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Transcription Factor Binding (transcription + factor_binding)
Terms modified by Transcription Factor Binding Selected AbstractsCyclooxygenase-2 Expression Induced by Photofrin Photodynamic Therapy Involves the p38 MAPK Pathway,PHOTOCHEMISTRY & PHOTOBIOLOGY, Issue 2 2008Marian Luna Photodynamic therapy (PDT), using the porphyrin photosensitizer Photofrin (PH), is approved for the clinical treatment of solid tumors. In addition to the direct cytotoxic responses of PH,PDT-mediated oxidative stress, this procedure also induces expression of angiogenic and prosurvival molecules including cyclooxygenase-2 (COX-2). In vivo treatment efficacy is improved when PH-PDT is combined with inhibitors of COX-2. In the current study we evaluated the signaling pathways involved with PH,PDT-mediated COX-2 expression in a mouse fibrosarcoma cell line. COX-2 promoter reporter constructs with mutated transcription elements documented that the nuclear factor kappa B (NF,B) element, cyclic-AMP response element 2 (CRE-2), CCAAT/enhancer binding protein (C/EBP) element and activator binding protein-1 (AP-1) element were responsive to PH-PDT. Transcription factor binding assays demonstrated that nuclear protein binding to NF,B, CRE-2, c-fos and c-jun elements were elevated following PH-PDT. Kinase phosphorylation upstream of COX-2 expression was also examined following PH-PDT. Stress-activated protein kinase/c-Jun N-terminal kinase (SAPK/JNK) and c-Jun were phosphorylated following PH-PDT but the SAPK/JNK inhibitor SP600125 failed to attenuate COX-2 expression. In contrast, p38 mitogen-activated protein kinase (MAPK), which activates CRE-2 binding, was phosphorylated following PH-PDT and inhibitors of p38 MAPK, SB203580 and SB202190, decreased PH,PDT-induced COX-2 expression at both the mRNA and protein levels. Extracellular signal-regulated kinase (ERK1/2) phosphorylation, which also increases CRE-2 binding activity, was initially high in untreated cells, decreased immediately following PH-PDT and then rapidly increased. MEK1/2 is immediately upstream of ERK1/2 and the MEK1 inhibitor PD98059 failed to attenuate COX-2 expression while the MEK1/2 inhibitor U0126 induced a slight decrease in COX-2 expression. The NF,B inhibitor SN50 failed to reduce COX-2 expression. These results demonstrate that multiple protein kinase cascades can be activated by oxidative stress and that the p38 MAPK signaling pathway and CRE-2 binding are involved in COX-2 expression following PH-PDT. [source] Structural and functional differences between the promoters of independently expressed killer cell Ig-like receptorsEUROPEAN JOURNAL OF IMMUNOLOGY, Issue 7 2005Bergen, Jeroen van Abstract Killer Ig-like receptors (KIR) are important for the recognition and elimination of diseased cells by human NK cells. Myeloid leukemia patients given a hematopoietic stem cell transplantation, for example, benefit from KIR-mediated NK alloreactivity directed against the leukemia cells. To establish an effective NK cell repertoire, most KIR genes are expressed stochastically, independently of the others. However, the sequences upstream of the coding regions of these KIR genes are highly homologous to the recently identified KIR3DL1 promoter (91.1,99.6% sequence identity), suggesting that they are regulated by similar if not identical mechanisms of transcriptional activation. We investigated the effects of small sequence differences between the KIR3DL1 promoter and other KIR promoters on transcription factor binding and promoter activity. Surprisingly, electrophoretic mobility shift assays and promoter-reporter assays revealed significant structural and functional differences in the cis-acting elements of these highly homologous KIR promoters, suggesting a key role for transcription factors in independent control of expression of specific KIR loci. Thus, the KIR repertoire may be shaped by a combination of both gene-specific and stochastic mechanisms. [source] Transcription of mammalian cytochrome c oxidase subunit IV-2 is controlled by a novel conserved oxygen responsive elementFEBS JOURNAL, Issue 21 2007Maik Hüttemann Subunit 4 of cytochrome c oxidase (CcO) is a nuclear-encoded regulatory subunit of the terminal complex of the mitochondrial electron transport chain. We have recently discovered an isoform of CcO 4 (CcO4-2) which is specific to lung and trachea, and is induced after birth. The role of CcO as the major cellular oxygen consumer, and the lung-specific expression of CcO4-2, led us to investigate CcO4-2 gene regulation. We cloned the CcO4-2 promoter regions of cow, rat and mouse and compared them with the human promoter. Promoter activity is localized within a 118-bp proximal region of the human promoter and is stimulated by hypoxia, reaching a maximum (threefold) under 4% oxygen compared with normoxia. CcO4-2 oxygen responsiveness was assigned by mutagenesis to a novel promoter element (5,-GGACGTTCCCACG-3,) that lies within a 24-bp region that is 79% conserved in all four species. This element is able to bind protein, and competition experiments revealed that, within the element, the four core bases 5,-TCNCA-3, are obligatory for transcription factor binding. CcO isolated from lung showed a 2.5-fold increased maximal turnover compared with liver CcO. We propose that CcO4-2 expression in highly oxygenated lung and trachea protects these tissues from oxidative damage by accelerating the last step in the electron transport chain, leading to a decrease in available electrons for free radical formation. [source] A survey of H2 gene sequences, including new wild-derived genesINTERNATIONAL JOURNAL OF IMMUNOGENETICS, Issue 1 2007N. A. Mitchison Summary A comprehensive collection of mouse major histocompatibility complex (MHC) promoter and exon 2 sequences is here presented and analysed. It covers the three best known class II genes and one class I gene, and includes new wild mouse sequences from the ,w' back-cross strains and from the Jackson collection. All sequences are in GenBank, and the new exon sequences largely confirm previous typing by serology and immune function. As in human leucocyte antigen (HLA), the overall nucleotide diversity is higher in the class II genes, in keeping with their more diverse function. Diversity along the promoters is highest in the region of known transcription factor binding, most notably in and around the CRE and rCAAT sequences. This distribution parallels that of maximum single nucleotide polymorphism impact previously obtained with reporter constructs. Taking into account the low nucleotide diversity of the CIITA promoter, we conclude that MHC promoters are likely to have diversified through co-evolution with their exons, while themselves also directly subject to natural selection. The H2Ebp alleles form a distinct group, associated with their lack of the recombination hot spot located between exon 2 and exon 3. The collection is expected to prove useful in guiding functional and evolutionary studies. [source] Programming the genome in embryonic and somatic stem cellsJOURNAL OF CELLULAR AND MOLECULAR MEDICINE, Issue 4 2007Philippe Collas ,,Introduction ,,Epigenetic makeup of embryonic stem cells: keeping chromatin loose -,DNA methylation and gene expression -,CpG methylation profiles in mouse ESCs -,CpG methylation patterns in human ESCs -,Both active and inactive histone modification marks on developmentally regulated genes in ESCs suggest transcriptional activation potential -,A regulatory role of histone H1 in gene expression in embryonic stem cells? -,Polycomb group proteins impose a transcriptional brake on lineage-priming genes ,,The epigenetic makeup of mesenchymal stem cells reflects restricted differentiation potential -,CpG methylation patterns on lineage-specific promoters in adipose stem cells -,CpG content affects the relationship between promoter DNA methylation and transcriptional activity -,Bivalent histone modifications on potentially active genes? ,,Linking DNA methylation to histone modifications, chromatin packaging and (re)organization of the nuclear compartment ,,Perspectives: towards remodelling the stem cell epigenome? Abstract In opposition to terminally differentiated cells, stem cells can self-renew and give rise to multiple cell types. Embryonic stem cells retain the ability of the inner cell mass of blastocysts to differentiate into all cell types of the body and have acquired in culture unlimited self-renewal capacity. Somatic stem cells are found in many adult tissues, have an extensive but finite lifespan and can differentiate into a more restricted array of cell types. A growing body of evidence indicates that multi-lineage differentiation ability of stem cells can be defined by the potential for expression of lineage-specification genes. Gene expression, or as emphasized here, potential for gene expression, is largely controlled by epigenetic modifications of DNA and chromatin on genomic regulatory and coding regions. These modifications modulate chromatin organization not only on specific genes but also at the level of the whole nucleus; they can also affect timing of DNA replication. This review highlights how mechanisms by which genes are poised for transcription in undifferentiated stem cells are being uncovered through primarily the mapping of DNA methylation, histone modifications and transcription factor binding throughout the genome. The combinatorial association of epigenetic marks on developmentally regulated and lineage-specifying genes in undifferentiated cells seems to define a pluripotent state. [source] Aging-dependent upregulation of IL-23p19 gene expression in dendritic cells is associated with differential transcription factor binding and histone modificationsAGING CELL, Issue 5 2009Rabab El Mezayen Summary Age-associated changes in immune response increase the risk of infection and promote inflammation and autoimmunity in older adults. The newly discovered cytokine IL-23 contributes to the maintenance and expansion of Th-17 cells, which promote proinflammatory responses. Our preliminary findings suggested that Th-17 responses are increased in aged mice. IL-23 consists of p40 and p19 subunits. Expression of the p19 subunit is regulated at the transcriptional level by NF-,B p65 and c-Rel transcription factors. Using bone-marrow-derived dendritic cells (DCs) from C57BL/6 mice, we show that IL-23 protein production and p19 subunit mRNA levels are significantly increased in DCs from aged mice after activation with TLR ligands (LPS + R848) when compared with DCs of young adult mice. We found that the increase in p19 expression in aged cells is associated with chromatin remodeling characterized by di- and tri-methylation of histone H3K4 and binding of mainly c-Rel at the p19 promoter. In young DCs, the promoter is tri-methylated only at H3K4 and bound by both p65 and c-Rel. C-Rel knockdown restores p65 binding in aged cells but does not activate p19 expression, suggesting that c-Rel is critical for p19 expression. In addition, p65 knockdown significantly increases c-Rel binding and p19 expression in young DCs to levels close to those detected in old cells. Furthermore, the decrease in p65 binding at the p19 promoter in old DCs was specific to the p19 gene since p65 binding to the IL-12p40 promoter was not significantly different between old and young DCs. Our results demonstrate that selective changes in H3K4 methylation, and c-Rel and p65 binding at the p19 promoter occur in DCs and contribute to the upregulation of the p19 subunit expression and IL-23 protein production observed in aged mice. This suggests epigenetic and transcriptional mechanisms contribute to dysregulated inflammatory and autoimmune responses associated with aging. [source] Phage display identifies novel peptides that bind extracellular-regulated protein kinase 2 to compete with transcription factor binding,JOURNAL OF PHYSICAL ORGANIC CHEMISTRY, Issue 6-7 2004Mark A. Rainey Abstract Extracellular-regulated protein kinase 2 (ERK2) is a serine/threonine-specific protein kinase capable of phosphorylating multiple protein substrates within a cell. In an attempt to identify novel peptides that bind and inhibit the function of an active form of ERK2, phage display was carried out using a disulfide-constrained peptide library (X2CX14CX2). Several phage clones were identified by an enzyme-linked immunosorbent assay (ELISA) that competed with both a protein substrate and adenosine triphosphate (ATP) for immobilized ERK2. A chemically synthesized peptide derived from these experiments, NH2 -KKKIRCIRGWTKDIRTLADSCQY-COOH, inhibited ERK2 phosphorylation of the protein substrate Ets,138, exhibiting competitive and mixed inhibition towards Ets,138 and MgATP2,, respectively. Surprisingly, the same peptide displayed equally potent inhibition towards the phosphorylation of ATF2 by p38 MAPK,, another MAP kinase that has ,46% sequence similarity to ERK2. This study indicates that active ERK2 can be targeted by phage display to find novel antagonists to kinase function and suggests that protein-binding sites within the MAPK family may contain conserved features that render them susceptible to ligand binding. Copyright © 2004 John Wiley & Sons, Ltd. [source] Hypoxia and glucocorticoid signaling converge to regulate macrophage migration inhibitory factor gene expressionARTHRITIS & RHEUMATISM, Issue 8 2009Laura M. Elsby Objective Macrophage migration inhibitory factor (MIF) is a proinflammatory mediator involved in the pathogenesis of rheumatoid arthritis. This study was undertaken to identify the MIF promoter elements responsible for regulating gene expression. Methods Luciferase reporter gene assays were used to identify the MIF promoter sequence responsible for basal activity. Bioinformatic analysis was used to predict transcription factor binding sites, and electrophoretic mobility shift assay (EMSA) was used to demonstrate transcription factor binding. Chromatin immunoprecipitation (ChIP) was used to demonstrate transcription factor loading on the MIF promoter. Results We identified the minimal promoter sequence required for basal MIF promoter activity that was also capable of conferring glucocorticoid-dependent inhibition in a T lymphocyte model cell line. Deletion studies and EMSA revealed 2 elements in the MIF promoter that were responsible for basal promoter activity. The 5, element binds CREB/activating transcription factor 1, and the 3, element is a functional hypoxia-responsive element binding hypoxia-inducible factor 1,. Further studies demonstrated that the cis elements are both required for glucocorticoid-dependent inhibition. ChIP demonstrated glucocorticoid-dependent recruitment of glucocorticoid receptor , to the MIF promoter in lymphocytes within 1 hour of treatment and a concomitant decrease in acetylated histone H3. Conclusion Our findings indicate that hypoxia and glucocorticoid signaling converge on a single element regulating MIF; this regulatory unit is a potential interacting node for microenvironment sensing of oxygen tension and glucocorticoid action in foci of inflammation. [source] Mediation of interleukin-1,,induced transforming growth factor ,1 expression by activator protein 4 transcription factor in primary cultures of bovine articular chondrocytes: Possible cooperation with activator protein 1ARTHRITIS & RHEUMATISM, Issue 6 2003R. Andriamanalijaona Objective Interleukin-1 (IL-1) and transforming growth factor ,1 (TGF,1) play major roles in osteoarticular diseases, exerting opposite effects on both the catabolism and anabolism of cartilage matrix. Previous findings suggest that IL-1 and TGF,1 could function in a feedback interaction. However, the effect exerted by IL-1 on expression of TGF, by articular chondrocytes is, so far, poorly understood. The present study was carried out to determine the influence of IL-1, on the expression of TGF,1 by bovine articular chondrocytes (BACs) in primary culture. Methods BAC primary cultures were treated with IL-1,, and TGF,1 messenger RNA (mRNA) steady-state levels and protein expression were measured by real-time reverse transcription,polymerase chain reaction and enzyme-linked immunosorbent assay, respectively. Transient transfection of TGF,1 gene promoter constructs was performed to delineate the DNA sequences that mediate the IL-1, effect. Electrophoretic mobility shift assays (EMSAs) and supershift analysis were used to characterize the transcription factors binding to these sequences. Results Cultured BACs responded to IL-1, exposure by exhibiting an increase of TGF,1 expression at both the mRNA and protein levels. The effect was found to be mediated by a major 80-bp sequence located between ,732 and ,652 upstream of the transcription initiation site. EMSA and supershift analysis revealed that the transcription factors activator protein 4 (AP-4) and AP-1 specifically bound to the ,720/,696 part of this sequence under IL-1, treatment. Overexpression of AP-4 in the BAC cultures resulted in stimulation of the transcriptional activity of the ,732/+11 TGF,1 promoter construct through the same IL-1,,responsive element. Conclusion IL-1, induces an increase of TGF,1 in articular chondrocytes through activation of AP-4 and AP-1 binding to the TGF,1 gene promoter. These findings may help us understand the role of IL-1, in the disease process. Notwithstanding its deleterious effect on cartilage, IL-1 could initiate the repair response displayed by injured cartilage in the early stages of osteoarthritis through its ability to enhance TGF,1 expression by local chondrocytes. [source] | ||||||||||||||||