Role des GTPases ARF dans la migration des cellules endotheliales et la secretion du monoxyde d'azote.

摘要

ARF6 and ARF1 are small GTPases of the ARF family(s) that regulate several signalling pathways including vesicles trafficking, lipid membrane remodelling and actin cytoskeleton reorganization. To date, the role of ARF6 and ARF1 in GPCR and RTK signalling, in endothelial cells, is little known. In this thesis, we aimed to characterize the role of ARF6 in the migration of endothelial cells induced by Enodothelin-1, and the role of ARF1 in the secretion of NO induced by VEGF. We show that ARF6 is essential for endothelial cell migration induced by endothelin-1. Inhibition of ARF6 expression using RNA interference markedly impaired basal and ET-1 stimulated cell migration. In this condition, FAK is found constitutively associated with Src. In contrast, depletion of ARF6 impairs the ability of GIT1 to form an agonist-promoted complex with FAK, thereby preventing disassembly of focal adhesions. As a consequence, adhesion of ARF6-depleted endothelial cells is increased and their motility is reduced.;Furthermore, our result shows that ARF1 GTPase is essential for the activation of eNOS and the secretion of NO following VEGFR2 activation in endothelial cells.;Inhibition of ARF1 expression using RNA interference markedly impaired the recruitment of Akt to the plasma membrane and its phosphorylation by the VEGF. As a consequence, the inhibition of Akt leads to an inhibition of eNOS, a well known downstream target, which in turn leads to inhibition of NO production.;All together, our results indicate that ARF6 and ARF1 are essential for the ETB and the VEGFR2 signalling leading to cell migration and NO secretion respectively, two required steps for angiogenesis.;Keywords. ARF6, ARF1, FAK, Src, GIT, Akt, eNOS, NO, ETB, VEGFR2, Migration.