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At least 181 records · Page 10Linked to original sources
PubMed1997

Regulation of gene expression directed by the long terminal repeats of feline leukemia viruses.

Repetitive structure of enhancer elements in the long terminal repeat (LTR) has been identified in feline leukemia viruses (FeLVs) integrated in lymphoid tumor cells in cats. In this study, promoter activities of the FeLV LTRs were measured in lymphoid and non-lymphoid cell lines in transient expression assays using plasmids containing the viral LTRs linked to the chloramphenicol acetyltransferase (CAT) gene. Promoter activity of the LTR with 3 enhancer repeats (pFTLTR) was significantly higher than that of the LTR with 1 enhancer (Glasgow-1 LTR) in feline (FT-1) and human (Jurkat) T-lymphoblastoid cell lines. Promoter activity of the pFTLTR was also significantly higher than that of its mutant (pFTLTR1E) in which 2 of the 3 enhancers were deleted in FT-1 and Jurkat cells. Both of these differences were not observed in a feline fibroblastic cell line (CrFK). Moreover, mutations affecting the consensus motifs for LVb, SV40, NF-1, GRE and FLV-1 resulted in decreased basal activity of the FeLV LTR (pFTLTR1E) in FT-1, Jurkat and CRFK cells. The decrease of the promoter activity was especially remarkable in FT-1 cells. The present study revealed the strong promoter activity of the FeLV LTR with 3 enhancer repeats and its modular enhancer elements positively regulating the transcription in a relatively tissue-specific manner.

Animals↗
PubMed1992

Hammerhead ribozyme-mediated cleavage of the long terminal repeat RNA of human immunodeficiency virus type 1.

Three ribozymes targeted against different sites of the long terminal repeat RNA (LTR RNA) of human immunodeficiency virus type 1 cleaved a LTR RNA transcript 1,000 x less efficiently than corresponding short synthetic oligoribonucleotide substrates. Varying the stem lengths of the ribozyme resulted in changes of the catalytic efficiency. Almost no cleavage was observed for a ribozyme forming only 10 base pairs instead of 14 with the LTR RNA. Increasing the base pairs to 16 or elongation of the stem formed within the ribozyme revealed only small changes in kcat/Km. The influence of chemical modifications within the ribozyme on the cleavage of the LTR RNA was also examined. 2'-Fluorocytidine substitutions as well as four terminal phosphorothioate internucleotidic linkages influenced the catalytic efficiency of ribozymes only negligibly. However, substitution of uridine by 2'-fluorouridine resulted in a 5-fold decrease of kcat/Km. A ribozyme containing all these modifications revealed only a 7-fold lower catalytic efficiency but a markedly increased stability in cell culture supernatant. These results demonstrate that it is possible to increase the stability of ribozymes toward nucleases without a serious loss in catalytic efficiency.

Base Sequence↗
PubMed1993

Mitomycin C stimulates the expression of human immunodeficiency virus long terminal repeat sequences in rat and human fibroblasts.

We have employed a recombinant plasmid, pBHIV1, carrying the long terminal repeat (LTR) sequences of the human immunodeficiency virus-1 (HIV-1) linked to the reporter chloramphenicol acetyl transferase (CAT) gene and to the aminoglycoside phosphotransferase (aph) gene as a selectable marker. We have introduced pBHIV1 into rat 208F and human MRCSV40TGR fibroblasts and obtained stable geneticin resistant RFBHIV1-1 and SVTGHIV1-1 cells, respectively. Both transfectant cells express CAT activity from the HIV LTR promoter. The response to the antineoplastic drug mitomycin C was studied on the LTR regulated CAT activity in both cell lines. It was found that mitomycin C at 10 micrograms/mL concentration stimulates the expression of CAT from the HIV LTR 77-fold in rat RFBHIV1-1 and 3.1-fold in human SVTGHIV1-1 cells.

Animals↗
PubMed1984

Molecular analysis of the envelope gene and long terminal repeat of Friend mink cell focus-inducing virus: implications for the functions of these sequences.

We sequenced the envelope (env) gene and 3' long terminal repeat of a Friend mink cell focus-inducing virus (F-MCFV). We also sequenced the gp70 coding regions for two cDNA clones of another F-MCFV. The deduced amino acid sequence of the env gene products of both F-MCFVs were compared to the corresponding sequences of other MCFVs and of ecotropic viruses. The env polypeptides of the different viruses showed long stretches of homology in the carboxy-terminal half of gp70 and in p15env ("constant region"). The amino-terminal half of gp70 was very similar in all MCFVs, but showed extensive variations relative to the ecotropic viruses ("differential region"). This differential region in all MCFVs is of endogeneous origin. We show evidence that this region carries determinants for ecotropic or polytropic host range. No indication could be found that the env gene products determine the histological type of disease caused by particular MCFVs. When the long terminal repeats of F-MCFV and Friend murine leukemia virus were compared with those of other viruses causing either lymphatic leukemia or erythroleukemia, several nucleotides were localized which might determine the histological type of disease caused by these viruses.

Amino Acid Sequence↗
PubMed1993

Differential interaction of cisplatin with the HIV-1 long terminal repeat in a resistant ovarian carcinoma cell line.

We constructed a recombinant plasmid, pBHIV1, carrying the long terminal repeat (LTR) sequences of HIV-1 linked to the reporter chloramphenicol acetyl transferase (CAT) gene and to the aminoglycoside phosphotransferase (aph) gene as a selectable marker. We have introduced pBHIV1 in a human ovarian cancer cell line A2780 and in a cisplatin resistant variant 2780CP, and obtained stable geneticin resistant A27HIV1-1 and 27CPHIV1-1 cells, respectively. Both transfectant cells expressed CAT activity from the HIV LTR promoter. The response to the anti-neoplastic drug cisplatin was studied on the LTR regulated CAT activity in both cell lines. It was found that cisplatin at 2.5 x 10(-5) M concentration stimulates the expression of CAT by 26-fold from the HIV LTR in A27HIV1-1, but requires a concentration of 5 x 10(-5) M to enhance expression by 4.1-fold in the cisplatin resistant 27CPHIV1-1 cells. Carboplatin, over a range of concentrations (1 x 10(-6) to 1 x 10(-4) M), does not stimulate expression of CAT from the HIV-1 LTR in either of the transfected cells.

Carboplatin↗
PubMed1994

The kinetics of HIV-1 long terminal repeat transcriptional activation resemble those of hsp70 promoter in heat-shock treated HeLa cells.

The long terminal repeat (LTR) of human immunodeficiency virus type 1 (HIV-1) is activated under different conditions including heat shock. By using transient transfection assays, we have compared the thermal activation of HIV-1 LTR to that of the promoter of the gene encoding the human stress protein hsp70 which is under the control of the heat shock transcription factor HSF. In these assays, the chloramphenicol acetyl transferase (Cat) gene was used as a reporter gene. Several parameters of the heat stress were analyzed such as the temperature, the duration of heat stress and that of the recovery period. Under every condition tested, we have found that the kinetics of activation of both promoters were very similar. In addition, both showed a similar inhibition by actinomycin D. These results were compared to those obtained with a DNA construct containing the early promoter of SV-40 virus coupled to the Cat gene. In this case, no heat-mediated accumulation of CAT protein was observed, indicating that the transcriptional activation of HIV-1 LTR by heat shock is specific. HIV-1 LTR contains two NF-kappa B binding elements, involved in the activation of this promoter during oxidative stress, which are sequence related to the heat shock element HSE. However, under all the heat shock conditions tested, we have been unable to detect the binding of any protein to kappa B elements, suggesting that this site is not directly involved in the thermal activation of HIV-1 LTR. These results indicate that the thermal transcriptional activation of HIV-1 LTR and hsp70 promoters occurs through different mechanisms that are triggered by similar heat shock conditions.

Base Sequence↗
PubMed1994

The kinetics of HIV-1 long terminal repeat transcriptional activation resemble those of hsp70 promoter in heat-shock treated HeLa cells.

The long terminal repeat (LTR) of human immunodeficiency virus type 1 (HIV-1) is activated under different conditions including heat shock. By using transient transfection assay, we have compared the thermal activation of HIV-1 LTR to that of the promoter of the gene encoding the human stress protein hsp70 which is under the control of the heat shock transcription factor HSF. In these assays, the chloramphenicol acetyl transferase (Cat) gene was used as a reporter gene. Several parameters of the heat stress were analyzed such as the temperature, the duration of heat stress and that of the recovery period. Under every condition tested, we have found that the kinetics of activation of both promoters were very similar. In addition, both showed a similar inhibition by actinomycin D. These results were compared to those obtained with a DNA construct containing the early promoter of SV-40 virus coupled to the Cat gene. In this case, no heat-mediated accumulation of CAT protein was observed, indicating that the transcriptional activation of HIV-1 LTR by heat shock is specific. HIV-1 LTR contains two NF-kappa B binding elements, involved in the activation of this promoter during oxidative stress, which are sequence related to the heat shock element HSE. However, under all the heat shock conditions tested, we have been unable to detect the binding of any protein to kappa B elements, suggesting that this site is not directly involved in the thermal activation of HIV-1 LTR. These results indicate that the thermal transcriptional activation of HIV-1 LTR and hsp70 promoters occurs through different mechanisms that are triggered by similar heat shock conditions.

Base Sequence↗
PubMed1988

Retinoic acid-induced differentiation of retrovirus-infected HL-60 cells is associated with enhanced transcription from the viral long terminal repeat.

I infected different human leukemic cell lines with an amphotropic retrovirus vector (designated PA317/N2) which confers G418 resistance and contains the Moloney murine leukemia virus long terminal repeat. In retrovirus-infected G418-resistant HL-60 cells, induction of granulocyte differentiation by retinoic acid was invariably accompanied by a marked increase (5- to 10-fold) in the transcriptional activity of the integrated retroviral long terminal repeat.

Blotting, Northern↗
PubMed1988

Trans-activation of the human immunodeficiency virus long terminal repeat by the hepatitis B virus X protein.

Human immunodeficiency virus 1 has been implicated as the main etiologic agent of the acquired immunodeficiency syndrome. However, other infectious agents may accelerate the progression of this disease. In particular, hepatitis B virus has been suggested as one such cofactor. Therefore, we have investigated the effects of hepatitis B virus gene products on expression of the human immunodeficiency virus I in transient transfection studies of Jurkat lymphoblastic T cells, using as reporter the chloramphenicol acetyltransferase gene coupled to the long terminal repeat of human immunodeficiency virus I. As measured by the amount of chloramphenicol acetyltransferase activity, gene expression directed by the human immunodeficiency virus I long terminal repeat increased approximately 10-fold in response to the hepatitis B virus X protein. This trans-activation by the X protein is multiplicative with the effect of phorbol esters and can be accounted for by an increase in the steady-state level of chloramphenicol acetyltransferase mRNA. Analysis of deletion and clustered point mutants in the long terminal repeat indicated that the X protein exerts its effect through multiple cis-acting sites. These results provide a possible molecular basis for the association of hepatitis B virus and the acquired immunodeficiency syndrome and confirm that the X protein is a transcriptional transactivator.

Acquired Immunodeficiency Syndrome↗
PubMed1983

Transcription of mouse mammary tumor virus: identification of a candidate mRNA for the long terminal repeat gene product.

We have examined an assortment of preneoplastic and neoplastic mouse mammary tissues for the presence of an mRNA which could encode the putative long terminal repeat gene product of mouse mammary tumor virus. We report here the detection of a novel mouse mammary tumor virus-specific, polyadenylic acid-containing transcript in certain preneoplastic and neoplastic mammary tissue of BALB/c mice. The molecule is 1.6 kilobases in length and contains sequences from the transcriptional leader and the U3 region of the proviral DNA. The upstream terminus of the 3' information lies 75 to 80 nucleotides from the beginning of the long terminal repeat open reading frame, in close proximity to a consensus splice acceptor in the DNA. The transcript was detected in hormonally or chemically induced neoplastic, preneoplastic, and lactating mammary tissue of BALB/c mice, but not in preneoplastic or tumor tissue induced by exogenous viruses in any strain of mice examined. This implies that the RNA we observed is transcribed from an endogenous provirus template.

Animals↗
PubMedDate not supplied

Hexamethylene bisacetamide and cis-platin stimulate the expression from the HIV-1 long terminal repeat sequence in human MCF-7 cells.

We have employed a recombinant plasmid pBHIV-1 carrying the Long Terminal Repeat sequences (LTR) of the Human Immunodeficiency Virus (HIV-1) linked to the reporter chloramphenicol acetyl transferase (CAT) gene and to the aminoglycoside phosphotransferase (aph) gene as a selectable marker. We have introduced pBHIV-1 DNA in human breast cancer MCF-7 cells and obtained the MCF-7HIV-1 cell line, resistant to geneticin. We have studied the effect of hexamethylene-bisacetamide (HMBA), cis-platin, interferon-aB, dexamethasone and tamoxifen on the LTR regulated CAT activity in MCF-7HIV-1 cells. It was found that HMBA and cis-platin stimulated CAT activity, whereas interferon-aB, dexamethasone and tamoxifen had no significant effect.

Acetamides↗
PubMed1991

The long terminal repeat is not a major determinant of the cellular tropism of human immunodeficiency virus type 1.

The long terminal repeats (LTRs) of human immunodeficiency virus type 1 (HIV-1) strains from the central nervous systems of four patients with AIDS and of an HIV-1 isolate which is highly macrophage-tropic were isolated by using the polymerase chain reaction. In transient transfection assays, these LTRs demonstrated no significant difference in basal or stimulated levels of transcription in any of a variety of cell lines tested, compared with expression directed from the LTR of a T-lymphocyte-tropic strain of HIV-1. Chimeric viruses were created with the LTRs of the macrophage-tropic and brain-derived viruses ligated to the viral backbone from a T-lymphocyte-tropic strain. No change in cellular tropism was demonstrated with these chimeric viruses. Thus, unlike the LTRs of some murine retroviruses, the LTR of HIV-1 does not appear to play a major role in determining cellular tropism.

Acquired Immunodeficiency Syndrome↗
PubMed1996

Strand displacement synthesis of the long terminal repeats by HIV reverse transcriptase.

According to the current model for retroviral replication, strand displacement of the long terminal repeat (LTR) is a necessary step during plus strand DNA synthesis in vivo. We have investigated the ability of human immunodeficiency virus reverse transcriptase (HIV-RT) to synthesize in vitro over a 634-nucleotide HIV LTR DNA template, having or lacking a single full-length DNA downstream primer. The presence of the downstream primer resulted in an approximately 12-fold reduction in the rate of upstream primer elongation. Addition of Escherichia coli single-stranded binding protein (SSB) or human replication protein A (RP-A) enhanced strand displacement synthesis; however, addition of HIV nucleocapsid protein (NC) did not. The presence of excess single-stranded DNA complementary to the downstream primer did not stimulate displacement synthesis. Interestingly, we observed that the elongating upstream primer could readily transfer to this DNA. This observation suggests that recombination is favored during strand displacement synthesis in vivo.

Base Sequence↗
PubMed1994

Identification and characterization of a human herpesvirus 6 gene segment capable of transactivating the human immunodeficiency virus type 1 long terminal repeat in an Sp1 binding site-dependent manner.

The human immunodeficiency virus type 1 (HIV-1) long terminal repeat (LTR) is transactivated by various extracellular signals and viral cofactors that include human herpesviruses. These transactivators are capable of transactivating the HIV-1 LTR through the transactivation response element, NF-kappa B, or other regulatory binding elements. Human herpesvirus 6 (HHV-6) is a potential cofactor of HIV-1. Here, we report that an HHV-6 gene segment, ZVH14, which can neoplastically transform NIH 3T3 and human keratinocytes, is capable of transactivating HIV-1 LTR chloramphenicol acetyltransferase constructs in an Sp1 binding site-dependent manner. Transactivation increased synergistically in the presence of multiple Sp1 sites and was dramatically reduced by cotransfection with oligomers designed to form triplex structures with HIV-1 LTR Sp1 binding sites. HIV-1 LTR NF-kappa B sites were not essential for ZVH14-mediated transactivation. A putative open reading frame in ZVH14, B115, which may encode a highly basic peptide consisting of 115 amino acid residues, showed transactivation capacity similar to that of ZVH14. This open reading frame also transactivated the HIV-1 LTR in an Sp1 site-dependent fashion in African green monkey kidney cells and human T cells. These data suggest that HHV-6 may stimulate HIV-1 replication via transactivation of Sp1 binding sites present in the HIV-1 promoter.

Amino Acid Sequence↗
PubMed1995

Identification of sequences in the long terminal repeat of the lymphoproliferative disease virus required for efficient transcription.

We analyzed the long terminal repeat (LTR) of the lymphoproliferative disease virus of turkeys for sequences that influence its promoter activity by using the chloramphenicol acetyltransferase assay. A series of LTR deletion mutants and recombinants between LTR and simian virus 40 regulatory sequences were used for these studies. Through transfection experiments, we identified a negative regulatory element residing at the 5' end of the U3. The two imperfect direct repeats (DRs) located at nt - 170 to - 125 upstream of the RNA transcription site were identified as enhancer elements which could stimulate transcription of a heterologous promoter in an orientation independent manner. Specific interaction of nuclear factors with the DRs element was identified. The two DRs contain cArg motifs which are suggested to play a role in tissue specific expression of several cellular genes.

Alpharetrovirus↗
PubMed1995

cis-acting regulatory elements in the bovine immunodeficiency virus long terminal repeat.

Functional cis-acting regulatory elements in the bovine immunodeficiency virus (BIV) long terminal repeat (LTR) were identified by deletion mapping and nuclear protein gel shift analysis using three BIV-infectible cell lines, Cf2Th, BLAC-20, and EREp. Deletion mapping studies indicated that putative NF-kappa B, GRE, AP-4, AP-1, CAAT, and ATF/CRE transcription factor elements positively contribute to LTR-directed gene expression in each cell line both in the presence and absence of the viral transactivator Tat. Sp1 and overlapping AP-3 and retroviral core enhancer elements had variable effects on LTR-directed gene expression depending on cell type and presence or absence of Tat. In addition, a sequence spanning the LTR U5 region and the untranslated viral leader was strongly repressive in all cell lines. Tat transactivated the LTR 25-fold over basal levels in a TAR-dependent manner in Cf2Th cells. In contrast, Tat transactivated the LTR only 2.5-fold over basal levels in EREp and BLAC-20 cells in a TAR-independent manner. Probes for putative NF-kappa B, GRE, Sp1, AP-4, AP-1, overlapping AP-3 and retroviral core enhancer, and juxtaposed CAAT and ATF-CRE elements specifically bound nuclear proteins from these three cell lines and HeLa cells, with the stoichiometry of binding being cell-type dependent. Probes for AP-4, AP-1, and juxtaposed CAAT and ATF/CRE elements exhibited greater protein binding with extracts from virally infected cells than with extracts from uninfected cells, suggesting that viral infection can modulate nuclear factor binding. The present studies indicate that several transcription factor elements in the BIV LTR have functional roles and that cell type can strongly determine the role they play in gene expression.

Animals↗
PubMed1992

Sequence variations and promoter activities of long terminal repeats from rat intracisternal A-particles.

Nucleotide sequences of three novel rat long terminal repeats (LTR) of intracisternal A-particles (IAP) were determined and compared with two previously published solitary rat IAP LTRs from the genomic clone H12 (Furter et al. (1989) J. Biol. Chem. 264, 18276-18279) and from the upstream region of the oncomodulin (OM) gene (Banville and Boie (1989) J. Mol. Biol. 207, 481-490). These five LTRs have a length of 286 to 370 bp and show the major variability within the U3 region. The CCAAT and the TATA boxes, the AATAAA polyadenylation signals and the CA polyadenylation sites are well conserved in sequence and position in all five LTRs, whereas several putative transcriptional factor binding sites in the U3 domain show considerable heterogeneity. The transcriptional activities of three LTRs were tested in transient gene expression assays using the human growth hormone (hGH) reporter gene in chemically transformed T14c cells which produce considerable amounts of oncomodulin. Promoter strengths of the three investigated LTRs varied considerably.

Animals↗