20-HETE Activates the Transcription of Angiotensin-Converting Enzyme via Nuclear Factor-κB Translocation and Promoter Binding

Victor Garcia, Brian Shkolnik, Laura Milhau, J R Falck, Michal Laniado Schwartzman

Research output: Contribution to journalArticle

16 Citations (Scopus)

Abstract

Increased vascular 20-hydroxyeicosatetraenoic acid (20-HETE), a cytochrome P450 arachidonic acid metabolite, promotes vascular dysfunction, injury, and hypertension that is dependent, in part, on the renin angiotensin system (RAS). We have shown that, in human microvascular endothelial cells, 20-HETE increases angiotensin-converting enzyme (ACE) mRNA, protein, and ACE activity via an epidermal growth factor receptor (EGFR)/ tyrosine kinase/mitogen-activated protein kinase (MAPK)/ inhibitor of κB kinase (IKK)b'mediated signaling pathway. In this work, we show that, similar to epidermal growth factor (EGF), 20-HETE (10 nM) activates EGFR by stimulating tyrosine phosphorylation; however, unlike 20-HETE, EGF does not induce ACE expression, and pretreatment with a neutralizing antibody against EGF does not prevent the 20-HETE-mediated ACE induction. Inhibition of nuclear factor κB (NF-κB) activation prevented the 4.58-fold (±0.78; P <0.05) 20-HETE-mediated induction of ACE. The 20-HETE increased NF-κB'binding activity in nuclear extracts and the activity of both the somatic and germinal ACE promoters by 4.37-fold (60.18; P <0.05) and 2.53-fold (60.24; P,0.05), respectively. The 20-HETE'stimulated ACE promoter activity was abrogated by the 20-HETE antagonist 20-hydroxy-6,15-eicosadienoic acid and by inhibitors of EGFR, MAPK, IKKb, and NF-κB activation. Sequence analysis demonstrated the presence of two and one putative NF-κB binding sites on the human somatic and germinal ACE promoters, respectively. Chromatin immunoprecipitation assay indicated that 20-HETE stimulates the translocation and subsequent binding of NF-κB to each of the putative binding sites (S1, 3.43 ± 0.3-fold enrichment versus vehicle; S2, 3.72 ± 0.68-fold enrichment versus vehicle; S3, 3.20 ± 0.18-fold enrichment versus vehicle; P <0.05). This is the first study to identify NF-κB as a transcriptional factor for ACE and to implicate a distinct EGFR/MAPK/IKK/NF-κB signaling cascade underlying 20-HETE'mediated transcriptional activation of ACE mRNA and stimulation of ACE activity.

Original languageEnglish (US)
Pages (from-to)525-533
Number of pages9
JournalJournal of Pharmacology and Experimental Therapeutics
Volume356
Issue number3
DOIs
StatePublished - Mar 1 2016

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Peptidyl-Dipeptidase A
Epidermal Growth Factor Receptor
Mitogen-Activated Protein Kinases
Epidermal Growth Factor
20-hydroxy-5,8,11,14-eicosatetraenoic acid
Binding Sites
Proto-Oncogene Proteins c-akt
Messenger RNA
Enzyme Induction
Chromatin Immunoprecipitation
Vascular System Injuries
Protein Kinase Inhibitors
Renin-Angiotensin System
Neutralizing Antibodies
Arachidonic Acid
Protein-Tyrosine Kinases
Cytochrome P-450 Enzyme System
Transcriptional Activation
Sequence Analysis
Blood Vessels

ASJC Scopus subject areas

  • Pharmacology
  • Molecular Medicine

Cite this

20-HETE Activates the Transcription of Angiotensin-Converting Enzyme via Nuclear Factor-κB Translocation and Promoter Binding. / Garcia, Victor; Shkolnik, Brian; Milhau, Laura; Falck, J R; Schwartzman, Michal Laniado.

In: Journal of Pharmacology and Experimental Therapeutics, Vol. 356, No. 3, 01.03.2016, p. 525-533.

Research output: Contribution to journalArticle

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