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Enhancer Binding Protein (enhancer + binding_protein)
Selected AbstractsHeregulin and forskolin-induced cyclin D3 expression in Schwann cells: Role of a CCAAT promoter element and CCAAT enhancer binding proteinGLIA, Issue 3 2004Luis Fuentealba Abstract Heregulin, a polypeptide growth factor, and forskolin, an adenylyl cyclase activator, synergistically stimulate expression of cyclin D3 and cell division in Schwann cells. Heregulin induces expression in Schwann cells of a luciferase reporter gene linked to the cyclin D3 promoter. Forskolin markedly augments reporter expression in the presence of heregulin. Deletion analysis identified several promoter sites that contribute to high-level reporter expression in heregulin- and forskolin-treated Schwann cells. A promoter fragment that contains 103 bp of 5,-flanking sequence produced significant reporter expression in heregulin- and forskolin-stimulated cells. Deletion of a consensus CCAAT site within this promoter fragment caused a nearly complete loss of reporter expression. Similar results were obtained when CCAAT site mutations were introduced into the promoter. Heregulin and forskolin increased steady-state levels of CCAAT/enhancer binding protein-, (C/EBP,) in Schwann cells. Mobility shift assays identified proteins in Schwann cell nuclear extracts that formed stable complexes with the cyclin D3 CCAAT promoter element and were disrupted by anti-C/EBP, antibody. Transfection of Schwann cells with C/EBP, cDNA increased cyclin D3 reporter expression. In contrast to these results, mutation of a cAMP response element in the cyclin D3 promoter had only a modest effect on heregulin- and forskolin-stimulated reporter expression. These findings demonstrate that C/EBP, plays a key role in the heregulin and cAMP-dependent regulation of cyclin D3 expression in Schwann cells. © 2003 Wiley-Liss, Inc. [source] MEK/ERK Signaling Controls Osmoregulation of Nucleus Pulposus Cells of the Intervertebral Disc by Transactivation of TonEBP/OREBP,JOURNAL OF BONE AND MINERAL RESEARCH, Issue 7 2007Tsung-Ting Tsai Abstract Earlier studies have shown that intervertebral disc cells express TonEBP, a transcription factor that permits adaptation to osmotic stress and regulates aggrecan gene expression. However, the mechanism of hyperosmotic activation of TonEBP in disc cells is not known. Results of this study show that hypertonic activation of ERK signaling regulates transactivation activity of TonEBP, modulating its function. Introduction: In an earlier report, we showed that tonicity enhancer binding protein (TonEBP) positively regulates aggrecan gene expression in disc cells, thereby autoregulating its osmotic environment. Although these studies indicated that the cells of the nucleus pulposus were optimally adapted to a hyperosmotic state, the mechanism by which the cells transduce the osmotic stress was not delineated. The primary goal of this study was to test the hypothesis that, in a hyperosmotic medium, the extracellular signal-regulated kinase (ERK) signaling pathway regulated TonEBP activity. Materials and Methods: Nucleus pulposus cells were maintained in isotonic or hypertonic media, and MAPK activation and TonEBP expression were analyzed. To study the role of MAPK in regulation of TonEBP function, gel shift and luciferase reporter assays were performed. ERK expression in cells was modulated by using expression plasmids or siRNA, and transactivation domain (TAD)-TonEBP activity was studied. Results: We found that hypertonicity resulted in phosphorylation and activation of ERK1/2 proteins and concomitant activation of C terminus TAD activity of ELK-1, a downstream transcription factor. In hypertonic media, treatment with ERK and p38 inhibitors resulted in downregulation of TonE promoter activity of TauT and HSP-70 and decreased binding of TonEBP to TonE motif. Similarly, forced expression of DN-ERK and DN-p38 in nucleus pulposus cells suppressed TauT and HSP-70 reporter gene activity. Finally, we noted that ERK was needed for transactivation of TonEBP. Expression of DN-ERK significantly suppressed, whereas, WT-ERK and CA-MEK1 enhanced, TAD activity of TonEBP. Experiments performed with HeLa cells indicated that the ERK signaling pathway also served a major role in regulating the osmotic response in nondiscal cells. Conclusions: Together, these studies showed that adaptation of the nucleus pulposus cells to their hyperosmotic milieu is dependent on activation of the ERK and p38- MAPK pathways acting through TonEBP and its target genes. [source] Extracellular matrix regulates alpha s1-casein gene expression in rabbit primary mammary cells and CCAAT enhancer binding protein (C/EBP) binding activityJOURNAL OF CELLULAR BIOCHEMISTRY, Issue 3 2001Geneviève Jolivet Abstract Previous studies have shown that both the signal transducer and activator of transcription 5 (STAT5) and the CCAAT enhancer binding proteins (C/EBPs) are involved in the regulation of casein gene expression by mammary epithelial cells. Prolactin (Prl) activation of STAT5 is necessary for casein gene expression. The extracellular matrix (ECM) regulates also casein gene expression. Here, we have investigated whether ECM regulates C/EBPs activity in primary rabbit mammary epithelial cells. Isolated primary mammary cells were cultured on plastic or on floating collagen I gel. Prolactin induced ,s 1-casein gene expression when cells were cultured on collagen but not on plastic. It is noteworthy that activated STAT5 was detected in both culture conditions. Several STAT5 isoforms (STAT5a, STAT5b, and other STAT5 related isoforms, some with lower molecular weight than the full-length STAT5a and STAT5b) were detected under the different culture conditions. However, their presence was not related to the expression of ,s 1-casein gene. The binding of nuclear factors to a C/EBP specific binding site and the protein level of C/EBP, differed in cells cultured on plastic or on collagen but these parameters were not modified by Prl. This suggests that C/EBP binding activity was regulated by ECM and not by Prl. Interestingly, these modifications were correlated to the expression of the ,s 1-casein gene. Hence, the activation of the ,s 1-casein gene expression depends on two independent signals, one delivered by Prl via the activation of STAT5, the other delivered by ECM via C/EBP. J. Cell. Biochem. 82:371,386, 2001. © 2001 Wiley-Liss, Inc. [source] The solution structure of ZNF593 from Homo sapiens reveals a zinc finger in a predominately unstructured proteinPROTEIN SCIENCE, Issue 3 2008Paulette L. Hayes Abstract Here, we report the solution structure of ZNF593, a protein identified in a functional study as a negative modulator of the DNA-binding activity of the Oct-2 transcription factor. ZNF593 contains a classic C2H2 zinc finger domain flanked by about 40 disordered residues on each terminus. Although the protein contains a high degree of intrinsic disorder, the structure of the zinc finger domain was resolved by NMR spectroscopy without a need for N- or C-terminal truncations. The tertiary structure of the zinc finger domain is composed of a ,-hairpin that positions the cysteine side chains for zinc coordination, followed by an atypical kinked ,-helix containing the two histidine side chain ligands. The structural topology of ZNF593 is similar to a fragment of the double-stranded RNA-binding protein Zfa and the C-terminal zinc finger of MBP-1, a human enhancer binding protein. The structure presented here will provide a guide for future functional studies of how ZNF593 negatively modulates the DNA-binding activity of Oct-2, a POU domain-containing transcription factor. Our work illustrates the unique capacity of NMR spectroscopy for structural analysis of folded domains in a predominantly disordered protein. [source] A sodium dodecyl sulfate,polyacrylamide gel electrophoresis,liquid chromatography tandem mass spectrometry analysis of bovine cartilage tissue response to mechanical compression injury and the inflammatory cytokines tumor necrosis factor , and interleukin-1,ARTHRITIS & RHEUMATISM, Issue 2 2008Anna L. Stevens Objective To compare the response of chondrocytes and cartilage matrix to injurious mechanical compression and treatment with interleukin-1, (IL-1,) and tumor necrosis factor , (TNF,), by characterizing proteins lost to the medium from cartilage explant culture. Methods Cartilage explants from young bovine stifle joints were treated with 10 ng/ml of IL-1, or 100 ng/ml of TNF, or were subjected to uniaxial, radially-unconfined injurious compression (50% strain; 100%/second strain rate) and were then cultured for 5 days. Pooled media were subjected to gel-based separation (sodium dodecyl sulfate,polyacrylamide gel electrophoresis) and analysis by liquid chromatography tandem mass spectrometry, and the data were analyzed by Spectrum Mill proteomics software, focusing on protein identification, expression levels, and matrix protein proteolysis. Results More than 250 proteins were detected, including extracellular matrix (ECM) structural proteins, pericellular matrix proteins important in cell,cell interactions, and novel cartilage proteins CD109, platelet-derived growth factor receptor,like, angiopoietin-like 7, and adipocyte enhancer binding protein 1. IL-1, and TNF, caused increased release of chitinase 3,like protein 1 (CHI3L1), CHI3L2, complement factor B, matrix metalloproteinase 3, ECM-1, haptoglobin, serum amyloid A3, and clusterin. Injurious compression caused the release of intracellular proteins, including Grp58, Grp78, ,4-actinin, pyruvate kinase, and vimentin. Injurious compression also caused increased release and evidence of proteolysis of type VI collagen subunits, cartilage oligomeric matrix protein, and fibronectin. Conclusion Overload compression injury caused a loss of cartilage integrity, including matrix damage and cell membrane disruption, which likely occurred through strain-induced mechanical disruption of cells and matrix. IL-1, and TNF, caused the release of proteins associated with an innate immune and stress response by the chondrocytes, which may play a role in host defense against pathogens or may protect cells against stress-induced damage. [source] Molecular markers associated with lymph node metastasis in pancreatic ductal adenocarcinoma by genome-wide expression profilingCANCER SCIENCE, Issue 1 2010Seiko Hirono Lymph node metastasis (LNM) is the most important prognostic factor in patients undergoing surgical resection of pancreatic ductal adenocarcinoma (PDAC). In this study, we aimed to identify molecular markers associated with LNM in PDAC using genome-wide expression profiling. In this study, laser microdissection and genome-wide transcriptional profiling were used to identify genes that were differentially expressed between PDAC cells with and without LNM obtained from 20 patients with PDAC. Immunohistochemical staining was used to confirm the clinical significance of these markers in an additional validation set of 43 patients. In the results, microarray profiling identified 46 genes that were differently expressed between PDAC with and without LNM with certain significance. Four of these biomarkers were validated by immunohistochemical staining for association with LNM in PDAC in an additional validation set of patients. In 63 patients with PDAC, significant LNM predictors in PDAC elucidated from multivariate analysis were low expression of activating enhancer binding protein 2 (AP2,) (P = 0.012) and high expression of mucin 17 (MUC17) (P = 0.0192). Furthermore, multivariate analysis revealed that AP2, -low expression and MUC17 -high expression are independent prognostic factors for poor overall survival (P = 0.0012, 0.0001, respectively). In conclusion, AP2, and MUC17 were independent markers associated with LNM of PDAC. These two markers were also associated with survival in patients with resected PDAC. We demonstrate that AP2, and MUC17 may serve as potential prognostic molecular markers for LNM in patients with PDAC. (Cancer Sci 2009) [source] Extracellular matrix regulates alpha s1-casein gene expression in rabbit primary mammary cells and CCAAT enhancer binding protein (C/EBP) binding activityJOURNAL OF CELLULAR BIOCHEMISTRY, Issue 3 2001Geneviève Jolivet Abstract Previous studies have shown that both the signal transducer and activator of transcription 5 (STAT5) and the CCAAT enhancer binding proteins (C/EBPs) are involved in the regulation of casein gene expression by mammary epithelial cells. Prolactin (Prl) activation of STAT5 is necessary for casein gene expression. The extracellular matrix (ECM) regulates also casein gene expression. Here, we have investigated whether ECM regulates C/EBPs activity in primary rabbit mammary epithelial cells. Isolated primary mammary cells were cultured on plastic or on floating collagen I gel. Prolactin induced ,s 1-casein gene expression when cells were cultured on collagen but not on plastic. It is noteworthy that activated STAT5 was detected in both culture conditions. Several STAT5 isoforms (STAT5a, STAT5b, and other STAT5 related isoforms, some with lower molecular weight than the full-length STAT5a and STAT5b) were detected under the different culture conditions. However, their presence was not related to the expression of ,s 1-casein gene. The binding of nuclear factors to a C/EBP specific binding site and the protein level of C/EBP, differed in cells cultured on plastic or on collagen but these parameters were not modified by Prl. This suggests that C/EBP binding activity was regulated by ECM and not by Prl. Interestingly, these modifications were correlated to the expression of the ,s 1-casein gene. Hence, the activation of the ,s 1-casein gene expression depends on two independent signals, one delivered by Prl via the activation of STAT5, the other delivered by ECM via C/EBP. J. Cell. Biochem. 82:371,386, 2001. © 2001 Wiley-Liss, Inc. [source] | |||||||||||||