functional studies including RNA interference showed that p73 inhibited Tat stimulation of the HIV-1 promoter. Furthermore, p73 prevented the interaction of Tat with cyclin T1 in vitro but not in vivo. These findings suggest possible new therapeutic approaches, using p73, for Tat-mediated AIDS pathogenesis.
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PMID:p73 Interacts with human immunodeficiency virus type 1 Tat in astrocytic cells and prevents its acetylation on lysine 28. 1613 3

ELDKWA on HIV-1 gp41 is a conserved epitope recognized by one broadly neutralizing monoclonal antibody 2F5, which is a promising candidate target for vaccine design. Here we report two ELDKWA-specific monoclonal antibodies (mAbs), 18F11 and 7E10, that were screened from the splenocytes of mice immunized by recombinant GST-(ELDKWA)4 protein. In further evaluation, these mAbs exhibited appreciable neutralizing activities against HIV-1 primary isolate 92US675 (clade B) with IC50 (50% inhibition concentration) of 6.84 +/- 0.36 microg/ml and 10.66 +/- 1.69 microg/ml, respectively. Unexpectedly, neither of these two murine mAbs could neutralize laboratory-adapted strain HIV-1 IIIB (clade B). As a control, human mAb 2F5 neutralized both primary and laboratory-adapted strains. These data strongly suggest that ELDKWA-specific antibodies induced by different antigenic formats show different neutralizing activities against HIV-1, which implies another complication in the development of effective vaccines.
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PMID:Neutralization of HIV-1 primary isolate by ELDKWA-specific murine monoclonal antibodies. 1632 2

The CXCR4 chemokine receptor is a G protein-coupled receptor that plays an important role in leukocyte homing, cancer metastasis, and human immunodeficiency virus infection. In response to ligand stimulation, chemokine receptors undergo endocytosis through clathrin-coated vesicle (CCV). Uncoating of CCV, a process involving heat shock cognate protein and several other proteins, is critical for fusion of CCV to endosomal compartments. The present study demonstrated that CXCR4 was associated with the 73-kDa heat shock cognate protein (Hsc73) in human embryonic kidney 293 cells in response to ligand stimulation. Truncation of the carboxyl terminal domain of CXCR4 reduced the association with Hsc73 and a glutathione S-transferase-CXCR4 carboxyl terminal fusion protein associated with Hsc73 in vitro, suggesting involvement of the carboxyl terminal domain of the receptor in the interaction. In response to ligand stimulation, CXCR4 underwent internalization and colocalization with Hsc73, but the receptor endocytosis was blocked by knockdown of Hsc73 with RNA interference. Moreover, Hsc73 knockdown significantly reduced the CXCR4-mediated chemotaxis of U87 glioma cell lines. These findings suggest that Hsc73 plays a role in chemokine receptor trafficking and the receptor-mediated chemotaxis.
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PMID:The 73-kDa heat shock cognate protein is a CXCR4 binding protein that regulates the receptor endocytosis and the receptor-mediated chemotaxis. 1636 78

Previous examination of the effect of TCF-4 on transcription of the human immunodeficiency virus type 1 (HIV-1) promoter in human astrocytic cells found that TCF-4 affects the HIV-1 promoter through the GC-rich domain (nt -80 to nt -68). Here, the physical interaction and a functional consequence of TCF4-Sp1 contact were characterized. It was shown that expression of TCF-4 in U-87 MG (human astrocytic) cells decreased basal and Sp1-mediated transcription of the HIV-1 promoter. Results from a GST pull-down assay, as well as combined immunoprecipitation and Western blot analysis of protein extracts from U-87 MG cells, revealed an interaction of Sp1 with TCF-4. Using in vitro protein chromatography, the region of Sp1 that contacts TCF-4 was mapped to aa 266-350. It was also found that, in cell-free extracts, TCF-4 prevented dsDNA-dependent protein kinase (DNA-PK)-mediated Sp1 phosphorylation. Surprisingly, TCF-4 failed to decrease Sp1-mediated transcription of the HIV-1 long terminal repeat (LTR) and Sp1 phosphorylation in cells expressing HIV-1 Tat. Results from immunoprecipitation/Western blotting demonstrated that TCF-4 lost its ability to interact with Sp1, but not with Tat, in Tat-transfected cells. Taken together, these findings suggest that activity at the HIV-1 promoter is influenced by phosphorylation of Sp1, which is affected by Tat and DNA-PK. Interactions among TCF-4, Sp1 and/or Tat may determine the level of viral gene transcription in human astrocytic cells.
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PMID:Human immunodeficiency virus type 1 Tat prevents dephosphorylation of Sp1 by TCF-4 in astrocytes. 1669 Sep 26

Human immunodeficiency virus type 1 (HIV-1) viral assembly is mediated by multiple protein-protein and protein-nucleic acid interactions. Human tRNA(Lys3) is used as the primer for HIV reverse transcription, and HIV Gag and GagPol are required for packaging of the tRNA into virions. Human lysyl-tRNA synthetase (LysRS) is also specifically packaged into HIV, suggesting a role for LysRS in tRNA packaging. Gag alone is sufficient for packaging of LysRS, and these two proteins have been shown to interact in vitro using glutathione S-transferase pull-down assays. In vitro pull-down assays using truncated constructs have also revealed that residues important for homodimerization of Gag and LysRS are critical for the Gag/LysRS interaction. In this work, we report further in vitro characterization of the interaction between HIV Gag and human LysRS using affinity pull-down assays, fluorescence anisotropy measurements and gel chromatography. An equilibrium binding constant of 310 +/- 80 nM was measured for the Gag/LysRS interaction. We also show that capsid alone binds to LysRS with a similar affinity as full-length Gag. Point mutations that disrupt the homodimerization of LysRS and Gag in vitro do not affect their interaction. These results suggest that dimerization of each protein per se is not required for the interaction but that residues involved in forming the homodimer interfaces contribute to heterodimer formation. Gel chromatography studies further support the formation of a Gag/LysRS heterodimer.
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PMID:In vitro characterization of the interaction between HIV-1 Gag and human lysyl-tRNA synthetase. 1670 15

Human immunodeficiency virus type 1 (HIV-1) infection of human macrophages can be inhibited by antibodies which bind to the tetraspanin protein CD63, but not by antibodies that bind to other members of the tetraspanin family. This inhibitory response was limited to CCR5 (R5)-tropic virus and was only observed using macrophages, but not T cells. Here, we show that recombinant soluble forms of the large extracellular domain (EC2) of human tetraspanins CD9, CD63, CD81, and CD151 produced as fusion proteins with glutathione S-transferase (GST) can all potently and completely inhibit R5 HIV-1 infection of macrophages with 50% inhibitory concentration values of 0.11 to 1.2 nM. Infection of peripheral blood mononuclear cells could also be partly inhibited, although higher concentrations of EC2 proteins were required. Inhibition was largely coreceptor independent, as macrophage infections by virions pseudotyped with CXCR4 (X4)-tropic HIV-1 or vesicular stomatitis virus (VSV)-G glycoproteins were also inhibited, but was time dependent, since addition prior to or during, but not after, virus inoculation resulted in potent inhibition. Incubation with tetraspanins did not decrease CD4 or HIV-1 coreceptor expression but did block virion uptake. Colocalization of fluorescently labeled tetraspanin EC2 proteins and HIV-1 virions within, and with CD4 and CXCR4 at the cell surfaces of, macrophages could be detected, and internalized tetraspanin EC2 proteins were directed to vesicular compartments that contained internalized dextran and transferrin. Collectively, the data suggest that the mechanism of inhibition of HIV-1 infection by tetraspanins is at the step of virus entry, perhaps via interference with binding and/or the formation of CD4-coreceptor complexes within microdomains that are required for membrane fusion events.
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PMID:Recombinant extracellular domains of tetraspanin proteins are potent inhibitors of the infection of macrophages by human immunodeficiency virus type 1. 1677 36

LEDGF/p75 binding-defective IN mutant viruses were previously characterized as replication-defective, yet RNAi did not reveal an essential role for the host factor in HIV-1 replication. Correlative analyses of protein binding and viral fitness were expanded here by targeting 12 residues at the IN-LEDGF/p75 binding interface. Whereas many of the resultant viruses were defective, the majority of the INs displayed wild-type in vitro integration activities. Though an overall trend of parallel loss of LEDGF/p75 binding and HIV-1 infectivity was observed, a strict correlation was not. His-tagged IN(A128Q), derived from a phenotypically wild-type virus, failed to pull-down LEDGF/p75, but IN(A128Q) was effectively recovered in a reciprocal GST pull-down assay. Under these conditions, IN(H171A), also derived from a phenotypically wild-type virus, interacted less efficiently than a previously described interaction-defective mutant, IN(Q168A). Thus, the relative affinity of the in vitro IN-LEDGF/p75 interaction is not a universal predictor of IN mutant viral fitness.
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PMID:Structure-based mutagenesis of the integrase-LEDGF/p75 interface uncouples a strict correlation between in vitro protein binding and HIV-1 fitness. 1695 83

The Tat protein from HIV-1 fused with heterologous proteins traverses biological membranes in a transcellular process called: protein transduction. This has already been successfully exploited in various biological models, but never in the nematode worm Caenorhabditis elegans. TAT-eGFP or GST-eGFP proteins were fed to C. elegans worms, which resulted in the specific localization of Tat-eGFP to epithelial intestinal cells. This system represents an efficient tool for transcellular transduction in C. elegans intestinal cells. Indeed, this approach avoids the use of tedious purification steps to purify the TAT fusion proteins and allows for rapid analyses of the transduced proteins. In addition, it may represent an efficient tool to functionally analyze the mechanisms of protein transduction as well as to complement RNAi/KO in the epithelial intestinal system. To sum up, the advantage of this technology is to combine the potential of bacterial expression system and the Tat-mediated transduction technique in living worm.
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PMID:Tat-mediated protein delivery in living Caenorhabditis elegans. 1714 Nov 80

Nef, an accessory protein of human and simian immunodeficiency viruses, is a critical determinant of pathogenesis that promotes the progression from infection to AIDS. The pathogenic effects of Nef are in large part dependent on its ability to downregulate the macrophage and T-cell coreceptor, CD4. It has been proposed that Nef induces downregulation by linking the cytosolic tail of CD4 to components of the host-cell protein trafficking machinery. To identify these components, we developed a novel Nef-CD4 downregulation system in Drosophila melanogaster S2 cells. We found that human immunodeficiency virus type 1 (HIV-1) Nef downregulates human CD4 in S2 cells and that this process is subject to the same sequence requirements as in human cells. An RNA interference screen targeting protein trafficking genes in S2 cells revealed a requirement for clathrin and the clathrin-associated, plasma membrane-localized AP2 complex in the downregulation of CD4. The requirement for AP2 was confirmed in the human cell line HeLa. We also used a yeast three-hybrid system and glutathione S-transferase pull-down analyses to demonstrate a robust, direct interaction between HIV-1 Nef and AP2. This interaction requires a dileucine motif in Nef that is also essential for downregulation of CD4. Together, these results support a model in which HIV-1 Nef downregulates CD4 by promoting its accelerated endocytosis by a clathrin/AP2 pathway.
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PMID:Downregulation of CD4 by human immunodeficiency virus type 1 Nef is dependent on clathrin and involves direct interaction of Nef with the AP2 clathrin adaptor. 1726

Human immunodeficiency virus type 2 (HIV-2) Vpx is required for nuclear translocation of the viral preintegration complex (PIC) in quiescent cells. In order to decipher the mechanism of action of Vpx, a cDNA library was screened with the yeast two-hybrid assay, resulting in the identification of heat shock protein 40, Hsp40/DnaJB6, as a Vpx-interactive protein. Interaction with Vpx was confirmed by glutathione S-transferase (GST) pull-down and coimmunoprecipitation assays. Overexpression of Hsp40/DnaJB6 enhanced Vpx nuclear import, whereas overexpression of a nuclear localization mutant of Hsp40/DnaJB6 (H31Q) or down-regulation of Hsp40/DnaJB6 by small interfering RNA (siRNA) reduced the nuclear import of Vpx. Hsp40/DnaJB6 competed with the Pr55(Gag) precursor protein for the binding of Vpx and incorporation into virus-like particles. Overexpression of Hsp40/DnaJB6 promoted viral PIC nuclear import, whereas siRNA down-regulation of Hsp40/DnaJB6 inhibited PIC nuclear import. These results demonstrate a role for Hsp40/DnaJB6 in the regulation of HIV-2 PIC nuclear transport.
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PMID:Hsp40 facilitates nuclear import of the human immunodeficiency virus type 2 Vpx-mediated preintegration complex. 1803 1


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