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Query: UMLS:C0021051 (immunodeficiency)
 71,517 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have characterized an inhibitory RNA element in the human immunodeficiency virus type 1 (HIV-1) gag coding sequence that prevents gag expression. The inhibition exerted by this element could be overcome by the presence of the Rev-responsive element in cis and of Rev protein in trans. To understand the mechanism of function, we inactivated the inhibitory element by mutagenesis while maintaining an intact gag coding region. A constitutive high level of Rev-independent gag expression was achieved only after the introduction of 28 point mutations over a large region of 270 nucleotides within the gag coding region. To our knowledge, this is the first demonstration of inactivation of a negative RNA element within a coding region without alteration of the expressed protein. Elimination of the inhibitory element in the p17gag region, named INS-1, offered the opportunity to detect a second inhibitory element in the gag-pol region. The presence of either INS element is sufficient to inhibit gag expression, demonstrating that multiple INS elements acting independently can inhibit HIV RNA expression. Expression of gag from Rous sarcoma virus, a retrovirus that does not require Rev-like regulatory proteins, revealed that the Rous sarcoma virus p19gag region does not contain inhibitory elements. These results demonstrate the presence of a strong inhibitory element acting at the level of mRNA and provide a general method for the removal of such elements from mRNA coding regions. The inhibitory element functions in the absence of any HIV-1 proteins, suggesting that cellular factors are responsible for this inhibition.
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PMID:Mutational inactivation of an inhibitory sequence in human immunodeficiency virus type 1 results in Rev-independent gag expression. 143 10

Expression of the human immunodeficiency virus (HIV) structural gene products is suppressed in the absence of the Rev protein. The block to expression reflects, in part, nuclear retention of those mRNAs which encode the structural proteins. The presence of intragenic cis-acting repressive sequences (CRS) and inefficient splicing of the primary viral transcript are thought to contribute to nuclear entrapment of viral RNA. To elucidate the mechanism for repression of HIV gene expression, the ability of a 270-bp segment of the pol gene shown previously to repress gene expression to interact with cellular factors was investigated. Incubation of RNA corresponding to the 270-bp CRS element with nuclear extract prepared from human T-cells revealed a strong and specific interaction with several cellular factors. Covalent cross-linking of the RNA-protein complex demonstrated the presence of at least three proteins, the predominant one having a molecular weight of approximately 42 kDa. A monoclonal antibody raised against hnRNP C, a component of the splicing machinery, recognized the CRS-protein complex, suggesting that hnRNP C or a closely related gene product interacts with CRS-containing RNA. Consistent with this conclusion, addition of RNA corresponding to a beta-globin intron sequence in the binding reaction completely blocked formation of the CRS-protein complex. These findings raise the possibility that the CRS elements elicit nuclear entrapment of viral RNA through formation of RNA-protein complexes that are not accessible to nuclear export pathways.
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PMID:Interaction of cellular factors with intragenic cis-acting repressive sequences within the HIV genome. 144 21

Individuals infected with human immunodeficiency virus type 1 (HIV-1) develop a humoral immune response to the virus's major structural gene products env, gag, and pol. The distribution of antibodies to env, gag, and pol proteins in Central African populations is of interest as they have a high level of immune system activation compared to non-African populations. Using the Western blot technique, we analyzed the isotypic distribution of anti-HIV antibodies in 45 HIV-1-infected individuals from Central Africa that were either symptomatic or asymptomatic. We observed two basic differences between the isotypic profile of individuals from Central Africa and non-African populations. Central African individuals had a strong polyisotypic response to gag and pol, which has only been observed for gag in American and European populations. In addition, individuals from Central Africa had a high frequency of IgG4 to gag and pol, 75 and 51%, respectively, as compared to 29 and 6% in a non-African population. The elevated IgG4 response may result from the high basal level of immune stimulation seen in Africans due to multiple and frequent exposures to viral, bacterial, and parasitic antigens.
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PMID:Isotypic distribution of HIV-1-specific antibodies in individuals from central Africa. 147 77

A simple method for the overproduction in Escherichia coli and purification of major core protein p24 of human immunodeficiency virus type 1 (HIV-1) was described. The gag-pol region encoding p24, p15, and protease was fused to 3' end of lacZ gene on plasmid. A LacZ-Gag fusion protein, the major primary product, is designed to be cleaved by the HIV-1 protease coexpressed through frameshifting. In fact, p24 and its immediate precursor, p25, were produced in the cells grown at 25C, but not at 37C. When the gag and pol frames were fused in-frame to express the protease without frameshifting, the main product, a LacZ-Gag-Pol fusion protein, was efficiently processed to give p24 exclusively both at 37C and 25C, suggesting more efficient expression of the protease. Recombinant p24 was purified to near homogeneity by a simple three-step procedure. The amino-terminal sequence of the recombinant p24 was the same as that of p24 deduced from nucleotide sequence, indicating that correct processing occurred in E. coli by the coexpressed protease. The method described here provides a means to obtain a large amount of highly pure p24, which is useful for crystallographic and functional studies, preparation of specific antibody, and diagnostic and prognostic uses.
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PMID:A simple method for overproduction and purification of p24 Gag protein of human immunodeficiency virus type 1. 147 33

The pol gene of all retroviruses is expressed as a gag-pol fusion protein which is proteolytically processed to produce all viral enzymes. In the human immunodeficiency virus (HIV), the gag and pol genes overlap by 241 nucleotides with pol in the -1 phase with respect to gag. The gag-pol fusion is produced via a -1 ribosomal frameshifting event that brings the overlapping, out-of-phase gag and pol genes into translational phase. Frameshifting occurs at a so called 'shift site' 8-10 nucleotides upstream of a hairpin loop which may play a role in the regulation of frameshifting. We have fused this region of HIV-1 to the 5' end of the firefly luciferase reporter gene in order to quantitatively measure ribosomal frameshifting both in cells and by in vitro translation. A series of 2'-O-methyl oligonucleotides was designed to specifically bind the sequences which flank the gag-pol hairpin. Ribosomal frameshifting is enhanced up to 6 fold by those oligonucleotides which bind the area just 3 to the stem. Oligonucleotides which bind 5' to the stem have no effect on frameshift efficiency. In addition, we have constructed a series of fusion genes which mimic the effect of the bound oligonucleotides with intramolecular hairpins. The results suggest that increasing RNA secondary structure downstream of the shift site increases the frequency of ribosomal frameshifting, and that this effect can be mimicked by antisense oligonucleotides.
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PMID:Enhancement of ribosomal frameshifting by oligonucleotides targeted to the HIV gag-pol region. 150 80

Synthetic peptide analog inhibitors of human immunodeficiency virus type 1 (HIV-1) protease were used to study the effects of inhibition of polyprotein processing on the assembly, structure, and infectivity of virions released from a T-cell line chronically infected with HIV-1. Inhibition of proteolytic processing of both Pr55gag and Pr160gag-pol was observed in purified virions from infected T cells after treatment. Protease inhibition was evident by the accumulation of precursors and processing intermediates of Pr55gag and by corresponding decreases in mature protein products. Electron microscopy revealed that the majority of the virion particles released from inhibitor-treated cells after a 24-h treatment had an immature or aberrant capsid morphology. This morphological change correlated with the inhibition of polyprotein processing and a loss of infectivity. The infectivity of virion particles purified from these chronically infected cell cultures was assessed following treatment with the inhibitor for 1 to 3 days. Virions purified from cultures treated with inhibitor for 1 or 2 days demonstrated a 95- to 100-fold reduction in virus titers, and treatment for 3 days resulted in complete loss of detectable infectivity. The fact that virions from treated cultures were unable to establish infection over the 7- to 10-day incubation period in the titration experiments strongly suggests that particles produced by inhibitor-treated cells were unable to reactivate to an infectious form when they were purified away from exogenous protease inhibitor. Thus, a block of HIV-1 protease processing of viral polyproteins by specific inhibitors results in a potent antiviral effect characterized by the production of noninfectious virions with altered protein structures and immature morphologies.
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PMID:Human immunodeficiency virus type 1 protease inhibitors irreversibly block infectivity of purified virions from chronically infected cells. 151 Apr 24

Patients infected with human immunodeficiency virus type 1 (HIV-1) develop a renal syndrome characterized by proteinuria, renal failure, and focal segmental glomerulosclerosis. By using a noninfectious HIV-1 DNA construct lacking the gag and pol genes, three transgenic mouse lines have been generated that develop a syndrome remarkably similar to the human disease. In the present study, we have characterized in detail one of these lines, Tg26. In Tg26 mice, proteinuria was detectable at approximately 24 days of age, followed by severe nephrotic syndrome and rapid progression to end-stage renal failure. Renal histology showed focal segmental glomerulosclerosis and microcystic tubular dilatation. Indirect immunofluorescence studies demonstrated increased accumulation of the basement membrane components laminin, collagen type IV, and heparan sulfate proteoglycan. The viral protein Rev was present in sclerotic glomeruli. Northern blot analysis of total renal RNA showed expression of viral genes prior to the appearance of histologic renal disease, with greatly diminished viral gene expression late in the disease course. Kidneys from transgenic mice expressed increased steady-state levels of collagen alpha 1(IV) mRNA when glomerulosclerosis was present. We conclude that the presence of HIV-1 genes is associated with progressive renal dysfunction and glomerulosclerosis in transgenic mice.
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PMID:Progressive glomerulosclerosis and enhanced renal accumulation of basement membrane components in mice transgenic for human immunodeficiency virus type 1 genes. 154 49

The human retroviruses human immunodeficiency virus type 1 (HIV-1) and human T-cell leukemia virus type I (HTLV-I) are characterized by complex regulation of gene expression. Each virus encodes a posttranscriptional regulator, the 19-kDa HIV-1 Rev protein and the 27-kDa HTLV-I Rex protein, which is required for viral replication. Expression of these trans activators results in the cytoplasmic accumulation of unspliced or singly spliced viral mRNA which encode the gag, pol, and env gene products. The finding that the HTLV-I Rex protein is able to functionally substitute for the Rev protein of HIV-1 indicates that HIV-1 Rev and HTLV-I Rex may interact with the same component of a cellular pathway involved in either mRNA splicing or transport. In this study, we have generated functional Rev/Rex hybrid proteins by domain exchange. We have defined, using in vivo and in vitro analyses, the activation domains of Rev and Rex which are the putative targets of a common host cell factor(s) required for Rev and Rex function.
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PMID:Definition of the human immunodeficiency virus type 1 Rev and human T-cell leukemia virus type I Rex protein activation domain by functional exchange. 154 84

A new, modular Western blot (immunoblot) system for human immunodeficiency virus (HIV) antibodies (ABN WesPage; Wellcome) was compared with enzyme immunoassays (Wellcome, Behringwerke, and Abbott) and with a U.S. Food and Drug Administration (FDA)-licensed Western blot (DuPont) in a multicenter study. A total of 649 serum samples from HIV patients at different stages of the disease, as well as from high-risk patients, from patients with conditions unrelated to AIDS, and from healthy blood donors, were used in the evaluation along with nine seroconversion panels. For evaluation of Western blot reactivity, both Centers for Disease Control (CDC) and FDA criteria were used. With the DuPont Western blot as the reference assay, the overall sensitivity and specificity of the ABN WesPage were 100 and 99.1%, respectively, when indeterminate results were not taken into account and when both tests were interpreted in accordance with CDC criteria. The DuPont Western blot detected significantly more antibodies to pol and gag gene products than the ABN WesPage. The ABN WesPage showed a higher positive rate of detection of viral envelope band gp160. When both Western blots were interpreted in accordance with CDC criteria, the ABN WesPage and the DuPont Western blot yielded 9.3 and 10.4% indeterminate results, respectively. When the DuPont Western blot was interpreted in accordance with the manufacturer's instructions (FDA criteria), 25.7% of the samples tested were regarded as indeterminate. The choice of interpretation criteria is of paramount importance for the evaluation of HIV Western blot patterns.
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PMID:Multicenter evaluation of the novel ABN Western blot (immunoblot) system in comparison with an enzyme-linked immunosorbent assay and a different Western blot. 155 87

Cells infected with a recombinant vaccinia virus carrying the gag and pol regions of the human immunodeficiency virus type 1 genome (Vac-gag/pol) released human immunodeficiency virus (HIV)-like particles containing HIV-specific RNA. However, cells infected with another recombinant vaccinia, Vac-gag/pol-dP, derived through the deletion of an 85-base region (nucleotide positions 679-763) of the HIV genome between the primer binding site and the gag initiation codon of Vac-gag/pol, produced HIV-like particles devoid of the HIV-specific RNA. This 85-base deletion was suggested to cause the collapse of a stable stem-loop structure of 46 bases (751-796) around the gag initiation codon. To examine the role of the stem-loop structure in the packaging of RNAs, we constructed a vaccinia vector plasmid that carried this 46-base sequence followed by the Sendai virus nucleocapsid (NP) gene. When both Vac-gag/pol-dP and this plasmid were introduced into cells, HIV-like particles released from the cells contained the NP gene RNA. However, another vaccinia vector plasmid, which carried the 46-base sequence in the midst of the NP gene, could not supply RNA for incorporation into HIV-like particles. Computer analysis of this plasmid sequence suggested that the 46-base sequence cannot form the stem-loop structure. These findings suggest that the stem-loop structure formed by the 46-base sequence is crucial as a packaging signal.
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PMID:RNA packaging signal of human immunodeficiency virus type 1. 158 35


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