SEARCH · PubMed Health

Results for “Long terminal repeats”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 955 records · Page 53Linked to original sources

PubMed1992

Characterization of a novel T lymphocyte protein which binds to a site related to steroid/thyroid hormone receptor response elements in the negative regulatory sequence of the human immunodeficiency virus long terminal repeat.

We have previously identified a T lymphocyte protein which binds to a site within the LTR of the human immunodeficiency virus type 1 (HIV-1) and exerts an inhibitory effect on virus gene expression. The palindromic site (site B) recognized by this protein is related to the palindromic binding sites of members of the steroid/thyroid hormone receptor family. Here we characterize the T cell protein binding to this site as a 100 kD protein which is most abundant in T cells and which binds to site B as a 200 kD complex. This protein is distinct from other members of the steroid/thyroid hormone receptor family including the COUP protein which has a closely related DNA binding specificity.

Base Sequence↗

PubMed1993

Blockade of human immunodeficiency virus type 1 production in CD4+ T cells by an intracellular CD4 expressed under control of the viral long terminal repeat.

A retroviral vector was constructed in which a gene encoding a mutated soluble CD4 protein that is retained in the endoplasmic reticulum (sCD4-KDEL) is expressed under control of human immunodeficiency virus type 1 (HIV-1) regulatory elements. HIV-1 infection of a human T-cell line transduced with this vector led to induction of sCD4-KDEL synthesis and a block in transport of the HIV envelope protein to the cell surface. There was a complete block to maturation of infectious HIV-1 in the transduced cells, no viral spread, and little or no syncytium formation. Infected cells gradually disappeared from the culture over a period of 2 months. This intracellular trap for HIV has potential application in gene therapy for AIDS.

Acquired Immunodeficiency Syndrome↗

PubMed1997

RNA-targeted activators, but not DNA-targeted activators, repress the synthesis of short transcripts at the human immunodeficiency virus type 1 long terminal repeat.

The human immunodeficiency virus type 1 (HIV-1) promoter directs the synthesis of two types of RNA molecules: full-length transcripts, whose synthesis is activated by the viral activator Tat, and short transcripts, whose synthesis is dependent on the inducer of short transcripts (IST), a bipartite DNA element located in large part downstream of the HIV-1 transcriptional start site. In the absence of Tat, short transcripts constitute the large majority of the RNA molecules synthesized from the HIV-1 promoter. In the presence of Tat, synthesis of the short transcripts is repressed and synthesis of the full-length transcripts is activated. Tat is unique among transcriptional activators in acting through an RNA target, the TAR element. However, Tat has been shown to activate transcription from a DNA target when fused to the appropriate DNA binding domain, raising the question of why Tat has been directed to the RNA. Here we have compared the abilities of Tat and other RNA- and DNA-bound activators to stimulate transcription from the HIV-1 promoter. We show that DNA-targeted activators, including DNA-targeted Tat, activate the synthesis of both short and long transcripts, while RNA-targeted Tat and another RNA-targeted activator activate the synthesis of full-length transcripts but specifically repress that of short transcripts. The unique ability of RNA-targeted activators to down-regulate short transcript synthesis suggests that Tat is directed to the RNA specifically for the purpose of repressing short transcripts.

Base Sequence↗

PubMed1995

X irradiation-induced transcription from the HIV type 1 long terminal repeat.

It has been previously shown in vitro and in vivo that the human immunodeficiency virus type 1 can be dramatically enhanced by certain heterologous viral, chemical, and physical (ultraviolet irradiation) agents. A common denominator shared by these agents is their ability to cause stress responses in cells. To analyze if a similar effect could occur by X irradiations, we tested the in vitro effect of X rays on HIV LTR-directed gene expression. The results demonstrate that the HIV-1 LTR is activated by X irradiation in a dose- and time-dependent manner, in all cell types tested, including epitheloid, fibroblast, and lymphoid cell lines. This study raises the possibility that exposure of AIDS patients to ionizing radiation (e.g., during treatment of epidemic Kaposi's sarcoma) could play a role in the activation of HIV-1 in vivo.

Cell Line↗

PubMed1990

The HIV-1 long terminal repeat contains an unusual element that induces the synthesis of short RNAs from various mRNA and snRNA promoters.

We describe an unusual element that activates the synthesis of short transcripts from a wide variety of mRNA and small nuclear RNA (snRNA) promoters, including the U6 RNA polymerase III promoter. This inducer of short transcripts (IST) is located between positions -5 and +82 relative to the cap site in the HIV-1 LTR. In the presence of IST, the total transcriptional activity of the different promoters is greatly increased, but the resulting additional RNA molecules are short, ending around position +60. IST is not the RNA target (TAR) for Tat trans-activation; however, because it relies entirely on cellular factors for activity, IST may serve to provide abundant RNA targets for Tat trans-activation without a requirement for full-length viral mRNA expression.

Base Sequence↗

PubMedDate not supplied

DNA elements target of transcriptional factors are not restricted to long terminal repeat of human immunodeficiency virus.

In this report we show that signals for transcriptional factors are not restricted to the HIV-1 LTR, but are present throughout the HIV-1 genome. Furthermore, we identified a sequence, AGAACAGATG, highly homologous to the X-box of class II MHC genes and located within the tat-IVS/env region of HIV-1. Double stranded oligonucleotides mimicking the HIV-1 region containing AGAACAGATG were synthesized and band shift experiments were performed demonstrating that this HIV-1 genomic region binds nuclear proteins. We further demonstrate that the binding of nuclear factors to this tat-IVS/env HIV-1 sequence is competed for, in the band-shift assay, by the highly homologous X-box of the promoter of the human HLA-DR alpha gene. The presence in the HIV-1 genome of DNA sequences homologous or identical to regulatory sequences of cellular genes represents a potential mechanism of predation of DNA elements recognized by DNA binding proteins.

Base Sequence↗

PubMed1995

Revertants and pseudo-revertants of human immunodeficiency virus type 1 viruses mutated in the long terminal repeat promoter region.

The TAR domain is an RNA secondary structure element within the leader transcript of the human immunodeficiency virus type 1 (HIV-1) virus. TAR RNA forms the binding site for the viral trans-activator protein Tat and cellular co-factors that are involved in induction of the LTR transcriptional promoter. Here, we report that mutations in the single-stranded bulge- and loop-domains of TAR RNA impair the ability of the virus to replicate in T cell lines. Revertant viruses were isolated upon prolonged culturing and analysed through sequencing. The reversion data confirm the importance of both bulge and loop as sequence-specific recognition motifs. We also analysed the replication phenotype of a mutant HIV-1 virus with a substitution in the -19/-3 promoter region. This mutant displayed delayed infection kinetics compared to the wild-type virus, and revertants with increased replication potential could be isolated. Interestingly, all revertants had acquired an additional mutation at position -2. Primer extension analyses revealed that an upstream shift in transcription start site usage was induced by the -19/-3 substitution. This effect was compensated for by the nucleotide substitution near the RNA start site.

Animals↗

PubMed1992

Sequences within the R region of the long terminal repeat activate basal transcription from the HIV-1 promoter.

The importance of the R region in basal human immunodeficiency virus type 1 (HIV-1) transcription was addressed by comparing a panel of HIV-1 R region mutants using in vitro and in vivo assays. Using deletion, base substitution mutants, and compensatory mutants, the precise R region sequences essential for basal HIV-1 promoter activity in vitro were mapped to sequences between +17 to +21. Within this regulatory domain, nucleotides +19 and +21 appear to be critical. The effect of these mutations on steady state RNA levels in transfected cells has been analyzed by S1 nuclease protection assay using uniformly labeled probes. Two main conclusions may be drawn from these studies. First, HIV-1 basal transcription is abundant, with the majority of correctly initiated transcripts truncated between sequences +57 to +70. Second, analysis of the compensatory mutants indicates the secondary structure of the nascent R region RNA is not an obligate requirement for the production of the truncated transcripts. Mutations in R region primary sequence that selectively abolish the production of the truncated transcripts in vivo also exhibit reduced promoter activity in vitro. The appearance of high levels of truncated transcripts raise the interesting possibility that-similar to c-myc, c-myb, and c-fos--basal HIV-1 expression is regulated by transcription elongation.

Gene Expression Regulation, Viral↗

PubMed1997

The effects of vinblastine on the expression of human immunodeficiency virus type 1 long terminal repeat.

Previous work by our group has demonstrated induction of the HIV-LTR following exposure of cells to various DNA-damaging agents such as ultraviolet (UV) light, cisplatin, and doxorubicin. The current experiments were designed to determine the relative effects of the anti-mitotic drug vinblastine on expression of the HIV-LTR. Using human cervical carcinoma (HeLa) cells stably transfected with the chloramphenicol acetyl transferase (CAT) reporter transcriptionally driven by the HIV-LTR promoter, we demonstrated a 9-10-fold induction at 48-72 h following vinblastine treatment. Previous experiments had demonstrated repression of cisplatin or doxorubicin-mediated HIV induction by treatment with salicylic acid. The vinblastine induction also was repressed by salicylic acid treatment, but not by treatment with indomethacin, suggesting a role for the NF kappa B pathway in the inductive response. When UV exposure was coupled to the vinblastine treatment, there was no additive or synergistic effect evident, suggesting similar paths of induction between the two agents. Northern blots demonstrated that these agents were operating at the level of transcription and that salicylic acid inhibited vinblastine-mediated induction of HIV-LTR-CAT mRNA only if administered at the same time as vinblastine; addition of salicylic acid 2 h later had no effect on transcript accumulation. All combinations of treatments with vinblastine and/or salicylic acid markedly reduced cell survival.

Cell Survival↗

PubMedDate not supplied

Curcumin and curcumin derivatives inhibit Tat-mediated transactivation of type 1 human immunodeficiency virus long terminal repeat.

The transcription of HIV1 provirus is regulated by both cellular and viral factors. Various evidence suggests that Tat protein secreted by HIV1-infected cells may have additional action in the pathogenesis of AIDS because of its ability to also be taken up by non-infected cells. Curcumin [diferuloylmethane or 1,7-bis-(4-hydroxy-3-methoxyphenyl)-1,6-heptadiene-3,5-dione] is the yellow pigment in turmeric Curcuma longa (Linn). It exhibits a variety of pharmacological effects including antiinflammatory and antiretroviral activities. Here, we demonstrated that curcumin used at 10 to 100 nM inhibited Tat transactivation of HIV1-LTR lacZ by 70 to 80% in HeLa cells. In order to develop more efficient curcumin derivatives, we synthesized and tested in the same experimental system the inhibitory activity of reduced curcumin (C1), which lacks the spatial structure of curcumin; allyl-curcumin (C2), which possesses a condensed allyl derivative on curcumin that plays the role of metal chelator; and tocopheryl-curcumin (C3), which enhances the antioxidant activity of the molecule. Results obtained with C1, C2 and C3 curcumin derivatives showed a significant inhibition (70 to 85%) of Tat transactivation. Despite the fact that tocopheryl-curcumin (C3) failed to scavenge O2.-, this curcumin derivative exhibited the most activity; 70% inhibition was obtained at 1 nM, while only 35% inhibition was obtained with the curcumin.

Curcumin↗

PubMed1995

Suppression of activation of the human immunodeficiency virus long terminal repeat by CD8+ T cells is not lentivirus specific.

CD8+ T lymphocytes of HIV-1-infected individuals can efficiently suppress HIV-1 replication in CD4+ T lymphocytes. To elucidate the molecular events underlying this suppression, we have used the HIV-1 LTR directing the chloramphenicol acetyltransferase gene (CAT) in transient transfection assays using human Jurkat T cells. In addition to supernatants of patient CD8+ T lymphocytes (CD4+ > 350/microliters), supernatant of a T cell clone derived by Herpesvirus saimiri (HVS)-mediated transformation of CD8+ T lymphocytes of a patient demonstrating inhibition of virus replication were examined. Similar levels of inhibition of LTR-mediated gene expression in response to Tat or mitogenic activation with phorbol ester and calcium ionophore were observed by supernatants of both sources. The inhibitory effect of CD8+ T lymphocytes was not exclusive to lentiviral LTRs since transcription of both the HTLV-I LTR and RSV LTR in response to mitogen was effectively inhibited. In examination of the influence of CD8+ T cell-derived supernatant on NF kappa B-mediated activation, a dimer of the HIV-1 NF kappa B elements directing CAT was markedly inhibited by supernatants of both patient CD8+ lymphocytes and the HVS-derived CD8+ clone. Thus the inhibitory nature of CD8+ T lymphocytes appears not to be specific to lentiviral promoters and may mediate an inhibitory effect via the NF kappa B element.

CD8-Positive T-Lymphocytes↗

PubMed1993

ETS family proteins activate transcription from HIV-1 long terminal repeat.

ets is a multigene family and its members share a common ETS DNA-binding domain. ETS proteins activate transcription via binding to a purine-rich GGAA core sequence located in promoters/enhancers of various genes, including several that are transcriptionally active in T cells. The ETS1, ETS2, and ERBG/Hu-FLI-1 gene expression pattern also suggests a role for these genes in cells of hematopoietic lineage. The HIV-1 LTR core enhancer contains two 10-base pair direct repeat sequences (left and right) that are required for regulation of HIV-1 mRNA expression by host transcription factors, including NF kappa B. Two ETS-binding sites are present in the core enhancer of all the HIV-1 isolates reported so far. In our studies, we utilized HIV-1 HXB2 and HIV-1 Z2Z6 core enhancers because the Z2Z6 strain has a single point mutation flanking the right ETS-binding site. We demonstrate that the ETS1, ETS2, and ERGB/Hu-FLI-1 proteins can trans-activate transcription from both the HXB2 and Z2Z6 core enhancer when linked to a reporter (cat) gene. In addition, we show that the DNA binding and trans-activation with the Z2Z6 core enhancer is at least 40-fold higher than that observed with the HXB2 core enhancer. Further, we provide evidence that the marked increase in binding and trans-activation with Z2Z6 core enhancer sequences is due to the substitution of a flanking T residue in HXB2 TGGAA) by a C residue in Z2Z6 (CGGAA) isolate, thus generating an optimal ETS-binding core (CGGAA) sequence.

Animals↗

PubMed1994

Activation of HIV type 1 long terminal repeat by ultraviolet light is serum and strain specific.

We have studied the UV responsiveness of xeroderma pigmentosum (XP) and HeLa cell lines transfected with a CAT reporter gene under the control of the HIV-1 LTR promoter. XP fibroblasts grown in 10% newborn bovine serum (NBS) were three times more responsive to UV radiation than cells grown in 10% fetal calf serum (FCS). Moreover, cocultivation of UV-irradiated XP cells with XP cells containing stable integrants of HIV-LTR CAT was found to be more than four times more effective in inducing the CAT activity when cells were maintained in 10% NBS than in 10% FCS. The level of induction was also dependent on the serum concentration. These data indicate that a serum component, possibly a cytokine(s), can enhance the UV response of both irradiated cells and unirradiated cells cocultivated with irradiated cells. The magnitude of UV responsiveness seemed also to be strain dependent. CAT activity for the HIV LTR promoter from the HTLV-IIIB (HIV-IIIB) strain was induced more than 30-fold by UV irradiation whereas activity from the LAV-1BRU strain was less than 2-fold. In contrast, both constructs were strongly induced by Tat expression. This indicates that there are differences in the induction mechanism for these two stimuli, even though UV radiation has been previously reported to induce a cellular Tat-like factor (Valerie K, et al., Nature [London] 1988;333:78-81).

Base Sequence↗

PubMed1996

Low temperature enhancement of reporter genes expression directed by human immunodeficiency virus type 1 long terminal repeat.

Bacteria and eukaryotic cells respond to cold stress by inducing and enhancing the synthesis of specific arrays of proteins. We describe here cold-induced enhancement of expression for two reporter genes; luciferase and beta-galactosidase, both under the control of HIV-1 LTR sequences, observed in mouse fibroblasts and human HeLa cells respectively. Increased expression of luciferase in fibroblasts when shifted to 25 degrees C was detectable at 30 degrees C but was not observed following cold shock at 4 degrees C. To sustain the cold-induced effect, cells had to be kept at subphysiological temperature. The observed enhancement of luciferase activity did not result from a particular site of integration of the reporter gene and was evident whether cold-stressed cells were stationary or growing. Cold-induced expression of luciferase was evidenced at the protein level, enzymatic activity and RNA level, furthermore, active transcription and translation were required for overexpression. The cold effect which has been generalized with the reporter gene beta-galactosidase appears to be a process involving, at least in part, the HIV-1 LTR sequences and might correspond to an increase in the half-life of mRNA. The cold-dependent enhanced expression of luciferase and beta-galactosidase reported here, together with data describing the activation of HIV-1 LTR by hyperthermia, point out the particular temperature sensitivity of these regulatory sequences. This potential thermal modulation may be useful in the comprehension of regulatory processes in latency and reactivation of viral expression during HIV-1 infection.

3T3 Cells↗

PubMed1993

CD8+ T lymphocyte-mediated inhibition of HIV-1 long terminal repeat transcription: a novel antiviral mechanism.

HIV-1 infection evokes a vigorous antiviral response that may participate in resolving the initial peak of plasma viremia and maintenance of the asymptomatic state. CD8+ T lymphocytes of HIV-1-infected individuals play a critical role in the cellular anti-HIV response. In agreement with previous reports, we observed a potent suppressive effect on HIV-1 production from autologous CD4+ T lymphocytes by CD8+ T lymphocytes from asymptomatic HIV-1-infected individuals. To elucidate the mechanism(s) of the nonlytic suppressive antiviral activity, we examined the effect of CD8+ T lymphocytes on the transcriptional activity of the HIV-1 promoter (HIV-LTR). CD8+ lymphocytes from HIV-1-infected asymptomatic individuals suppressed tat-mediated HIV-LTR transcription in CD4+ lymphocytes. HIV-LTR transcriptional activity was suppressed by CD8 lymphocytes to an extent similar to tat-mediated transcription whereas CMV immediate early gene promoter activity was not affected. In contrast to the suppressive effect seen with CD8+ lymphocytes from HIV-1-infected individuals, CD8+ lymphocytes from uninfected individuals did not significantly inhibit tat-mediated or HIV-LTR transcription. The transcriptional inhibitory activity was not MHC class I restricted and could be mediated by a soluble factor(s). Supernatants from some CD8+ T lymphocyte cultures from HIV-1+ individuals exerted an inhibitory effect on tat-mediated HIV-LTR transcription comparable to that seen with CD8+ cells. In conclusion, CD8+ lymphocytes from asymptomatic HIV-1+ individuals could suppress virus production by inhibiting HIV-1 gene expression.(ABSTRACT TRUNCATED AT 250 WORDS)

CD8 Antigens↗

PubMed1998

Mycobacterium tuberculosis mannose-capped lipoarabinomannan can induce NF-kappaB-dependent activation of human immunodeficiency virus type 1 long terminal repeat in T cells.

Tuberculosis has emerged as an epidemic, extended by the large number of individuals infected with human immunodeficiency virus type 1 (HIV-1). The major goal of this study was to determine whether the mycobacterial cell wall component mannose-capped lipoarabinomannan (ManLAM) of Mycobacterium tuberculosis (M. tuberculosis) could activate transcription of HIV-1 in T cells with the use of an in vitro cell culture system. These experiments are of prime importance considering that CD4-expressing T lymphocytes represent the major virus reservoir in the peripheral blood of infected individuals. Using the 1G5 cell line harbouring the luciferase reporter gene under the control of the HIV-1 LTR, it was first found that culture protein filtrates (CFP) from M. tuberculosis or purified ManLAM could activate HIV-1 LTR-dependent gene expression unlike similarly prepared CFP extracts devoid of ManLAM. The implication of protein tyrosine kinase(s), protein kinase A and/or protein kinase C was highlighted by the abrogation of the ManLAM-mediated activation of HIV-1 LTR-driven gene expression using herbimycin A and H7. It was also determined, using electrophoresis mobility shift assays, that M. tuberculosis ManLAM led to the nuclear translocation of the transcription factor NF-kappaB. M. tuberculosis ManLAM resulted in clear induction of the luciferase gene placed under the control of the wild-type, but not the kappaB-mutated, HIV-1 LTR region. Finally, the ManLAM-mediated activation of HIV-1 LTR transcription was found to be independent of the autocrine or paracrine action of endogenous TNF-alpha. The results suggest that M. tuberculosis can upregulate HIV-1 expression in T cells and could thus have the potential to influence the pathogenesis of HIV-1 infection.

Binding Sites↗

PubMed1997

The kappaB sites in the human immunodeficiency virus type 1 long terminal repeat enhance virus replication yet are not absolutely required for viral growth.

The dependence of human immunodeficiency virus type 1 (HIV-1) on its NF-kappaB binding sites (kappaB sites) for replication in transformed and primary T-cell targets was examined by infecting cells with HIV-1 reporter viruses containing kappaB site enhancer mutations. Viral transcription was measured either with luciferase-expressing HIV-1 that infects for a single round or by flow cytometric analyses with HIV-1 expressing placental alkaline phosphatase (PLAP) or green-fluorescent protein (GFP). Both PLAP- and GFP-expressing viruses spread from cell to cell and allowed analysis of viral gene expression patterns in single cells. Infection of a panel of T-cell lines with different basal levels of NF-kappaB demonstrated a direct correlation between the amount of constitutive nuclear NF-kappaB and the degree to which a wild-type virus outperformed kappaB site mutants. One T-cell line with a constitutively high level of nuclear NF-kappaB, PM1, showed a 20-fold decrease in transcription when its kappaB sites were mutated. In contrast, in a T-cell line with a low basal level of NF-kappaB, SupT1, mutation of the kappaB site in the enhancer had no effect on viral transcription or growth rate. Phytohemagglutinin-activated peripheral blood mononuclear cells showed a large dependence on the kappaB sites for optimal virus growth. Viruses without marker genes corroborated the finding that mutations to the kappaB sites impair virus production in cells with a high basal level of NF-kappaB. These data show that in T cells, HIV-1 can use NF-kappaB to enhance its growth but the virus is clearly able to grow in its absence.

Base Sequence↗

PubMed1995

Replication of HIV type 1 in rabbit cell lines is not limited by deficiencies in tat, rev, or long terminal repeat function.

HIV-1 infection has been documented in rabbits, but infection proceeds slowly in this species. Human and rabbit cell lines were compared in order to identify barriers to efficient HIV-1 infection of rabbit cells. A direct comparison of human and rabbit CD4 as receptor for HIV-1 indicated that the rabbit CD4 homolog did not function well even when expressed by human cells. Examination of viral RNA production indicated that the major HIV transcripts were produced in HIV-infected rabbit cells, but were present at levels significantly lower than those found for human cells. Ability of HIV-1 LTRs to direct protein expression in human and rabbit cells was compared using gene constructs with the chloramphenicol acetyltransferase (cat) gene flanked by HIV-1 LTRs. Chloramphenicol acetyltransferase protein expression was equivalent in rabbit and human cell lines transfected with the HIV-1/CAT constructs and cotransfections with the HIV-1 tat gene led to similar increases in CAT expression. Subsequent transfections with an infectious molecular HIV clone yielded approximately equal levels of HIV protein expression in rabbit and human cell lines, suggesting that major barriers to virus production in rabbit lines exist at steps prior to transcription of the viral genome. Because HTLV-I replicates with high efficiency in rabbit cells, a chimeric virus clone was constructed consisting of the 5' portion of HIV-1 through the nef coding sequence followed by the 3' HTLV-I LTR. Transfection of most rabbit cell lines with the chimera produced levels of p24gag protein higher than those transfected with the parent HIV-1 clone. By contrast, the unmodified HIV clone replicated more efficiently in all human cell lines tested.

Animals↗