Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:1.14.14.3 (luciferase)
 38,195 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

To gain insight into mechanisms of cell type-specific transcription of class mu-glutathione S-transferase genes, the gene encoding the Yb3 subunit was cloned. Yb3 subunits are selectively expressed at high levels in rat brain and testis but not in liver or kidney. The Yb3 subunit gene spans over 6 kb and consists of 8 exons and 7 introns and a sequence consisting of tandem direct repeat consensus octamer DNA binding motifs separated by a 6 base pair (bp) spacer was identified in its 5'-flanking region. Gel shift assays with a 40 bp segment of DNA containing the two consensus octamer sequences, revealed the presence of specific binding proteins in nuclear extracts of rat brain, testis and C6 glioma cells. DNA binding activity was greatly reduced in liver, kidney and HTC cells. Reporter vectors carrying segments of the 5'-flanking region of the Yb3 subunit gene fused to a luciferase gene were introduced into C6 glioma cells which express high levels of Yb3 subunits, and into HTC cells which do not. The plasmids consisting of the Yb3 gene promoter up to, but not including, the octamer motifs did not support luciferase transcription in the C6 glioma cells, but larger fragments that included the octamer repeat sequences, effectively directed transcription in the C6 glioma cells. With mutated octameric sequences transcriptional activity was greatly reduced, and none of the same Yb3 constructs directed substantial luciferase transcription in the HTC cells. The results show that octamer motifs in the 5'-flanking region of the Yb3 subunit gene are functional and are the principal cis-acting elements that account for its discrete cell type-selective expression. This gene is one of the few known targets for octamer DNA binding transcription factors in brain.
 ...
PMID:Brain and testis selective expression of the glutathione S-transferase Yb3 subunit is governed by tandem direct repeat octamer motifs in the 5'-flanking region of its gene. 770 76

Modulation of the expression of the D2-dopamine receptor gene is involved in several pathological and developmental circumstances. The gene and the corresponding promoter regions of the rat D2 receptor were isolated and partly characterized to study its regulation. The rat D2-receptor gene spans at least 50 kb, and possesses eight exons; its organization was compared to those of the other dopamine-receptor genes in a phylogenetic perspective. The gene contains two transcription-start sites: the major one is located about 320 bp upstream from the 3' end of the first exon, and a minor site is 70 bp further upstream. Transient-expression assays with fusion constructs comprising fragments of the D2-promoter region and the luciferase reporter gene confirmed the existence of two independent, TATA-lacking promoters. Both promoters separately induced transcription of the luciferase gene in C6 glioma, primary fibroblasts, GH3 and MMQ pituitary cell lines, among which only the MMQ cells normally express the D2 receptor. Transcription is enhanced by the reunion of the two promoters, and modified by the addition of upstream sequences. Thus the 1-kb promoter region analysed does not contain all the elements necessary to confer tissue-specific expression of the gene, but does carry some positive and negative regulatory elements, which remain to be characterized.
 ...
PMID:Transcription of the rat dopamine-D2-receptor gene from two promoters. 812 17

Regulation of steroidogenesis in classic endocrine tissues is mediated by transcriptional regulation of the P450scc gene, which encodes the first and rate-limiting cholesterol side-chain cleavage enzyme. We previously showed that P450scc messenger RNA is regionally expressed in the adult rat brain, primary glial cultures, and C6 glioma cells. Expression of P450scc in the brain results in the de novo synthesis of neurosteroids, a class of steroid hormones that are active at gamma-aminobutyric acidA and N-methyl-D-aspartate receptors. We determined whether P450scc expression is transcriptionally regulated in neural cells, using the same DNA sequences and nuclear proteins as classic steroidogenic adrenal and Leydig cells. The transcriptional activity of deletional mutants of 2.5 kilobases of the 5'-flanking regulatory region of the rat P450scc gene cloned into a luciferase reporter gene was assessed in mouse adrenocortical Y-1, mouse Leydig MA-10, rat C6 glioma, rat GC somatotrope, and mouse GT1-7 neurosecretory cell lines. P450scc was transcriptionally regulated in Y-1, MA-10, and C6 glioma cells, but not in GC or GT1-7 cells. In one region (-94/-35), putative steroidogenic factor-1-binding sites appeared to be critical for the basal transcriptional activity and cAMP responsiveness in steroidogenic Y-1 and MA-10 cells, but had no function in rat C6 cells. DNA sequences between -94/-130 mediated both basal and cAMP-inducible transcriptional activity in C6 cells. Gel mobility shift assays showed that one nuclear protein binding to DNA sequences between -54 and -35 was abundant in MA-10 and Y-1 cells, but was absent from C6 cells, whereas another nuclear protein, binding to DNA sequences between -94 and -130 was abundant in C6 cells, but was rare in MA-10 cells and absent from Y-1 and other adrenocortical cells. Although the DNA sequence between -94 and -130 contains an Sp1 site, Sp1 did not bind to this site. Nevertheless, this GC-rich region was critical for nuclear protein binding and for basal and cAMP-induced transcriptional regulation in both C6 and MA-10 cells. These observations demonstrate that the rat P450scc gene is transcriptionally regulated in glioma cells, but its regulation in glial cells involves a DNA element different from those used in classic steroidogenic tissues. The results further suggest that steroidogenic factor-1 is not involved in regulating neurosteroidogenesis.
 ...
PMID:Transcriptional regulation of P450scc gene expression in neural and steroidogenic cells: implications for regulation of neurosteroidogenesis. 858 34

The posttranscriptional regulation of glucose transporter GLUT1 gene expression may be mediated by specific interactions of cytosolic proteins and regulatory cis-elements within the untranslated regions (UTRs) of the GLUT1 mRNA. These putative cis/trans interactions were examined in the present studies with RNase T1 protection assays using 32P-labeled GLUT1 3'-UTR prepared from transcription plasmids and cytosolic proteins from C6 rat glioma cells. RNase T1 mapping studies localized a cis-element to nucleotides 2,170-2,207 on the bovine GLUT1 mRNA 3'-UTR. Ultraviolet cross-linking of RNA/protein complexes identified two complexes having molecular masses of 88 and 44 kDa. Competition studies with synthetic RNA and oligodeoxynucleotides showed the 88-kDa complex reacted with nucleotides 2,180-2,197 and that the 44-kDa complex reacted with sequences within nucleotides 1,717-2,132 of the bovine GLUT1 mRNA. The GLUT1 3'-UTR between nucleotides 2,100 and 2,300 was generated by polymerase chain reaction and subcloned at a unique Pfl/MI site within the 3'-UTR of a luciferase gene within the mammalian expression vector pGL2. Transfection of C6 rat glioma cells with the luciferase expression vector containing this portion of the GLUT1 3'-UTR resulted in a sixfold increase in luciferase gene expression in C6 cells. The identification of these cis/trans mechanisms provides support for the hypothesis that the posttranscriptional regulation of GLUT1 gene expression may be mediated by the interaction of specific cytosolic proteins with the GLUT1 mRNA 3'-UTR.
 ...
PMID:Cis-element/cytoplasmic protein interaction within the 3'-untranslated region of the GLUT1 glucose transporter mRNA. 859 13

We have demonstrated the presence of a rat prion protein (RaPrP) gene promoter upstream of multiple initiation sites. A 0.1-kb fragment upstream of the 5'-untranslated region contains specific DNA motifs characteristic of promoter elements including an AP-1 binding site, an inverted CCAAT motif and three inverted Sp-1 binding sites. This fragment directs transcription of a luciferase reporter gene in pheochromocytoma cells (PC12) and rat glioma cells (C6), suggesting that it contains the promoter for the RaPrP gene. To more precisely localize the transcription regulatory elements in this region, a series of 5'-deletion mutants were generated. Deletion analysis showed that an inverted CCAAt and adjoining Sp-1 binding sequences may play an important role in transcription of the RaPrP gene.
 ...
PMID:Identification of a promoter region in the rat prion protein gene. 861 25

Expression of mitogenic basic fibroblast growth factor (bFGF) in the central nervous system is inhibited by direct cell contact and is implicated in reactive and neoplastic transformation of astrocytes. The molecular mechanisms controlling expression of bFGF were examined in cultures of human astrocytes. Cell-density-dependent depletion of bFGF mRNA levels parallels changes in bFGF gene protein. Regulation of transcription of a bFGF luciferase reporter gene containing an upstream region (bp -1800 to +314) of the bFGF gene promoter mimicks the density-dependent regulation of the endogenous bFGF gene in transfected astrocytes. Deletion analysis has identified a fragment (bp -650 to -513) and sequences further downstream (bp -274 to +314) as the regions required for the regulation of bFGF gene activity by cell density. Unlike in astrocytes, changing the cell density of glioma cell cultures does not affect the levels of bFGF protein and mRNA. bFGF luciferase constructs were expressed at the same level in high- or low-density cultures of glioma cells, indicating altered regulation of the bFGF gene promoter. Electrophoretic mobility shift assays showed binding of nuclear proteins to a fragment of bFGF gene promoter from bp -650 to -453. This binding was abolished by a deletion of the upstream cell-density-responsive region (bp -650 to -512). Binding was observed with nuclear extracts from subconfluent astrocytes but was reduced in extracts from confluent astrocytes. Our results indicate that induction of bFGF in astrocytes upon reduction of cell density is mediated transcriptionally by positive trans-acting factors interacting with bFGF promoter. In contrast, nuclear proteins from glioma cells bind to the promoter region from bp -650 to -453 independent of cell density. Thus, the constitutive binding of trans-acting factor(s) to the region of the bFGF promoter from bp -650 to -453 may be responsible for the continuous expression of bFGF that leads to the uncontrolled growth of glioma cells.
 ...
PMID:Promoter regions involved in density-dependent regulation of basic fibroblast growth factor gene expression in human astrocytic cells. 863 98

Herein we describe experiments showing that the early growth response gene 1 (EGR-1) promoter is sufficient to confer selective expression of the luciferase gene (Luc) in glioma cell lines exposed to ionizing radiation. Activity of the EGR-1 promoter was investigated in human glioblastoma cells using the plasmid vector, pEGR-Luc. The EGR-1 promoter gene directed radiosensitive expression of luciferase. This promoter showed high levels of activity (10-fold) in irradiated glioma cell lines as compared to basal levels of activity in nonirradiated cell lines. Maximum activation was detectable at 1-3 hr after stimulation with 20 Gy. The results also demonstrate that cells modified to contain the herpes simplex virus-thymidine kinase (HSV-tk) gene under control of the EGR-1 promoter become sensitive to treatment with the antiviral agent ganciclovir (GCV), whereas nonirradiated cells and nontransfected cells were unaffected by this agent. This results suggest that therapeutic genes can be expressed selectively in irradiated glioma cells. The results also indicate that the EGR-1 promoter can be used to induce exogenous genes selectively in radiation fields used for the treatment of malignant brain tumors.
 ...
PMID:Activation of the radiosensitive EGR-1 promoter induces expression of the herpes simplex virus thymidine kinase gene and sensitivity of human glioma cells to ganciclovir. 866 75

IGF-I gene transcription is regulated by two promoters--the major promoter which is active in all tissues and regulates transcription of IGF-I mRNAs that contain exon 1 and a second promoter which regulates transcription of IGF-I mRNAs that contain exon 2 and from which significant transcription is restricted to the liver. The major promoter is a TATAA-less promoter that lacks both a CAAT box and SP1 binding sites and that utilizes multiple transcription initiation sites. The current studies were designed to delineate the functional elements of the major promoter. Transient transfection assays using rat C6 glioma cells and rat dermal fibroblasts in primary culture demonstrated that basal activity of the major promoter was located between -18 (with +1 defined as the most 5' transcription initiation site in exon 1) and +78 of exon 1. DNase I footprinting, which was performed using nuclear extracts from rat C6 glioma cells, demonstrated protein binding to a sequence that extended from -10 to +9 (termed IGFI-FP1). In gel shift assays, binding of C6 cell nuclear proteins to a 34-basepair (bp) oligonucleotide that contained IGFI-FP1 resulted in the formation of three specific protein-DNA complexes. The functional role of protein binding to IGFI-FP1 was examined by mutating the sequences between either -4 and -2 or -9 and -7 in IGF-I-luciferase fusion genes that contained either 412 or 18 bp of 5'-flanking region and 302 bp of exon 1. Both of these mutations altered protein binding to IGFI-FP1 as demonstrated by gel shift analysis. Transfection of the wild-type and mutant fusion genes into C6 glioma cells demonstrated that mutation of the nucleotides between -4 and -2 decreased luciferase activity to approximately 50% of wild-type activity, whereas mutation of the nucleotides between -9 and -7 decreased luciferase activity to 11-35% of wild-type activity. These data demonstrate that: (i) basal activity of the major promoter of the rat IGF-I gene is localized to the region between -18 and +78 of exon 1; (ii) protein binding sites are present within this region of the major promoter; and (iii) protein binding to this region contributes to basal expression of the IGF-I gene.
 ...
PMID:The major promoter of the rat insulin-like growth factor-I gene binds a protein complex that is required for basal expression. 867 54

ICER (inducible cyclic AMP early repressor), a member of the cyclic AMP response element (CRE) modulator (CREM) family of transcription factors, is a powerful repressor of cyclic AMP-mediated transactivation. Our studies characterize the regulation of ICER in C6 glioma cells and investigate its role in repressing transcription of the beta1-adrenergic receptor (beta1AR) gene. Incubation with isoproterenol (100 nM) results in a rapid induction in levels of mRNA for ICER and its splice variant ICERgamma, with maximal induction occurring after 2 h of treatment. Incubation with isoproterenol also increased levels of CREM isoforms within 1 h; this was unexpected given previous reports that these isoforms are not rapidly induced. Increased expression of ICER and CREM was accompanied by induction of two CRE-binding complexes. The presence of ICER in these two CRE-binding complexes is demonstrated by their disruption with CREM antibody and by their comigration with recombinant ICER. Because the time course for isoproterenol induction of ICER mRNA and CRE binding corresponds to that for down-regulation of beta1AR mRNA levels in C6 glioma cells, the influence of ICER beta1AR transcription was directly examined. Coexpression of ICER significantly decreased transcriptional activity of a rat beta1AR promoter-luciferase reporter construct that contains a CRE. In contrast, coexpression of ICER did not influence two truncated rat beta1AR promoter constructs that lack the CRE site. These data demonstrate that ICER can interact at the beta1AR promoter to repress transcription.
 ...
PMID:Adrenergic regulation of ICER (inducible cyclic AMP early repressor) and beta1-adrenergic receptor gene expression in C6 glioma cells. 876 72

In this study, we investigated whether the regulation and the copy number of the herpes simplex virus thymidine kinase (HSVtk) gene increased the sensitization to ganciclovir (GCV) of glioma cell lines (Rat C6 and human U118-MG) using liposome-mediated gene transfer. Three recombinant plasmids carrying the HSVtk gene driven by the thymidine kinase promoter in single (pAGo) and double copy (pYED) or by the human cytomegalovirus promoter (pCMVtk) were used for the transfection. The DNA delivery was optimized by screening a panel of cationic liposomes using Lac-Z and luciferase as reporter genes. The efficiency of transfection reached 33% to 36% in vitro but only 18.6% in vivo after an intratumoral injection of DNA-liposome complexes. Moreover, after transfection of the three plasmids, the cell-killing effect of GCV was evaluated. A significant enhancement (four- to fivefold) of the cell sensitivity to GCV was shown in pCMVtk and pYED as compared with pAGo-transfected cells in both cell lines. According to the plasmid, the effect of the HSVtk/GCV system was confirmed by in vivo experiments and was objectified by a higher tumor weight reduction with pCMVtk (49%) than pAGo (27%). From these results, we conclude that (1) the gene transfer can be achieved by cationic liposomes both in vitro and in vivo and that (2) using this type of vector, the antitumor effect of the HSVtk/GCV system could be potentiated by the up-regulation of HSVtk gene duplication.
 ...
PMID:Liposomal delivery of the herpes simplex virus thymidine kinase gene in glioma: improvement of cell sensitization to ganciclovir. 898 41


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