title: Nitric oxide decreases the expression of endothelin-converting enzyme-1 through mRNA destabilization.
abstract: OBJECTIVE
  Endothelial function depends on the equilibrium in the synthesis of vasoactive endothelial factors. It is well known that endothelin and nitric oxide (NO) exhibit reciprocal regulation. We assessed the ability of NO to regulate endothelin-converting enzyme-1 (ECE-1) expression in vascular endothelial cells.
  
  
  RESULTS
  Bovine aortic endothelial cells were incubated with 2 different NO donors as well as with a cyclic-GMP analog, dibutyryl-cGMP (dB-cGMP). ECE-1 protein content and mRNA expression were evaluated by Western blot and Northern blot, respectively, promoter activity by transfection experiments, ECE-1 activity by ELISA, and cGMP production by radioimmunoassay. Both NO donors decreased ECE-1 protein content, mRNA expression, and ECE-1 activity. ODQ, an inhibitor of soluble guanylyl cyclase, blocked those effects. NO donors raised cGMP levels, and dB-cGMP mimicked their effects on ECE-1 expression, which were blocked by KT5823, a nonspecific PKG inhibitor. The changes on ECE-1 expression were due to a destabilization on 3'-untranslated region (3'-UTR) of this mRNA, because the activity of a luciferase reporter construct containing the 3'-UTR of the ECE-1 gene was reduced by dB-cGMP in a PKG-dependent manner. The biological relevance of this regulation was confirmed in bovine aortic endothelial cells coincubated with macrophages in the presence of lipopolysaccharide, in eNOS-deficient mice, and in Wistar rats treated with NO donors. In every case, an inverse relationship was observed between NO and ECE-1 protein content.
  
  
  CONCLUSIONS
  Our results support that NO regulates ECE-1 expression through a cGMP/PKG-dependent regulatory mechanism at the post-transcriptional level via the 3'-UTR of the ECE-1 gene.
authors.affiliation: Research Unit and Nephrology Section, Hospital Universitario Príncipe de Asturias, Alcalá de Henares, Spain.
authors.forename: Viviana
authors.initials: V
authors.lastname: Raoch
authors.forename: Fernando
authors.initials: F
authors.lastname: Rodríguez-Pascual
authors.forename: Vanesa
authors.initials: V
authors.lastname: López-Martínez
authors.forename: Diana
authors.initials: D
authors.lastname: Medrano-Andrés
authors.forename: Manuel
authors.initials: M
authors.lastname: Rodríguez-Puyol
authors.forename: Santiago
authors.initials: S
authors.lastname: Lamas
authors.forename: Diego
authors.initials: D
authors.lastname: Rodríguez-Puyol
authors.forename: Susana
authors.initials: S
authors.lastname: López-Ongil
mesh_terms: D000818:Animals; D001011:Aorta / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D016282:Aspartic Acid Endopeptidases / Q000235:genetics / Q000378:metabolism*; D002417:Cattle; D002460:Cell Line; D018920:Coculture Techniques; D006152:Cyclic GMP / Q000378:metabolism / Q000494:pharmacology; D017869:Cyclic GMP-Dependent Protein Kinases / Q000378:metabolism; D015536:Down-Regulation / Q000187:drug effects* / Q000502:physiology; D000072396:Endothelin-Converting Enzymes; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D006162:Guanylate Cyclase / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D007668:Kidney / Q000166:cytology / Q000187:drug effects / Q000196:embryology / Q000378:metabolism; D008264:Macrophages / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D008666:Metalloendopeptidases / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D018345:Mice, Knockout; D023421:Models, Animal; D009569:Nitric Oxide / Q000378:metabolism*; D020030:Nitric Oxide Donors / Q000494:pharmacology*; D052250:Nitric Oxide Synthase Type III / Q000172:deficiency / Q000235:genetics; D012333:RNA, Messenger / Q000187:drug effects / Q000378:metabolism*; D017208:Rats, Wistar
chemical_list: D020030:Nitric Oxide Donors; D012333:RNA, Messenger; D009569:Nitric Oxide; D052250:Nitric Oxide Synthase Type III; C496315:Nos3 protein, mouse; D017869:Cyclic GMP-Dependent Protein Kinases; D016282:Aspartic Acid Endopeptidases; D008666:Metalloendopeptidases; C000606923:ECE1 protein, human; C000606925:Ece1 protein, mouse; D000072396:Endothelin-Converting Enzymes; D006162:Guanylate Cyclase; D006152:Cyclic GMP