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

Although multiple forms of the HIV-1 Tat protein are synthesized during infection from alternatively spliced mRNAs, only 72 amino acid residues encoded in the first Tat exon are necessary for full transactivation activity. We used limited proteolytic digestions of proteins expressed in vitro to study the structures of three Tat proteins from isolate HXB2: 72R Tat (first Tat exon); 86R Tat (first and second Tat exons), Tev or TNV (first Tat exon plus Env and Rev exons). For the 86R and Tev proteins, either trypsin or chymotrypsin cleaved the majority of carboxyl residues from the first exon. Moreover, when released from carboxyl residues, the first exon of 86R and Tev was relatively resistant to subsequent proteolysis. The entire 72R Tat protein was relatively resistant to proteolysis. The protease-resistant first exon in all Tat proteins was abolished by EDTA treatment, suggesting that divalent cations are required for its conformation. Our results suggest that the first exon in the 86R, 72R, and Tev proteins is folded into a similar structure which, as defined by partial proteolysis, acts as a single biochemical domain.
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PMID:Limited proteolytic digestions identify common structural features of HIV-1 Tat proteins expressed during infection from alternatively spliced mRNAs. 845 40

Standard methods for determining absolute CD3+CD4+ T-lymphocyte concentrations using combined results from flow cytometry and hematology analyzers are expensive and require fresh (< 18 h old) blood. We evaluated four new "alternative" methods for determining absolute CD4+ T-lymphocyte counts; (a) the FACSCount System (Becton Dickinson Immunocytometry Systems), (b) VCS Technology/Coulter Cyto-Spheres (Coulter Corporation), (c) Zymmune CD4/CD8 Cell Monitoring Kit (Zynaxis, Inc.), and (d) TRAx CD4 Test Kit (T Cell Diagnostics). EDTA-anticoagulated whole-blood specimens from HIV-seropositive patients and anti-HIV-negative adult blood donors and staff were tested by the standard flow cytometry method on fresh blood (0-6 postphlebotomy) and by the four alternative methods at 0 to 6 h and 24 to 30 h postphlebotomy. Representative specimens (< 6 h old) were tested five times with each system to evaluate reproducibility. Correlation coefficients for absolute CD4+ counts between standard flow cytometry and the alternative methods ranged from 0.84 to 0.92 for both fresh and 24- to 30-h-old specimens. Average coefficient of variation for reproducibility ranged from 4.5 to 7.1%. The four alternative CD4+ T-lymphocyte counting methods performed well relative to standard methods. Each alternative method offers advantages over standard flow cytometry with respect to sample throughout, required technical expertise, and cost per result. These methods should facilitate wider availability of low-cost CD4 counts.
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PMID:Evaluation of four alternative methodologies for determination of absolute CD4+ lymphocyte counts. The National Heart, Lung, and Blood Institute Retrovirus Epidemiology Donor Study. 854 31

This investigation was performed to determine whether HTLV-I can activate complement, since previous studies show that complement activation by some viruses, including HIV-1, can enhance binding to, and infection of complement receptor-positive (CR+) cells. Complement treatment increased binding of HTLV-I to CR+ HPB-ALL cells by approximately 5-fold. In contrast, increased binding was not observed with H9 cells, which lack CR. Heat inactivation or EDTA treatment of complement blocked this increased binding while EGTA treatment only partially blocked binding. Anti-CR2 antibody significantly blocked binding of complement-treated HTLV-I to HPB-ALL cells. Since previous studies showed that HIV-1 could activate complement, activation of complement by this virus was compared with HTLV-I. It was observed that binding of HTLV-I to HPB-ALL cells was enhanced by highly dilute complement (> or = 1:810) while HIV-1 required much higher concentrations of complement (> or = 1:30), indicating that HTLV-I is a much stronger complement activator. Treatment with complement transiently increased the ability of HTLV-I to infect CR+ cell lines as judged by provirus formation (4- to 8-fold increase) and p24 production (5- to 10-fold increase). In contrast, complement treatment did not increase infection of CR- cells. In conclusion this study shows that HTLV-I activates complement leading to increased binding to, and transiently increased infection of, CR+ cells. This complement-mediated increased binding of HTLV-I may dramatically affect viral trafficking and immunological reactivity of virus in vivo.
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PMID:HTLV-I activates complement leading to increased binding to complement receptor-positive cells. 855 9

ACH-2 cells, an immortalized human T-cell line, contain a single integrated copy of the HIV-1 provirus. Here, the structure of HIV-1 chromatin was probed using a DNA cleavage reagent. Nuclei were isolated from ACH-2 cells and treated with methidiumpropyl-EDTA (MPE)-iron(II) to produce limited DNA cleavage. Primers were selected at approximately 300 bp intervals along the HIV-1 DNA, and sites of preferential cleavage were mapped by carrying out 50 cycles of primer extension using a thermo-stable DNA polymerase in the presence of [32P]dATP. By comparing the resulting cleavage pattern with patterns derived from human cell lines not containing HIV-1 sequences, it was possible to map the arrangement of nucleosomes across the integrated HIV-1 genome. Particularly regular spacing was seen in the 3' end of the pol and env coding regions, and several extended blocks spared of nucleosomes were found in gag and pol, the largest being an approximately 450 bp region in gag. For comparison, and to examine nucleosome placement on HIV-1 DNA when it is not integrated, overlapping segments of HIV-1 DNA were cloned into an EBV-oriP plasmid, grown as stable episomes in a human B lymphoblastoid cell line, and the same analysis using MPE-iron(II) cleavage and primer extension carried out. The major features of nucleosome placement on these EBV/HIV minichromosomes was very similar to that observed in the integrated HIV-1 genome arguing for a strong sequence dependence for nucleosome placement along HIV-1 DNA.
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PMID:Nucleosomal arrangement of HIV-1 DNA: maps generated from an integrated genome and an EBV-based episomal model. 860 34

Recombinant glycoprotein 120 (rgp120) of human immunodeficiency virus type-1 (HIV-1) activates the human complement system in the absence of anti-gp120 antibodies. HIV-1 glycoprotein gp120 can dissociate from the viral envelope either spontaneously or after binding of HIV-1 to the CD4 molecule. As a consequence, gp120 can circulate in the patient's serum and attach to the surface of uninfected CD4+ T cells. Complement activation by cell-bound HIV-1 glycoprotein gp120 with subsequent opsonization may represent a mechanism for the elimination of uninfected CD4+ cells by the reticuloendothelial system, thereby enhancing the progression of HIV disease. In the current study, the complement proteins C4,C3,C5,C9, and properdin were found to bind to a synthetic peptide covering positions 233-251 of the gp120BRU sequence on incubation with normal human serum. Complement activation by the peptide was comparable with that induced by aggregated IgG, complete rgp120, and the previously described complement-activating gp41-peptide 609-623. Activation occurred via the classical pathway and was abrogated in the presence of EDTA, Mg2+/EGTA, or C4-deficient human serum. Peptides partly overlapping the sequence 233-251 activated complement to a lesser extent. The complement-activating capacity of the gp120 sequence 233-251 was not restricted to the HIV-1BRU isolate, because a peptide from the corresponding sequence of the HIV-1MN strain was also capable of activating complement. An additional strong complement-activating site was identified in the gp120 sequence 321-360 of the HIV-1MN strain. These data indicate that distinct sites in gp120 are able to activate human serum complement via the classical pathway in the absence of anti-gp120 and independent of glycosylation.
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PMID:Identification of complement activation sites in human immunodeficiency virus type-1 glycoprotein gp120. 863 Mar 95

The relationship between cytomegalovirus (CMV) DNA in leucocytes and CMV disease in AIDS patients was sought. In 195 HIV-1 infected, mostly AIDS patients, CMV nPCR was performed in 477 peripheral EDTA blood samples which were collected also for CD4 cell counts (403), classic (410) and rapid virus isolation (270), and antigenemia tests (190). Most patients who died were autopsied. Immunohistopathology for CMV was performed. The first 43 patients were classified clinically according to having (A) verified organ involvement of CMV (15), (B) suspected CMV disease due to symptoms (4), or no CMV-associated disease (24). CMV-DNA was detected in the majority of samples (66%) and patients (68%). In contrast, CMV in the samples was detected in only 16% by classical and 11% by rapid isolation and in 8.4% by the antigenemia test. Acquisition of CMV DNA in leucocytes became more common as the CD4 cell counts fell. Detection of CMV DNA was significantly associated with CMV-associated symptoms and later mortality. In conclusion, CMV PCR of DNA in leucocytes is a sensitive and early marker of CMV disease in HIV-infected AIDS patients. It might be a marker to be added to CD4 cell counts for initiation of preemptive therapy.
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PMID:CMV PCR in leucocytes as an early marker for the development of CMV disease in AIDS patients. 866 50

To investigate whether HTLV-I induces the development of complement-dependent cytotoxic antibodies in humans, sera of asymptomatic HTLV-I carriers and of patients suffering from tropical spastic paraparesis/HTLV-I-associated myelopathy (TSP/HAM) or adult T cell leukaemia (ATL) were used in a cytotoxicity assay against a panel of target cells. This panel included uninfected cell lines (CEM, Jurkat, Molt and H9), cell lines chronically infected with HTLV-I (MT2, MT4, C9IPL and HUT102), as well as lines H36 (H9 infected with HTLV-I), H9-IIIB (H9 infected with HIVms) and H9-MN (H9 infected with HIMVMN). HTLV-I+ sera induced lysis of H36 and of lines expressing HTLV-I antigens in the presence of rabbit complement, but did not lyse cells in presence of human complement. The HTLV-I+ sera also failed to lyse the HTLV-I- lines and H9 cells, suggesting that lysis was specific for HTLV-I. H36 cell lysis was prevented by IgG depletion of the sera and by dialysis of rabbit complement against EGTA or EDTA. Rabbit complement-dependent cytotoxic antibodies were present in the sera of 14/14 HTLV-I-infected individuals; the highest titres were predominantly found in the sera of the TSP/HAM patients. Such antibodies were also detected in 5/5 individuals coinfected with HIV-1 and HTLV-I, although no cytotoxic antibody could be found against HIV-infected cells. Vice versa, sera of HIV-1-infected individuals did not exert a lytic effect in the presence of complement (of human or rabbit origin) against HIV-1- or HTLV-I-infected cells. Incubation of the sera of four HTLV-I-infected patients with HTLV-I env-specific synthetic peptides demonstrated that some of the complement-dependent cytotoxic antibodies recognized epitopes located on gp46 between amino acids 190 and 209. There is no correlation of rabbit complement-dependent cytotoxic HTLV-I antibodies with the development of disease.
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PMID:Complement-dependent cytotoxic antibodies to human T lymphotropic virus type I (HTLV-I)-infected cells in the sera of HTLV-I-infected individuals. 869 33

The synthesis of a C-5 modified uridine phosphoramidite which contains a primary amino group protected with Fmoc is described. During cleavage and deprotection of chemically synthesized RNA, the Fmoc protecting group is removed to yield a free amino group at a predetermined position in the RNA sequence that can be covalently modified with any reporter group, small structural probes, and biological molecules. This modified uridine phosphoramidite was used to incorporate a reactive primary amino group at position 24 in the HIV-1 Tat binding site of a trans-activation responsive (TAR) RNA sequence during chemical syntheses. Modified RNA phosphoramidite was incorporated into RNA oligomers with more than 97% coupling efficiencies. RNA containing modified uridine was cleaved from the support, deprotected, and desalted according to standard procedures. After deprotection and gel purification, nuclease digestion and HPLC analysis were performed to confirm the incorporation of C-5-aminouridine into the RNA sequence. The effect of modified uridine on TAR RNA structure was analyzed by CD spectroscopy and protein binding assays. Site-specific incorporation of EDTA was accomplished by treating primary amine bearing TAR RNA with an isothiocyanato derivative of nitrobenzyl-EDTA.
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PMID:Incorporation of an artificial protease and nuclease at the HIV-1 Tat binding site of trans-activation responsive RNA. 881 49

Fragile X syndrome is a frequent cause of mental retardation resulting from the absence of FMRP, the protein encoded by the FMR1 gene. FMRP is an RNA-binding protein of unknown function which is associated with ribosomes. To gain insight into FMRP function, we performed immunolocalization analysis of FMRP truncation and fusion constructs which revealed a nuclear localization signal (NLS) in the amino terminus of FMRP as well as a nuclear export signal (NES) encoded by exon 14. A 17 amino acid peptide containing the FMRP NES, which closely resembles the NES motifs recently described for HIV-1 Rev and PKI, is sufficient to direct nuclear export of a microinjected protein conjugate. Sucrose gradient analysis shows that FMRP ribosome association is RNA-dependent and FMRP is found in ribonucleoprotein (RNP) particles following EDTA treatment. These data are consistent with nascent FMRP entering the nucleus to assemble into mRNP particles prior to export back into the cytoplasm and suggests that fragile X syndrome may result from altered translation of transcripts which normally bind to FMRP.
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PMID:The fragile X mental retardation protein is a ribonucleoprotein containing both nuclear localization and nuclear export signals. 884 25

The analytical variability of the new commercially available Reverse Transcriptase-Polymerase Chain Reaction (RT-PCR) assay, Amplicor HIV-1 Monitor, has been assessed to establish criteria for assessing the significance of HIV-1 RNA level measurements. Estimations of the standard deviations (SD) of log-copies in inter-assay (mean 0.09 log) and in inter-laboratory (mean 0.14 log) reproducibility experiments demonstrated that the assay can discriminate with 95% confidence between 3-fold (inter-assay) and 5-fold differences (inter-laboratory). The inter-lot reproducibility (mean 0.10 log) was similar to the inter-assay reproducibility. The HIV-1 RNA concentrations measured in plasma collected in potassium EDTA anticoagulant were slightly higher than those measured in plasma collected in sodium citrate. The HIV-1 RNA concentrations measured in sera were about 50% of the HIV-1 RNA concentrations measured in paired plasma samples. However, there was a strong correlation between these two measurements (P < 0.0001). The assay was used to measure viral RNA in the plasma of 50 HIV-1 positive individuals at different stages of infection. All the individuals had detectable HIV-1 RNA (300-957000 copies/ml). There was no correlation between HIV-1 RNA and Immune Complex Dissociated (ICD) p24 antigen, but HIV-1 RNA was correlated with CD4+ cell counts (P < 0.0001) and the clinical stage (P = 0.0042), with higher HIV-1 RNA concentrations in patients with a more advanced stage of the disease. The significant association of HIV-1 RNA with major markers of HIV infection and the reliability of this sensitive, easy-to-use RT-PCR assay indicate its suitability for use in clinical trials and suggest that this assay is appropriate for routine clinical applications.
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PMID:Assessment of a standardized reverse-transcriptase PCR assay for quantifying HIV-1 RNA in plasma and serum. 884 17


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