Lipopolysaccharide induces 5-lipoxygenase-activating protein gene expression in THP-1 cells via a NF-κB and C/EBP-mediated mechanism

2005 ◽  
Vol 288 (5) ◽  
pp. C1125-C1133 ◽  
Author(s):  
Kenneth J. Serio ◽  
K. Veera Reddy ◽  
Timothy D. Bigby
Keyword(s):  
Gene Expression ◽  
Chromatin Immunoprecipitation ◽  
Protein C ◽  
Protein Gene ◽  
Prolonged Exposure ◽  
Human Monocyte ◽  
Dominant Negative ◽  
Mononuclear Phagocytes ◽  
Nuclear Extracts ◽  
Enhancer Binding Protein

We examined induced expression of the 5-lipoxygenase-activating protein (FLAP), which is critical for leukotriene synthesis in mononuclear phagocytes. Prolonged exposure to the bacterial component, lipopolysaccharide (LPS), increased FLAP gene transcription, mRNA expression, and protein expression in the human monocyte-like THP-1 cell line. Activation and inhibition of the NF-κB pathway modulated LPS induction of FLAP gene expression. An NF-κB-mediated mechanism of action was supported by overexpression of dominant-negative IκBα and p50/p65 proteins. EMSA/supershift and DNase I footprint analyses revealed that p50 binds to an NF-κB site located in the proximal FLAP promoter, while chromatin immunoprecipitation assays demonstrated that LPS induced binding of p50 but not of p65. Moreover, EMSA/supershift analyses demonstrated that LPS induced time-dependent binding of THP-1 nuclear extracts (containing p50) to this promoter region. Mutation of the NF-κB site decreased basal promoter activity and abolished the p50- and p65-associated induction. EMSA/supershift analyses also demonstrated that LPS induced binding of THP-1 nuclear extracts [containing CCAAT/enhancer binding protein (C/EBP)-α, -δ, and -ε] to a C/EBP site located adjacent to the NF-κB site in the FLAP promoter. We conclude that LPS enhances FLAP gene expression via both NF-κB- and C/EBP-mediated transcriptional mechanisms in mononuclear phagocytes.

scholarly journals Epigenetic Changes of the Cyp11a1 Promoter Region in Granulosa Cells Undergoing Luteinization During Ovulation in Female Rats

Endocrinology ◽  
2016 ◽  
Vol 157 (9) ◽  
pp. 3344-3354 ◽  
Author(s):  
Maki Okada ◽  
Lifa Lee ◽  
Ryo Maekawa ◽  
Shun Sato ◽  
Takuya Kajimura ◽  
...  
Keyword(s):  
Gene Expression ◽  
Dna Methylation ◽  
Granulosa Cells ◽  
Chromatin Immunoprecipitation ◽  
Promoter Region ◽  
Binding Protein ◽  
Female Rats ◽  
Proximal Promoter ◽  
Lh Surge ◽  
Enhancer Binding Protein

The ovulatory LH surge induces rapid up-regulation of Cyp11a1 in granulosa cells (GCs) undergoing luteinization during ovulation. This study investigated in vivo whether epigenetic controls including histone modifications and DNA methylation in the promoter region are associated with the rapid increase of Cyp11a1 gene expression after LH surge. GCs were obtained from rats treated with equine chorionic gonadotropin (CG) before (0 h) and 4 h and 12 h after human (h)CG injection. Cyp11a1 mRNA levels rapidly increased after hCG injection, reached a peak at 4 hours, and then remained elevated until 12 hours. DNA methylation status in the Cyp11a1 proximal promoter region was hypomethylated and did not change at any of the observed times after hCG injection. Chromatin immunoprecipitation assays revealed that the levels of trimethylation of lysine 4 on histone H3 (H3K4me3), an active mark for transcription, increased, whereas the levels of H3K9me3 and H3K27me3, which are marks associated with repression of transcription, decreased in the Cyp11a1 proximal promoter after hCG injection. Chromatin condensation, which was analyzed using deoxyribonuclease I, decreased in the Cyp11a1 proximal promoter after hCG injection. Chromatin immunoprecipitation assays also showed that the binding activity of CAATT/enhancer-binding protein-β to the Cyp11a1 proximal promoter increased after hCG injection. Luciferase assays revealed that the CAATT/enhancer-binding protein-β-binding site had transcriptional activity and contributed to basal and cAMP-induced Cyp11a1 expression. These results suggest that changes in histone modification and chromatin structure in the Cyp11a1 proximal promoter are involved in the rapid increase of Cyp11a1 gene expression in GCs undergoing luteinization during ovulation.

cAMP activation of phosphoenolpyruvate carboxykinase transcription in renal LLC-PK1-F+ cells

1996 ◽  
Vol 271 (2) ◽  
pp. F347-F355 ◽  
Author(s):  
X. Liu ◽  
N. P. Curthoys
Keyword(s):  
Transcriptional Activation ◽  
Protein Phosphatase ◽  
Protein C ◽  
Phosphoenolpyruvate Carboxykinase ◽  
Fold Increase ◽  
Dominant Negative ◽  
Expression Vectors ◽  
Activation Of Transcription ◽  
Enhancer Binding Protein ◽  
F Cells

Phosphoenolpyruvate carboxykinase (PCK) is a key regulatory enzyme in renal ammoniagenesis and gluconeogenesis. LLC-PK1-F+ cells are porcine renal proximal tubule-like cells that express significant levels of the cytosolic PCK. Treatment of subconfluent LLC-PK1-F+ cells with 0.1 mM 8-(4-chlorophenylthio)-adenosine 3',5'-cyclic monophosphate (CPT-cAMP) for 8 h causes a 21-fold increase in PCK mRNA. This response is very rapid and is not inhibited by 0.5 mM cycloheximide, indicating that ongoing protein synthesis is not required. Similarly, cells transfected with PCK(-490)CAT exhibit an 8- to 10-fold increase in chloramphenicol acetyltransferase (CAT) activity when treated with cAMP for 24 h. The addition of okadaic acid, a protein phosphatase inhibitor, both stimulated the CAT activity and potentiated the cAMP effect by twofold, suggesting that phosphorylation may contribute to the transcriptional activation. Assays using a series of PCK-CAT constructs containing specific deletions or block mutations established that the CRE-1 the P3(II) elements are required for the cAMP response. Cotransfection experiments using dominant negative expression vectors indicated that a CCAAT enhancer binding protein (C/EBP) transcription factor, and not CREB, mediates cAMP activation of transcription in LLC-PK1-F+ cells.

Growth-dependent inhibition of CCAAT enhancer-binding protein (C/EBP alpha) gene expression during hepatocyte proliferation in the regenerating liver and in culture

1992 ◽  
Vol 12 (6) ◽  
pp. 2553-2560
Author(s):  
D Mischoulon ◽  
B Rana ◽  
N L Bucher ◽  
S R Farmer
Keyword(s):  
Gene Expression ◽  
Mrna Expression ◽  
Protein C ◽  
Binding Protein ◽  
Regenerating Liver ◽  
Differentiated Cells ◽  
Enhancer Binding Protein ◽  
Ccaat Enhancer Binding Protein ◽  
Alpha Gene ◽  
Rat Tail

As an approach to understanding physiological mechanisms that control the proliferation of highly differentiated cells, we are addressing whether certain hepatic transcription factors participate in mechanisms that control the growth of hepatocytes. We have focused on CCAAT enhancer-binding protein (C/EBP alpha), a transcription factor which is highly abundant in normal liver and is considered to regulate expression of many genes, including some involved in energy metabolism (S. L. McKnight, M. D. Lane, and S. Gluecksohn-Walsh. Genes Dev. 3:2021-2024, 1989). Using Northern (RNA) blot analysis, we have examined the expression of C/EBP alpha mRNA during liver regeneration and in primary cultures of hepatocytes. C/EBP alpha mRNA levels decrease 60 to 80% within 1 to 3 h after partial hepatectomy as the cells move from G0 to G1 and decrease further when cells progress into S phase. Run-on transcription analysis is in agreement with the Northern blot data, thus suggesting that C/EBP alpha is transcriptionally regulated in regenerating liver. C/EBP alpha mRNA expression also decreases dramatically during the growth of freshly isolated normal hepatocytes cultured under conventional conditions (on dried rat tail collagen; stimulated to proliferate by epidermal growth factor [EGF] and insulin). Cultures of hepatocytes on rat tail collagen in the presence or absence of EGF clearly show that within 3 h, EGF depresses C/EBP alpha mRNA expression and that this effect is substantially greater by 4 h. Inhibition of protein synthesis in the liver by cycloheximide or in cultured hepatocytes by puromycin or cycloheximide effectively blocks the down-regulation of C/EBP alpha gene expression, apparently by stabilizing the normal rapid turnover of the C/EBP alpha mRNA (half-life of <2 h). This drop in C/EBP alpha gene expression in response to activation of hepatocyte growth is consistent with the proposal that C/EBP alpha has an antiproliferative role to play in highly differentiated cells (R. M. Umek, A. D. Friedman, and S. L. McKnight, Science 251: 288-292, 1991).

scholarly journals A Novel Functional Polymorphism in the CCAAT/Enhancer Binding Protein (C/EBP), Epsilon (CEBPE) Gene Promoter Influences Acute Lymphoblastic Leukemia Risk Via Interaction with IKZF1

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 489-489
Author(s):  
Kyle M. Walsh ◽  
Adam J. de Smith ◽  
Jianqiao Xiao ◽  
Marcus O. Muench ◽  
Marina Fomin ◽  
...  
Keyword(s):  
B Cell ◽  
Chromatin Immunoprecipitation ◽  
Protein C ◽  
Risk Allele ◽  
Lymphoblastic Leukemia ◽  
Gene Promoter ◽  
Expression Vectors ◽  
Genome Wide ◽  
Enhancer Binding Protein ◽  
Increased Risk

Abstract Purpose: We sought to identify and characterize the functional polymorphism in the CCAAT/enhancer binding protein (C/EBP), epsilon (CEBPE) gene, which is known to harbor a genetic risk modifier for childhood acute lymphoblastic leukemia (ALL). Background: Association between ALL risk and polymorphic variation at the CEBPE gene, a zinc-finger hematopoietic and apoptotic regulator expressed exclusively in myeloid cells, was established in a genome-wide association study and replicated in other populations. The actual functional genetic variation that actually contributes the leukemia risk is not known. The discovery and functional characterization of the functional variant may lead to risk stratification, prevention, and/or therapeutic modalities. Methods: Utilizing California Childhood Leukemia Study genome-wide SNP data from Hispanic subjects, we refine the genetic signal to a specific single nucleotide polymorphism (SNP) within the CEBPE gene promoter (rs2239635). This SNP was identified first using genetic imputation using 1000 Genomes data as a framework, and further validated in additional subjects using Taqman genotyping. Functional work to characterize the impact of the polymorphism was performed using prediction methods (database searches), chromatin immunoprecipitation assays from normal subjects heterozygous for rs2239635, and luciferase expression vectors carrying the SNP. Results: The minor allele of rs2239635 conferred a 1.87-fold increased risk of B-cell ALL (95% CI = 1.51-2.30; P = 6.2x10-9) and a 2.94-fold increased risk of high hyperdiploid B-cell ALL (95% CI = 2.12-4.06; P = 8.2x10-11), but only a 1.51-fold increased risk of non-hyperdiploid B-cell ALL (95% CI = 1.11-2.04; P = 8.8x10-3). Adjusting for rs2239635 attenuated all SNP associations in the region, including the original GWAS allele, strongly suggest that the GWAS signal in CCLS Hispanics is attributable to rs2239635. We also demonstrate, using published functional data and our own chromatin immunoprecipitation assays, that this SNP disrupts a canonical transcription factor binding site for Ikaros (IKZF1), an essential hematopoietic transcription factor. Using expression vectors, we found the polymorphic locus to have transcriptional repressive properties that are diminished in the presence of the ALL risk allele. Additionally, a heritable risk allele within IKZF1 interacts with rs2239635 in a multiplicative fashion (P = 0.02) in case-control association analyses. Discussion: In sum, evidence for a functional CEBPE polymorphism links two hematopoietic developmental transcriptional factors and highlights feedback control between them. The minor, risk-associated variant of CEBPE may alter the ability of IKZF1 to repress CEBPE expression in precursor B-cells, causing lineage confusion and enhanced pre-B cell leukemia risk. This risk primarily impacts risk of pediatric ALL that exhibits the high hyperdiploid phenotype. Disclosures No relevant conflicts of interest to declare.

scholarly journals KIT oncogene inhibition drives intratumoral macrophage M2 polarization

2013 ◽  
Vol 210 (13) ◽  
pp. 2873-2886 ◽  
Author(s):  
Michael J. Cavnar ◽  
Shan Zeng ◽  
Teresa S. Kim ◽  
Eric C. Sorenson ◽  
Lee M. Ocuin ◽  
...  
Keyword(s):  
Gene Expression ◽  
Protein C ◽  
Cancer Microenvironment ◽  
M2 Polarization ◽  
Enhancer Binding Protein ◽  
Activating Mutation ◽  
Similar Gene Expression Profile ◽  
And Function ◽  
Human Sarcoma ◽  
Similar Gene

Tumor-associated macrophages (TAMs) are a major component of the cancer microenvironment. Modulation of TAMs is under intense investigation because they are thought to be nearly always of the M2 subtype, which supports tumor growth. Gastrointestinal stromal tumor (GIST) is the most common human sarcoma and typically results from an activating mutation in the KIT oncogene. Using a spontaneous mouse model of GIST and 57 freshly procured human GISTs, we discovered that TAMs displayed an M1-like phenotype and function at baseline. In both mice and humans, the KIT oncoprotein inhibitor imatinib polarized TAMs to become M2-like, a process which involved TAM interaction with apoptotic tumor cells leading to the induction of CCAAT/enhancer binding protein (C/EBP) transcription factors. In human GISTs that eventually developed resistance to imatinib, TAMs reverted to an M1-like phenotype and had a similar gene expression profile as TAMs from untreated human GISTs. Therefore, TAM polarization depends on tumor cell oncogene activity and has important implications for immunotherapeutic strategies in human cancers.

scholarly journals Growth-dependent inhibition of CCAAT enhancer-binding protein (C/EBP alpha) gene expression during hepatocyte proliferation in the regenerating liver and in culture.

1992 ◽  
Vol 12 (6) ◽  
pp. 2553-2560 ◽  
Author(s):  
D Mischoulon ◽  
B Rana ◽  
N L Bucher ◽  
S R Farmer
Keyword(s):  
Gene Expression ◽  
Mrna Expression ◽  
Protein C ◽  
Binding Protein ◽  
Regenerating Liver ◽  
Differentiated Cells ◽  
Enhancer Binding Protein ◽  
Ccaat Enhancer Binding Protein ◽  
Alpha Gene ◽  
Rat Tail

As an approach to understanding physiological mechanisms that control the proliferation of highly differentiated cells, we are addressing whether certain hepatic transcription factors participate in mechanisms that control the growth of hepatocytes. We have focused on CCAAT enhancer-binding protein (C/EBP alpha), a transcription factor which is highly abundant in normal liver and is considered to regulate expression of many genes, including some involved in energy metabolism (S. L. McKnight, M. D. Lane, and S. Gluecksohn-Walsh. Genes Dev. 3:2021-2024, 1989). Using Northern (RNA) blot analysis, we have examined the expression of C/EBP alpha mRNA during liver regeneration and in primary cultures of hepatocytes. C/EBP alpha mRNA levels decrease 60 to 80% within 1 to 3 h after partial hepatectomy as the cells move from G0 to G1 and decrease further when cells progress into S phase. Run-on transcription analysis is in agreement with the Northern blot data, thus suggesting that C/EBP alpha is transcriptionally regulated in regenerating liver. C/EBP alpha mRNA expression also decreases dramatically during the growth of freshly isolated normal hepatocytes cultured under conventional conditions (on dried rat tail collagen; stimulated to proliferate by epidermal growth factor [EGF] and insulin). Cultures of hepatocytes on rat tail collagen in the presence or absence of EGF clearly show that within 3 h, EGF depresses C/EBP alpha mRNA expression and that this effect is substantially greater by 4 h. Inhibition of protein synthesis in the liver by cycloheximide or in cultured hepatocytes by puromycin or cycloheximide effectively blocks the down-regulation of C/EBP alpha gene expression, apparently by stabilizing the normal rapid turnover of the C/EBP alpha mRNA (half-life of <2 h). This drop in C/EBP alpha gene expression in response to activation of hepatocyte growth is consistent with the proposal that C/EBP alpha has an antiproliferative role to play in highly differentiated cells (R. M. Umek, A. D. Friedman, and S. L. McKnight, Science 251: 288-292, 1991).

CCAAT/Enhancer-Binding Protein   Gene Expression Is Mediated by APRF/STAT3

1997 ◽  
Vol 121 (4) ◽  
pp. 731-738 ◽  
Author(s):  
T. Yamada ◽  
K. Tobita ◽  
S. Osada ◽  
T. Nishihara ◽  
M. Imagawa
Keyword(s):  
Gene Expression ◽  
Protein Gene ◽  
Binding Protein ◽  
Enhancer Binding Protein ◽  
Ccaat Enhancer Binding Protein ◽  
Binding Protein Gene
Sign in / Sign up

Export Citation Format

Share Document