Transcriptional and post-transcriptional regulation of preproendothelin-1 by plaque-prone hemodynamics

OBJECTIVE: Plaque-prone areas are exposed to a particular hemodynamic environment characterized by a low mean shear stress value and a cyclic reversal flow. This mechanical environment, also termed oscillatory shear stress (OSS), induces the expression of several pro-atherogenic genes in the endothelial cells including the preproendothelin-1 (ppET-1) gene. The present paper investigates the molecular mechanisms of this induction. METHODS AND RESULTS: Several deletional mutants of ppET-1 gene promoter were cloned upstream of a luciferase gene and transiently transfected in bovine arterial endothelial cells that were further exposed to plaque-prone hemodynamics. After 24h of flow exposure, analysis of the transfected cells showed that a proximal promoter of 156 base pairs length retained OSS responsiveness. Mutation of an activator protein-1 (AP-1) binding site present in this minimal promoter completely abolished its activation by OSS. Consistently, electrophoresis mobility shift assay revealed a sustained activation of AP-1 transcription factor in endothelial cells exposed to OSS. In addition to the transcriptional activation, we demonstrated that OSS also induces a stabilization of ppET-1mRNA through the 3'-untranslated region (3'-UTR) of this gene. Fluvastatin, a drug known to improve endothelial function, was shown to prevent OSS up-regulation of the ppET-1 gene expression. Under this flow condition, fluvastatin affects ppET-1 gene expression via inhibition of its promoter activity without affecting ppET-1mRNA stability. CONCLUSIONS: The present study demonstrate that plaque-prone hemodynamic induces ppET-1 gene expression by both transcriptional and post-transcriptional mechanisms via an activation of AP-1 transcriptional factor and stabilization of mRNA. The transcriptional up-regulation of ppET-1 was shown to be fluvastatin sensitive