Abstract 191: NF-?B p65 Methylation by Protein Arginine Methyltransferase 5 (PRMT5) is Necessary for the Transcriptional Induction of CXCL10 by TNF-a

2013 ◽  
Vol 33 (suppl_1) ◽  
Author(s):  
Daniel P Harris
Keyword(s):  
Endothelial Cells ◽  
Mrna Expression ◽  
Chromatin Immunoprecipitation ◽  
Proximal Promoter ◽  
Protein Arginine Methyltransferase ◽  
Arginine Methyltransferase ◽  
Expression Of Genes ◽  
Tnf Α ◽  
Affect Expression ◽  
Transcriptional Induction

TNF-α initiates the expression of genes involved in the recruitment, adhesion, and transmigration of leukocytes to sites of inflammation. Here, we report that the protein arginine methyltransferase PRMT5 is required for the transcriptional induction of the pro-inflammatory chemokine CXCL10 (IP-10) in endothelial cells. Depletion of PRMT5 by siRNA results in significantly diminished TNF-α-induced CXCL10 mRNA expression, but does not affect expression of other chemokines, such as MCP-1 or IL-8. Chromatin immunoprecipitation experiments of the CXCL10 proximal promoter show the presence of symmetrical dimethylated arginine (sDMA)-containing proteins upon exposure to TNF-α. This methylation is completely lost when PRMT5 is removed from cells by siRNA. Using immunoprecipitation, we show that PRMT5 enhances CXCL10 expression by methylating the RelA (p65) subunit of NF-κB. In summary, we have identified that PRMT5 is a novel regulator of CXCL10 expression. Further, we have discovered that PRMT5 methylates NF-κB, a finding which may further knowledge of the post-translational code governing NF-κB regulation and target specificity.

scholarly journals NF-κB regulates thrombin-induced ICAM-1 gene expression in cooperation with NFAT by binding to the intronic NF-κB site in the ICAM-1 gene

2009 ◽  
Vol 38 (1) ◽  
pp. 42-53 ◽  
Author(s):  
Jiaping Xue ◽  
Prabhakar B. Thippegowda ◽  
Guochang Hu ◽  
Kurt Bachmaier ◽  
John W. Christman ◽  
...  
Keyword(s):  
Gene Expression ◽  
Endothelial Cells ◽  
Chromatin Immunoprecipitation ◽  
Binding Sites ◽  
Leukocyte Adhesion ◽  
Reporter Expression ◽  
Mobility Shift ◽  
Intron 1 ◽  
Tnf Α ◽  
Protease Activated Receptor 1

Activation of NF-κB is essential for protease-activated receptor-1 (PAR-1)-mediated ICAM-1 expression in endothelial cells. Here we show that PAR-1 activation induces binding of both p65/RelA and NFATc1 to the NF-κB binding site localized in intron-1 of the ICAM-1 gene to initiate transcription in endothelial cells. We discovered the presence of two NF-κB binding sites in intron-1 (+70, NF-κB site 1; +611, NF-κB site 2) of the human ICAM-1 gene. Chromatin immunoprecipitation results showed that thrombin induced binding of p65/RelA and of NFATc1 specifically to intronic NF-κB site 1 of the ICAM-1 gene. Electrophoretic mobility shift and supershift assays confirmed the binding of p65/RelA and NFATc1 to the intronic NF-κB site 1 in thrombin-stimulated cells. Thrombin increased the expression of ICAM-1-promoter-intron 1-reporter (−1,385 to +234) construct ∼25-fold and mutation of intronic NF-κB site 1 markedly reduced thrombin-induced reporter expression. Moreover, inhibition of calcineurin, knockdown of either NFATc1 or p65/RelA with siRNA significantly reduced thrombin-induced ICAM-1 expression and polymorphonuclear leukocyte adhesion to endothelial cells. In contrast, NFATc1 knockdown had no effect on TNF-α-induced ICAM-1 expression. Thus these results suggest that p65/RelA and NFATc1 bind to the intronic NF-κB site 1 sequence to induce optimal transcription of the ICAM-1 gene in response to thrombin in endothelial cells.

scholarly journals The expression of Drosha, DGCR8, Dicer and Ago-2 genes are upregulated in human umbilical vein endothelial cells under hyperglycemic condition

2018 ◽  
Vol 52 (3) ◽  
pp. 123-127 ◽  
Author(s):  
Farhad Ghadiri Soufi ◽  
Ali Akbar Poursadegh Zonouzi ◽  
Ebrahim Eftekhar ◽  
Kamila Kamali ◽  
Sara Aghakhani Chegeni ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Mrna Expression ◽  
Umbilical Vein ◽  
Control Group ◽  
Human Umbilical Vein ◽  
Expression Levels ◽  
Expression Of Genes ◽  
Mirnas Expression ◽  
Umbilical Vein Endothelial Cells ◽  
Mrna Expression Levels

AbstractObjectives. It has been shown that dysregulation of miRNAs expression contributes to the pathogenesis and progression of the diabetes and diabetes-related complications. Drosha, DGCR8, Dicer, and Ago-2 are involved in the miRNA maturation. The aim of the present study was to investigate the mRNA expression levels of these genes in the human umbilical vein endothelial cells (HUVECs) under hyperglycemic condition.Methods. HUVECs were cultured in normo-(5 mM) and hyperglycemic (25 mM) conditions for 24 h. As osmotic control, cells were treated with D-mannitol (25 mM, for 24 h). The mRNA expression levels of Drosha, DGCR8, Dicer and Ago-2 were evaluated using quantitative real-time PCR.Results. The expression level of Drosha, DGCR8, Dicer, and Ago-2 were increased in hyperglycemic HUVECs compared to the control group.Conclusion. Our results show that under hyperglycemic condition, expression of genes involved in the miRNA maturation was significantly increased in HUVECs. Upregulation of these genes may have role in diabetic complications through the dysregulation of the miRNA expression.

scholarly journals Physiological concentrations of dietary polyphenols regulate vascular endothelial cell expression of genes important in cardiovascular health

2009 ◽  
Vol 103 (10) ◽  
pp. 1398-1403 ◽  
Author(s):  
Sonja K. Nicholson ◽  
Gregory A. Tucker ◽  
John M. Brameld
Keyword(s):  
Gene Expression ◽  
Endothelial Cells ◽  
Mrna Expression ◽  
Cardiovascular Health ◽  
Umbilical Vein ◽  
Vascular Endothelial Cell ◽  
Vascular Endothelial ◽  
Potential Health ◽  
Dietary Polyphenols ◽  
Expression Of Genes

Previous cell culture-based studies have shown potential health beneficial effects on gene expression of dietary polyphenols, including those found in red wine and green tea. However, these studies have tended to use higher concentrations (2–100 μm) than those observed in blood (0·1–1 μm) after consuming polyphenol-rich foods or beverages. The present study investigated effects of physiological concentrations of different classes of dietary polyphenol on the expression of genes important in cardiovascular health (endothelial NO synthase (eNOS), endothelin-1 (ET-1) and vascular endothelial growth factor (VEGF)) by cultured vascular endothelial cells (human umbilical vein endothelial cells) in the absence or presence of H2O2. Resveratrol and quercetin (0·1–1 μm) increased eNOS and VEGF mRNA expression particularly in the absence of H2O2 (50 μm) and decreased H2O2-induced ET-1 mRNA expression (P < 0·001 for polyphenol × H2O2 interactions). Similarly, resveratrol and quercetin decreased endothelin secretion into the media, blocking the stimulatory effect of 50 μm-H2O2 (P < 0·001 for polyphenol × H2O2 interaction). Of the nine other polyphenols tested, only epigallocatechin gallate had similar effects on both the eNOS and ET-1 mRNA expression, but to a lesser extent than resveratrol at an equimolar concentration (0·1 μm). The observed effects on gene expression would be expected to result in vasodilation and thereby reduced blood pressure. Since only three of the eleven polyphenols tested had biological activity, it is unclear whether particular structures are important or whether the effects might relate to the relatively high antioxidant capacities of the three active polyphenols.

scholarly journals An oxidative DNA “damage” and repair mechanism localized in the VEGF promoter is important for hypoxia-induced VEGF mRNA expression

2015 ◽  
Vol 309 (11) ◽  
pp. L1367-L1375 ◽  
Author(s):  
Viktor Pastukh ◽  
Justin T. Roberts ◽  
David W. Clark ◽  
Gina C. Bardwell ◽  
Mita Patel ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Pulmonary Artery ◽  
Mrna Expression ◽  
Transcriptional Activation ◽  
Chromatin Immunoprecipitation ◽  
Oxidative Dna Damage ◽  
Vegf Mrna ◽  
Base Modifications ◽  
Inducible Genes ◽  
Vegf Mrna Expression

In hypoxia, mitochondria-generated reactive oxygen species not only stimulate accumulation of the transcriptional regulator of hypoxic gene expression, hypoxia inducible factor-1 (Hif-1), but also cause oxidative base modifications in hypoxic response elements (HREs) of hypoxia-inducible genes. When the hypoxia-induced base modifications are suppressed, Hif-1 fails to associate with the HRE of the VEGF promoter, and VEGF mRNA accumulation is blunted. The mechanism linking base modifications to transcription is unknown. Here we determined whether recruitment of base excision DNA repair (BER) enzymes in response to hypoxia-induced promoter modifications was required for transcription complex assembly and VEGF mRNA expression. Using chromatin immunoprecipitation analyses in pulmonary artery endothelial cells, we found that hypoxia-mediated formation of the base oxidation product 8-oxoguanine (8-oxoG) in VEGF HREs was temporally associated with binding of Hif-1α and the BER enzymes 8-oxoguanine glycosylase 1 (Ogg1) and redox effector factor-1 (Ref-1)/apurinic/apyrimidinic endonuclease 1 (Ape1) and introduction of DNA strand breaks. Hif-1α colocalized with HRE sequences harboring Ref-1/Ape1, but not Ogg1. Inhibition of BER by small interfering RNA-mediated reduction in Ogg1 augmented hypoxia-induced 8-oxoG accumulation and attenuated Hif-1α and Ref-1/Ape1 binding to VEGF HRE sequences and blunted VEGF mRNA expression. Chromatin immunoprecipitation-sequence analysis of 8-oxoG distribution in hypoxic pulmonary artery endothelial cells showed that most of the oxidized base was localized to promoters with virtually no overlap between normoxic and hypoxic data sets. Transcription of genes whose promoters lost 8-oxoG during hypoxia was reduced, while those gaining 8-oxoG was elevated. Collectively, these findings suggest that the BER pathway links hypoxia-induced introduction of oxidative DNA modifications in promoters of hypoxia-inducible genes to transcriptional activation.

scholarly journals The LIM Protein AJUBA Recruits Protein Arginine Methyltransferase 5 To Mediate SNAIL-Dependent Transcriptional Repression

2008 ◽  
Vol 28 (10) ◽  
pp. 3198-3207 ◽  
Author(s):  
Zhaoyuan Hou ◽  
Hongzhuang Peng ◽  
Kasirajan Ayyanathan ◽  
Kai-Ping Yan ◽  
Ellen M. Langer ◽  
...  
Keyword(s):  
Transcriptional Repression ◽  
Epithelial Mesenchymal Transition ◽  
Arginine Methylation ◽  
Proximal Promoter ◽  
Dependent Manner ◽  
Protein Arginine Methyltransferase ◽  
Arginine Methyltransferase ◽  
Mesenchymal Transition ◽  
E Cadherin ◽  
Protein Arginine Methyltransferase 5

ABSTRACT The SNAIL transcription factor contains C-terminal tandem zinc finger motifs and an N-terminal SNAG repression domain. The members of the SNAIL family have recently emerged as major contributors to the processes of development and metastasis via the regulation of epithelial-mesenchymal transition events during embryonic development and tumor progression. However, the mechanisms by which SNAIL represses gene expression are largely undefined. Previously we demonstrated that the AJUBA family of LIM proteins function as corepressors for SNAIL and, as such, may serve as a platform for the assembly of chromatin-modifying factors. Here, we describe the identification of the protein arginine methyltransferase 5 (PRMT5) as an effector recruited to SNAIL through an interaction with AJUBA that functions to repress the SNAIL target gene, E-cadherin. PRMT5 binds to the non-LIM region of AJUBA and is translocated into the nucleus in a SNAIL- and AJUBA-dependent manner. The depletion of PRMT5 in p19 cells stimulates E-cadherin expression, and the SNAIL, AJUBA, and PRMT5 ternary complex can be found at the proximal promoter region of the E-cadherin gene, concomitant with increased arginine methylation of histones at the locus. Together, these data suggest that PRMT5 is an effector of SNAIL-dependent gene repression.

scholarly journals Heme induces mRNA expression and activation of tissue factor by TLR4 dependent mechanisms

2020 ◽  
Author(s):  
B.W. Hounkpe ◽  
C.R.P. Moraes ◽  
M.N.N. do Santos ◽  
F. F. Costa ◽  
E.V. De Paula
Keyword(s):  
Endothelial Cells ◽  
Mrna Expression ◽  
Tissue Factor ◽  
Mononuclear Cells ◽  
Cell Disease ◽  
Coagulation Activation ◽  
Coagulation Activity ◽  
Human Mononuclear Cells ◽  
Tnf Α ◽  
Dependent Pathway

AbstractIntroductionHemolytic diseases such as Sickle Cell Disease (SCD) are characterized by a natural propensity for both arterial and venous thrombosis. Evidence showing that heme can induce tissue factor (TF) expression in endothelial cells and TF-dependent coagulation activation in animal models of SCD suggest that heme can contribute to hypercoagulability in this condition. We recently demonstrated that heme can induce coagulation activation in whole blood of healthy volunteers in a TF-dependent fashion.MethodsHerein, we aimed to evaluate whether this heme-induced coagulation activity was dependent on the expression and/or activation of hematopoietic TF in human mononuclear cells. TF mRNA expression was evaluated by qPCR and TF procoagulant activity was evaluated using a 2-stage assay based on the generation of FXa.ResultsHeme was capable of inducing TF expression and activation in a TLR4-dependent pathway. This activity was further amplified after TNF-α-priming.ConclusionOur results provide additional evidences on the mechanisms by which heme is involved in the pathogenesis of hypercoagulability in hemolytic diseases.

Sign in / Sign up

Export Citation Format

Share Document