scholarly journals Promoter elements in the transcriptional activation of the human stromelysin-1 gene by the inflammatory cytokine, interleukin 1

1994 ◽  
Vol 302 (2) ◽  
pp. 471-477 ◽  
Author(s):  
S Quinones ◽  
G Buttice ◽  
M Kurkinen
Keyword(s):  
Transcriptional Activation ◽  
Inflammatory Cytokine ◽  
Interleukin 1 ◽  
Gene Promoter ◽  
Flanking Sequence ◽  
Tissue Remodelling ◽  
Native Promoter ◽  
Cell Extracts ◽  
Extracellular Matrix Components ◽  
The Absolute

Stromelysin-1, a tissue-remodelling metalloproteinase synthesized by fibroblasts, has proteolytic activity against a variety of extracellular matrix components. Stromelysin-1 gene transcription is induced by the inflammatory cytokine interleukin (IL)-1. In fibroblasts transiently transfected with constructs containing 5′-deletion mutants of the human stromelysin-1 gene promoter, IL-1-induced transcriptional activity was abolished with the removal of region -102 to -54. This region includes an AP-1 binding site at positions -70 to -64. The AP-1 site alone increased the basal activity of and conferred minimal IL-1 inducibility onto the heterologous gene promoter of thymidine kinase. Interestingly, although the removal of the AP-1 site from the native promoter (-1303 to +4) affected the absolute levels of IL-1-induced and basal promoter activity, it did not alter their ratio, indicating the involvement of regions outside the AP-1 site in the IL-1 response. Of the stromelysin-1 5′ flanking sequence examined, only the region -274 to -54 could confer IL-1 inducibility to a heterologous promoter independently of the AP-1 site. This region also bound specific nuclear factors. Further analysis revealed that the region composed of -86 to -71 and -63 to -54 could independently respond to IL-1 and bind protein of whole cell extracts. Protein binding to this region and to the AP-1 site was modestly induced by IL-1 treatment. From these results we conclude that, in fibroblasts, the AP-1 site (-70 to -64) is not necessary for the IL-1 response; however, it probably interacts through protein associations with the responsive region immediately surrounding it in the absolute transcriptional activation of the human stromelysin-1 gene by IL-1.

scholarly journals CpG methylation at the USF-binding site is important for the liver-specific transcription of the chipmunk HP-27 gene

2006 ◽  
Vol 395 (1) ◽  
pp. 203-209 ◽  
Author(s):  
Gen Fujii ◽  
Yuki Nakamura ◽  
Daisuke Tsukamoto ◽  
Michihiko Ito ◽  
Tadayoshi Shiba ◽  
...  
Keyword(s):  
Binding Site ◽  
Transcriptional Activation ◽  
Methylation Status ◽  
Gene Promoter ◽  
Cpg Methylation ◽  
Flanking Sequence ◽  
Upstream Stimulatory Factor ◽  
Cpg Dinucleotides ◽  
E Box ◽  
Cpg Dinucleotide

The chipmunk hibernation-specific HP-27 gene is expressed specifically in the liver and has a CpG-poor promoter. To reveal how the liver-specific transcription of the HP-27 gene is regulated, we performed yeast one-hybrid screening of a chipmunk liver cDNA library. A 5′-flanking sequence of the HP-27 gene, extending from −170 to −140 and containing an E-box (5′-CACGTG-3′), is essential for the liver-specific transcription of HP-27. We used this sequence as bait and found that a ubiquitously expressed transcription factor, USF (upstream stimulatory factor), bound to the E-box. In COS-7 cells, USF activated transcription from the HP-27 gene promoter. We then used bisulphite genomic sequencing to analyse the methylation status of the four CpG dinucleotides that lie in the 5′-flanking sequence of the HP-27 gene up to −450, to investigate how the ubiquitously expressed USF activates transcription of the HP-27 gene only in the liver, while its transcription is repressed elsewhere. The only difference in methylation in the tissues tested was in the CpG dinucleotide in the USF-binding site, which was hypomethylated in the liver, but highly methylated in the kidney and heart. The specific methylation of the CpG dinucleotide at the USF-binding site impeded both the binding of USF and its transcriptional activation of the HP-27 gene. Chromatin immunoprecipitation using anti-USF antibodies revealed that USF bound to the HP-27 gene promoter in the liver, but not in the kidney or heart. Thus CpG methylation at the USF-binding site functions in establishing and maintaining tissue-specific transcription from the CpG-poor HP-27 gene promoter.

scholarly journals Analysis by cell-free transcription of the liver-specific pyruvate kinase gene promoter.

1989 ◽  
Vol 9 (10) ◽  
pp. 4409-4415 ◽  
Author(s):  
S Vaulont ◽  
N Puzenat ◽  
A Kahn ◽  
M Raymondjean
Keyword(s):  
Binding Site ◽  
Pyruvate Kinase ◽  
Gene Promoter ◽  
Specific Factor ◽  
Nuclear Factor ◽  
Flanking Sequence ◽  
Kinase Gene ◽  
Cell Extracts ◽  
Nuclear Factor 1

A DNA fragment spanning nucleotides -183 to -4 with respect to the cap site of the rat L-type pyruvate kinase (L-PK) gene contains at least four binding sites for putative transcriptional factors: hepatocyte nuclear factor 1 (HNF1), liver factor A1 (LF-A1), nuclear factor 1 (NF1), and major late transcription factor (MLTF). This fragment was used to direct transcription of a reporter sequence (a G-free cassette) in cell extracts. This L-PK promoter was active in liver nuclear extracts, but not in extracts from nonhepatic tissues. A reduction of 50% of the activity was obtained with a deleted L-PK promoter containing only the HNF1-binding site. In contrast, deletion of the HNF1-binding site inactivated the promoter by more than 90%. These results were confirmed by titration experiments with synthetic oligonucleotides. Titration of HNF1 resulted in an 85% decrease of transcriptional activity, while titration of LF-A1 resulted in only a 40% decrease. The influence of NF1 and MLTF seemed to be marginal in this system. The proximal 5'-flanking sequence of the L-PK gene therefore appears to function in vitro as an efficient liver-specific promoter which requires the binding of the liver factor HNF1 and which is also stimulated by the binding of another liver-specific factor, LF-A1.

Analysis by cell-free transcription of the liver-specific pyruvate kinase gene promoter

1989 ◽  
Vol 9 (10) ◽  
pp. 4409-4415
Author(s):  
S Vaulont ◽  
N Puzenat ◽  
A Kahn ◽  
M Raymondjean
Keyword(s):  
Binding Site ◽  
Pyruvate Kinase ◽  
Gene Promoter ◽  
Specific Factor ◽  
Nuclear Factor ◽  
Flanking Sequence ◽  
Kinase Gene ◽  
Cell Extracts ◽  
Nuclear Factor 1

A DNA fragment spanning nucleotides -183 to -4 with respect to the cap site of the rat L-type pyruvate kinase (L-PK) gene contains at least four binding sites for putative transcriptional factors: hepatocyte nuclear factor 1 (HNF1), liver factor A1 (LF-A1), nuclear factor 1 (NF1), and major late transcription factor (MLTF). This fragment was used to direct transcription of a reporter sequence (a G-free cassette) in cell extracts. This L-PK promoter was active in liver nuclear extracts, but not in extracts from nonhepatic tissues. A reduction of 50% of the activity was obtained with a deleted L-PK promoter containing only the HNF1-binding site. In contrast, deletion of the HNF1-binding site inactivated the promoter by more than 90%. These results were confirmed by titration experiments with synthetic oligonucleotides. Titration of HNF1 resulted in an 85% decrease of transcriptional activity, while titration of LF-A1 resulted in only a 40% decrease. The influence of NF1 and MLTF seemed to be marginal in this system. The proximal 5'-flanking sequence of the L-PK gene therefore appears to function in vitro as an efficient liver-specific promoter which requires the binding of the liver factor HNF1 and which is also stimulated by the binding of another liver-specific factor, LF-A1.

scholarly journals PGC-1-Related Coactivator, a Novel, Serum-Inducible Coactivator of Nuclear Respiratory Factor 1-Dependent Transcription in Mammalian Cells

2001 ◽  
Vol 21 (11) ◽  
pp. 3738-3749 ◽  
Author(s):  
Ulf Andersson ◽  
Richard C. Scarpulla
Keyword(s):  
Transcriptional Activation ◽  
Mammalian Cells ◽  
Laser Scanning ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Peroxisome Proliferator ◽  
Nuclear Respiratory Factor ◽  
Nuclear Respiratory Factor 1 ◽  
Cell Extracts

ABSTRACT The thermogenic peroxisome proliferator-activated receptor γ (PPAR-γ) coactivator 1 (PGC-1) has previously been shown to activate mitochondrial biogenesis in part through a direct interaction with nuclear respiratory factor 1 (NRF-1). In order to identify related coactivators that act through NRF-1, we searched the databases for sequences with similarities to PGC-1. Here, we describe the first characterization of a 177-kDa transcriptional coactivator, designated PGC-1-related coactivator (PRC). PRC is ubiquitously expressed in murine and human tissues and cell lines; but unlike PGC-1, PRC was not dramatically up-regulated during thermogenesis in brown fat. However, its expression was down-regulated in quiescent BALB/3T3 cells and was rapidly induced by reintroduction of serum, conditions where PGC-1 was not detected. PRC activated NRF-1-dependent promoters in a manner similar to that observed for PGC-1. Moreover, NRF-1 was immunoprecipitated from cell extracts by antibodies directed against PRC, and both proteins were colocalized to the nucleoplasm by confocal laser scanning microscopy. PRC interacts in vitro with the NRF-1 DNA binding domain through two distinct recognition motifs that are separated by an unstructured proline-rich region. PRC also contains a potent transcriptional activation domain in its amino terminus adjacent to an LXXLL motif. The spatial arrangement of these functional domains coincides with those found in PGC-1, supporting the conclusion that PRC and PGC-1 are structurally and functionally related. We conclude that PRC is a functional relative of PGC-1 that operates through NRF-1 and possibly other activators in response to proliferative signals.

Novel genetic association between ulcerative colitis and the anti-inflammatory cytokine interleukin-1 receptor antagonist

1994 ◽  
Vol 106 (3) ◽  
pp. 637-642 ◽  
Author(s):  
John C. Mansfield ◽  
Hazel Holden ◽  
Joanna K. Tarlow ◽  
Francesco S. Di Giovine ◽  
Tarra L. McDowell ◽  
...  
Keyword(s):  
Ulcerative Colitis ◽  
Receptor Antagonist ◽  
Genetic Association ◽  
Inflammatory Cytokine ◽  
Interleukin 1 ◽  
Interleukin 1 Receptor Antagonist ◽  
Anti Inflammatory ◽  
Anti Inflammatory Cytokine

scholarly journals In vitro expression of long and short ovine prolactin receptors: activation of Jak2/STAT5 pathway is not sufficient to account for prolactin signal transduction to the ovine beta-lactoglobulin gene promoter

1999 ◽  
Vol 23 (2) ◽  
pp. 125-136 ◽  
Author(s):  
C Bignon ◽  
N Daniel ◽  
L Belair ◽  
J Djiane
Keyword(s):  
Tyrosine Phosphorylation ◽  
Transcriptional Activation ◽  
Target Genes ◽  
Gene Promoter ◽  
Janus Kinase ◽  
Prolactin Receptors ◽  
Milk Protein Gene ◽  
Prolactin Signaling ◽  
Beta Lactoglobulin

The recent finding that sheep had long (l-oPRLR) and short (s-oPRLR) prolactin receptors provided new tools to further explore prolactin signaling to target genes. Here we used CHO cells transfected with l-oPRLR or s-oPRLR cDNAs to compare the activation of known key steps of prolactin signaling by the two receptors. We found that prolactin stimulated l-oPRLR tyrosine phosphorylation, although it lacked the last tyrosine residue found in other long prolactin receptors. In addition, l-oPRLR and s-oPRLR both responded to prolactin stimulation by (1) Janus kinase 2 (Jak2) tyrosine phosphorylation, (2) DNA-binding activation of signal transducer and activator of transcription 5 (STAT5), (3) stimulation of transcription from a promoter made of six repeats of STAT5-responsive sequence. However, although it contains STAT5-binding consensus sequences, the ovine beta-lactoglobulin promoter (-4000 to +40) was transactivated by l-oPRLR, but not by s-oPRLR. Taken together, our results indicate that activation of Jak2/STAT5 pathway alone is not sufficient to account for prolactin-induced transcription of this milk protein gene, and that sequences of its promoter, other than STAT5-specific sequences, account for the opposite transcriptional activation capabilities of l-oPRLR and s-oPRLR.

scholarly journals A Protective Role for Interleukin-1 Signaling during Mouse Adenovirus Type 1-Induced Encephalitis

2016 ◽  
Vol 91 (4) ◽  
Author(s):  
Luiza A. Castro-Jorge ◽  
Carla D. Pretto ◽  
Asa B. Smith ◽  
Oded Foreman ◽  
Kelly E. Carnahan ◽  
...  
Keyword(s):  
Inflammatory Cytokine ◽  
Time Course ◽  
Interleukin 1 ◽  
Adenovirus Type ◽  
Mouse Strains ◽  
Complex Nature ◽  
Type I ◽  
Viral Loads ◽  
The Brain

ABSTRACT Interleukin-1β (IL-1β), an inflammatory cytokine and IL-1 receptor ligand, has diverse activities in the brain. We examined whether IL-1 signaling contributes to the encephalitis observed in mouse adenovirus type 1 (MAV-1) infection, using mice lacking the IL-1 receptor (Il1r1 −/− mice). Il1r1 −/− mice demonstrated reduced survival, greater disruption of the blood-brain barrier (BBB), higher brain viral loads, and higher brain inflammatory cytokine and chemokine levels than control C57BL/6J mice. We also examined infections of mice defective in IL-1β production (Pycard −/− mice) and mice defective in trafficking of Toll-like receptors to the endosome (Unc93b1 −/− mice). Pycard −/− and Unc93b1 −/− mice showed lower survival (similar to Il1r1 −/− mice) than control mice but, unlike Il1r1 −/− mice, did not have increased brain viral loads or BBB disruption. Based on the brain cytokine levels, MAV-1-infected Unc93b1 −/− mice had a very different inflammatory profile from infected Il1r1 −/− and Pycard −/− mice. Histological examination demonstrated pathological findings consistent with encephalitis in control and knockout mice; however, intranuclear viral inclusions were seen only in Il1r1 −/− mice. A time course of infection of control and Il1r1 −/− mice evaluating the kinetics of viral replication and cytokine production revealed differences between the mouse strains primarily at 7 to 8 days after infection, when mice began succumbing to MAV-1 infection. In the absence of IL-1 signaling, we noted an increase in the transcription of type I interferon (IFN)-stimulated genes. Together, these results indicate that IL-1 signaling is important during MAV-1 infection and suggest that, in its absence, increased IFN-β signaling may result in increased neuroinflammation. IMPORTANCE The investigation of encephalitis pathogenesis produced by different viruses is needed to characterize virus and host-specific factors that contribute to disease. MAV-1 produces viral encephalitis in its natural host, providing a good model for studying factors involved in encephalitis development. We investigated the role of IL-1 signaling during MAV-1-induced encephalitis. Unexpectedly, the lack of IL-1 signaling increased the mortality and inflammation in mice infected with MAV-1. Also, there was an increase in the transcription of type I IFN-stimulated genes that correlated with the observed increased mortality and inflammation. The findings highlight the complex nature of encephalitis and suggests that IL-1 has a protective effect for the development of MAV-1-induced encephalitis.

scholarly journals Postrecruitment Function of Yeast Med6 Protein during the Transcriptional Activation by Mediator Complex

2018 ◽  
Vol 2018 ◽  
pp. 1-9
Author(s):  
Gwang Sik Kim ◽  
Young Chul Lee
Keyword(s):  
Gene Expression ◽  
Reporter Gene ◽  
Transcriptional Activation ◽  
Reporter Gene Expression ◽  
Temperature Sensitive ◽  
Internal Deletion ◽  
Cell Extracts ◽  
Evolutionarily Conserved

Med6 protein (Med6p) is a hallmark component of evolutionarily conserved Mediator complexes, and the genuine role of Med6p in Mediator functions remains elusive. For the functional analysis ofSaccharomyces cerevisiaeMed6p (scMed6p), we generated a series of scMed6p mutants harboring a small internal deletion. Genetic analysis of these mutants revealed that three regions (amino acids 33–42 (Δ2), 125–134 (Δ5), and 157–166 (Δ6)) of scMed6p are required for cell viability and are located at highly conserved regions of Med6 homologs. Notably, the Med6p-Δ2 mutant was barely detectable in whole-cell extracts and purified Mediator, suggesting a loss of Mediator association and concurrent rapid degradation. Consistent with this, the recombinant forms of Med6p having these mutations partially (Δ2) restore or fail (Δ5 and Δ6) to restore in vitro transcriptional defects caused by temperature-sensitivemed6mutation. In an artificial recruitment assay, Mediator containing a LexA-fused wild-type Med6p or Med6p-Δ5 was recruited to thelexAoperator region with TBP and activated reporter gene expression. However, the recruitment of Mediator containing LexA-Med6p-Δ6 tolexAoperator region resulted in neither TBP recruitment nor reporter gene expression. This result demonstrates a pivotal role of Med6p in the postrecruitment function of Mediator, which is essential for transcriptional activation by Mediator.

scholarly journals ADR1-Mediated Transcriptional Activation Requires the Presence of an Intact TFIID Complex

1998 ◽  
Vol 18 (10) ◽  
pp. 5861-5867 ◽  
Author(s):  
Philip B. Komarnitsky ◽  
Edward R. Klebanow ◽  
P. Anthony Weil ◽  
Clyde L. Denis
Keyword(s):  
Transcriptional Activation ◽  
Yeast Cells ◽  
Binding Studies ◽  
Whole Cell ◽  
Transactivation Domains ◽  
Cell Extracts ◽  
Genes Expression ◽  
Tfiid Complex

ABSTRACT The yeast transcriptional activator ADR1, which is required forADH2 and other genes’ expression, contains four transactivation domains (TADs). While previous studies have shown that these TADs act through GCN5 and ADA2, and presumably TFIIB, other factors are likely to be involved in ADR1 function. In this study, we addressed the question of whether TFIID is also required for ADR1 action. In vitro binding studies indicated that TADI of ADR1 was able to retain TAFII90 from yeast extracts and TADII could retain TBP and TAFII130/145. TADIV, however, was capable of retaining multiple TAFIIs, suggesting that TADIV was binding TFIID from yeast whole-cell extracts. The ability of TADIV truncation derivatives to interact with TFIID correlated with their transcription activation potential in vivo. In addition, the ability of LexA-ADR1-TADIV to activate transcription in vivo was compromised by a mutation in TAFII130/145. ADR1 was found to associate in vivo with TFIID in that immunoprecipitation of either TAFII90 or TBP from yeast whole-cell extracts specifically coimmunoprecipitated ADR1. Most importantly, depletion of TAFII90 from yeast cells dramatically reducedADH2 derepression. These results indicate that ADR1 physically associates with TFIID and that its ability to activate transcription requires an intact TFIID complex.

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