Caveolin 1 is required for the activation of endothelial nitric oxide synthase in response to 17beta-estradiol.
Evidence suggests that estrogen mediates rapid endothelial nitric oxide synthase (eNOS) activation via estrogen receptor-a (ERalpha) within the plasma membrane of endothelial cells (EC). ERalpha is known to colocalize with caveolin 1, the major structural protein of caveolae, and caveolin 1 stimulates the translocation of ERalpha to the plasma membrane. However, the role played by caveolin 1 in regulating 17beta-estradiol-mediated NO signaling in EC has not been adequately resolved. Thus, the purpose of this study was to explore how 17beta-estradiol stimulates eNOS activity and the role of caveolin 1 in this process. Our data demonstrate that modulation of caveolin 1 expression using small interfering RNA or adenoviral gene delivery alters ERalpha localization to the plasma membrane in EC. Further, before estrogen stimulation ERalpha associates with caveolin 1, whereas stimulation promotes a pp60(Src)-mediated phosphorylation of caveolin 1 at tyrosine 14, increasing ERalpha-PI3 kinase interactions and disrupting caveolin 1-ERalpha interactions. Adenoviral mediated overexpression of a phosphorylation-deficient mutant of caveolin (Y14FCav) attenuated the ERalpha/PI3 kinase interaction and prevented Akt-mediated eNOS activation. Furthermore, Y14FCav overexpression reduced eNOS phosphorylation at serine1177 and decreased NO generation after estrogen exposure. Using a library of overlapping peptides we identified residues 62-73 of caveolin 1 as the ERalpha-binding site. Delivery of a synthetic peptide based on this sequence decreased ERalpha plasma membrane translocation and reduced estrogen-mediated activation of eNOS. In conclusion, caveolin 1 stimulates 17beta-estradiol-induced NO production by promoting ERalpha to the plasma membrane, which facilitates the activation of the PI3 kinase pathway, leading to eNOS activation and NO generation.
Sud
Neetu
N
Pulmonary Vascular Disease Program, Vascular Biology Center, Medical College of Georgia, Augusta, Georgia 30912, USA.
Wiseman
Dean A
DA
Black
Stephen M
SM
D000818:Animals; D051242:Caveolin 1 / Q000235:genetics / Q000378:metabolism*; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000187:drug effects / Q000378:metabolism; D004958:Estradiol / Q000494:pharmacology*; D047628:Estrogen Receptor alpha / Q000378:metabolism; D009569:Nitric Oxide / Q000378:metabolism; D052250:Nitric Oxide Synthase Type III / Q000235:genetics / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D011651:Pulmonary Artery / Q000187:drug effects / Q000378:metabolism; D012756:Sheep; D015398:Signal Transduction / Q000187:drug effects / Q000235:genetics
D051242:Caveolin 1; D047628:Estrogen Receptor alpha; D009569:Nitric Oxide; D004958:Estradiol; D052250:Nitric Oxide Synthase Type III; D019869:Phosphatidylinositol 3-Kinases; D051057:Proto-Oncogene Proteins c-akt