Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

---

Query: EC:1.14.14.3 (luciferase)
38,195 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Expression of a gene encoding the diphtheria toxin A (DT-A) chain, under the control of human immunodeficiency virus-1 (HIV-1) proteins Tat and Rev, has previously been shown to confer on cells an impaired ability to produce HIV. That work was done in HeLa cell lines that had stably integrated the regulated DT-A gene in a plasmid context. To increase the efficiency with which the HIV-regulated DT-A gene could be introduced into cells, we studied a recombinant, amphotropic murine leukemia virus containing the HIV-regulated DT-A transcription unit. Here we demonstrate that such recombinant retroviruses can be packaged, for both wild-type DT-A and an attenuated version, tox 176. In transient transfection assays, the proviral constructs exhibited similar basal and trans-activated levels of DT-A expression to the parental plasmids. Transduced H9 cells expressed the integrated DT-A gene upon transfection with plasmids encoding Tat and Rev, as assayed by decreased expression of a cotransfected luciferase reporter gene. Furthermore, the transduced H9 cells were substantially impaired in their ability to produce HIV, as demonstrated by p24 assays of culture supernatants following either transfection with an HIV proviral clone or infection with HIV-IIIB. These data demonstrate that basal expression of the regulated DT-A gene has been reduced to a tolerable level, both in packaging cells and transduced H9 cells. The use of HIV-regulated retroviruses encoding the highly lethal DT-A product may eventually be applicable as a gene therapy approach for the acquired immunodeficiency syndrome (AIDS).
...
PMID:Inhibition of human immunodeficiency virus-1 production resulting from transduction with a retrovirus containing an HIV-regulated diphtheria toxin A chain gene. 132 91

Recent findings have indicated that megakaryocytes may be susceptible to human immunodeficiency virus (HIV) infection, suggesting a potential role for megakaryocytes as viral reservoirs in HIV-infected patients. We report that the megakaryocytic cell line Dami could be productively infected with the HTLV III-B strain of HIV-1, in 26 different experiments (results of 16 experiments are reported); productive infection lasted up to 30 weeks. Despite a lack of detectable surface expression of the CD4 molecule and very low levels of CD4 mRNA, between 40% and 60% of megakaryocytic cells produced viral proteins after contact with HIV-1. Neither cytopathogenic effects nor syncytial formation was observed. Production of high levels of functional viral particles was indicated by analysis of p24 protein levels, reverse transcriptase activity, ultrastructural studies, and the capacity of supernatants from infected Dami cells to infect the Molt-4 T-lymphocytic cell line. HIV-1 RNA and protein levels in infected Dami cells were enhanced by treatment with tumor necrosis factor-alpha (TNF-alpha), and decreased by treatment with interferon-alpha (IFN-alpha) and IFN-gamma. Transient transfection of the megakaryocytic cells with various constructs of the HIV-1 promoter (LTR) linked to the luciferase reporter gene suggested that the effect of TNF-alpha was related, as in monocytic and T-cell lines, to transactivation of the enhancer region of the HIV-1 LTR. These findings indicate that signals provided by the immune system may modulate HIV-1 expression in cells of the megakaryocytic lineage.
...
PMID:Productive human immunodeficiency virus-1 infection of megakaryocytic cells is enhanced by tumor necrosis factor-alpha. 158 16

By using human CD4+ lymphoblastoid T cells transiently cotransfected with human immunodeficiency virus (HIV) and cytomegalovirus (CMV), we tested whether modulation of T-cell activation through the protein kinase C (PKC) or the protein kinase A (PKA) pathway synergized with CMV immediate-early (IE) proteins in HIV long terminal repeat (LTR) transactivation. Stimulation with phorbol myristate acetate, tumor necrosis factor, or cross-linked antibodies to CD3 and CD28 resulted in modest enhancement (two- to fourfold) of the activity of a luciferase expression vector under control of the HIV LTR. Cotransfection of a vector expressing the CMV IE1 and IE2 proteins under the control of their own promoter enhanced HIV LTR activity 16- to 49-fold. Combination of any one of the above stimuli and CMV IE expression amplified HIV LTR activity 99- to 624-fold. Stimulation of PKA-dependent pathways with forskolin, 8-bromo cyclic AMP, or prostaglandin E2 had a minimal effect on HIV LTR activity, whereas such stimuli resulted in synergistic amplification in cells cotransfected with CMV IE (three- to fivefold increases over the effects of CMV IE alone). This synergism was independent of the NF-kappa B binding motifs within the HIV LTR. CMV IE2, but not IE1, protein induced HIV transactivation and synergized with signals modulating T-cell activation. The intense synergism observed was superior to the increase in IE protein expression following PKC activation by phorbol myristate acetate. Treatment of cells with PKC inhibitor GF109203X blocked most of the observed synergism.(ABSTRACT TRUNCATED AT 250 WORDS)
...
PMID:Modulation of T-cell activation through protein kinase C- or A-dependent signalling pathways synergistically increases human immunodeficiency virus long terminal repeat induction by cytomegalovirus immediate-early proteins. 165 49

Construction of a DNA plasmid that expresses a diphtheria toxin A chain (DT-A) gene under control of human immunodeficiency virus (HIV-1) proteins Tat and Rev has been described. Here the generation of HeLa cell clones containing integrated, HIV-regulated DT-A sequences is reported. Five such clones were identified by their decreased expression of a luciferase reporter gene transiently cotransfected with Tat- and Rev-encoding plasmids. The decreased luciferase expression most probably was due to activation of the integrated DT-A gene because higher luciferase activity could be restored by introducing either DT antitoxin or a gene encoding a mutant, DT-resistant elongation factor 2 (the intracellular target for DT-A). Analysis by polymerase chain reaction (PCR) indicated that all clones expressed DT-A encoding RNA. The clones were then transfected with an HIV proviral clone and were tested for HIV production; all five clones demonstrated substantially impaired HIV production compared with parental HeLa cells, as shown by p24 assays of culture supernatants. Our success in generating these cell lines indicates that extremely low basal expression has been achieved in view of the high cellular lethality of DT-A. HIV-regulated expression of DT-A may be applicable as a gene therapy approach for the acquired immune deficiency syndrome (AIDS), to bring about selective suicide of HIV-infected cells before production of viral progeny.
...
PMID:Inhibition of HIV production in cells containing an integrated, HIV-regulated diphtheria toxin A chain gene. 173 39

Expression of a gene encoding the diphtheria toxin A (DT-A) fragment, controlled by tissue specific regulatory elements, has previously been used to kill selected cell populations. Here, we have examined the feasibility of controlling DT-A expression using regulatory systems from the human immunodeficiency virus (HIV-1) genome. Plasmids were constructed which express either DT-A or, as a model system, the luciferase (luc) reporter gene, under control of HIV-1 long terminal repeat (LTR) sequences (-167 to +80). While trans-activation by expression of the viral protein Tat was demonstrated, significant basal expression was observed. To reduce basal expression, cis-acting negative regulatory elements from the env region of the HIV-1 genome were inserted in the 3' untranslated region of both the luc and DT-A constructs. This dramatically reduced basal expression from the HIV LTR, and now both viral regulatory proteins Tat and Rev were required for maximal trans-activation. Such regulation of DT-A expression might be therapeutically applied to selectively kill HIV-infected cells in acquired immunodeficiency syndrome (AIDS) and AIDS-related complex (ARC).
...
PMID:Activation of a diphtheria toxin A gene by expression of human immunodeficiency virus-1 Tat and Rev proteins in transfected cells. 186 40

UV irradiation has been shown to activate the human immunodeficiency virus type 1 (HIV-1) long terminal repeat (LTR) in cell culture; however, only limited studies have been described in vivo. UV light has been categorized as UV-A (400 to 315 nm), -B (315 to 280 nm), or -C (less than 280 nm); the longer wavelengths are less harmful but more penetrative. Highly penetrative UV-A radiation constitutes the vast majority of UV sunlight reaching the earth's surface but is normally harmless. UV-B irradiation is more harmful but less prevalent than UV-A. In this report, the HIV-1 LTR-luciferase gene in the skin of transgenic mice was markedly activated when exposed to UV-B irradiation. The LTR in the skin of transgenic mice pretreated topically with a photosensitizing agent (psoralen) was also activated to similar levels when exposed to UV-A light. A 2-h exposure to sunlight activated the LTR in skin treated with psoralen, whereas the LTR in skin not treated with psoralen was activated after 7 h of sunlight exposure. The HIV-1 LTR-beta-galactosidase reporter gene was preferentially activated by UV-B irradiation in a small population of epidermal cells. The transgenic mouse models carrying HIV-1 LTR-luciferase and LTR-beta-galactosidase reporter genes have been used to demonstrate the in vivo UV-induced activation of the LTR and might be used to evaluate other environmental factors or pharmacologic substances that might potentially activate the HIV-1 LTR in vivo.
...
PMID:In vivo activation of human immunodeficiency virus type 1 long terminal repeat by UV type A (UV-A) light plus psoralen and UV-B light in the skin of transgenic mice. 190 29

Since human immunodeficiency virus (HIV) nef has been suggested to exert regulatory effects on HIV long terminal repeat (LTR) activity, we transiently transfected HIV LTR chloramphenicol acetyltransferase or luciferase expression vectors into a human astrocytoma clone (U-373nef) that constitutively expresses the HIV nef gene. In these cells, basal HIV LTR activity, as well as tumor necrosis factor-induced or tat-driven activity, was similar to that in control cells. Lack of any detectable effect of HIV nef on LTR activity was not the result of mutations in integrated nef DNA, as was shown by polymerase chain reaction. These data suggest that the role of nef in HIV genome transcription does not necessarily involve a direct influence on HIV LTR activity.
...
PMID:Constitutive expression of human immunodeficiency virus (HIV) nef protein in human astrocytes does not influence basal or induced HIV long terminal repeat activity. 218 77

A microtransfection method, using either the DEAE-dextran or the Ca.phosphate procedure has been developed. A plasmid expressing the luciferase-encoding gene under the control of the human immunodeficiency virus (HIV) LTR promoter was constructed. Transfections were performed in 96-well plates, allowing statistical evaluation of the results. This microtransfection method requires the use of 100- to 1000-fold less plasmid and cells than in a conventional chloramphenicol acetyl transferase (CAT) assay. A Luciferase index which takes into account cell viability after transfection has been defined using a semi-automated absorbance assay. A 20-h incubation period post-transfection is sufficient for optimal results. Basal long terminal repeat activity and autologous Tat transactivation were studied in various lymphoid, monocytic and adherent human cell lines. Infection of microtransfected cells by HIV activated luc expression. This assay can thus also be used for rapid detection and quantitation of HIV. Antiviral activities of drugs can be assessed in a two-day test.
...
PMID:A microtransfection method using the luciferase-encoding reporter gene for the assay of human immunodeficiency virus LTR promoter activity. 218 84

An attenuated vaccinia virus mutant with specific genetic lesions has been used to develop a vehicle for safer live recombinant virus vaccines. The mutant virus 48-7 has an 8-MDa deletion starting 2.2 MDa from the left end of the viral genome and point mutations in the gene encoding the 14-kDa fusion protein that determines the plaque-size phenotype of the virus. Using the highly sensitive reporter gene luciferase, we have shown that this mutant can generate recombinant viruses that infect cultured cells and animals with normal vaccinia virus tropism. Insertion of the envelope and gag genes of human immunodeficiency virus type 1 into the attenuated vaccinia mutant resulted in their efficient expression and precursor processing in infected cultured cells. Infection of mice with human immunodeficiency virus-vaccinia recombinant viruses elicited human immunodeficiency virus-specific antibodies. Using mice pretreated with cyclophosphamide as a model for immunosuppression, the reduced virulence of the mutant recombinant virus was clearly evident. These findings demonstrate that the highly attenuated vaccinia virus mutant 48-7 can be used to generate effective and safer vaccines.
...
PMID:Highly attenuated vaccinia virus mutants for the generation of safe recombinant viruses. 278 4

Retroviral integration is the step which leads to establishment of the provirus, cis- and trans-acting regions of the human immunodeficiency type 1 (HIV-1) retrovirus genome, including the attachment site (att) at the ends of the unintegrated viral DNA and the conserved domains within the integrase (IN) protein, have been identified as being important for integration. We investigated the role of each of these regions in the context of an infectious HIV-1 molecular clone through point mutagenesis of the att site and the zinc finger-like and catalytic domains of IN. The effect of each mutation on integration activity was examined by using a single-step infection system with envelope-pseudotype virus. The relative integration efficiency was estimated by monitoring the levels of viral DNA over time in the infected cells. The integration activities of catalytic domain point mutants and att site deletion mutants were estimated to be 0.5 and 5% of wild-type activity, respectively. However, in contrast with previous in vitro cell-free integration studies, alteration of the highly conserved CA dinucleotide resulted in a mutant which still retained 40% of wild-type integration activity. The relative levels of expression of each mutant, as measured by a luciferase reporter gene, correlated with levels of integration. This observation is consistent with those of previous studies indicating that integration is an obligatory step for retroviral gene expression. Interestingly, we found that three different HIV-1 constructs bearing point mutations in the zinc finger-like domain synthesized much lower levels of viral DNA after infection, suggesting impairment of these mutants before or at the initiation of reverse transcription. Western blot (immunoblot) analysis demonstrated wild-type levels of reverse transcriptase within the mutant virions. In vitro endogenous reverse transcription assays indicated that all three mutants in the zinc finger-like domain had wild-type levels of reverse transcriptase activity. These data indicate that in addition to integration, IN may have an effect on the proper course of events in the viral life cycle that precede integration.
...
PMID:Genetic analysis of human immunodeficiency virus type 1 integrase and the U3 att site: unusual phenotype of mutants in the zinc finger-like domain. 747 78


1 2 3 4 5 6 7 8 9 10 Next >>

---