Characterization of endothelial to mesenchymal transition induced by Notch

Abstract

Notch activation in aortic endothelial cells (ECs) takes place at embryonic stages during cardiac valve formation and induces an endothelial-to-mesenchymal transition (EndMT). Using aortic ECs, we show here that active Notch expression promotes EndMT resulting in down-regulation of VE-cadherin and up-regulation of mesenchymal genes such as Fibronectin and Snail1/2. In these cells, TGF-β1 exacerbates Notch effects increasing Snail1 and Fibronectin activation. When Notch-downstream pathways were analyzed, we detected an increase in GSK-3β phosphorylation and inactivation what facilitates Snail1 nuclear retention and protein stabilization. However, the total activity of Akt was down-regulated. The discrepancy between Akt activity and GSK-3β phosphorylation is explained because Notch induces a switch in the Akt isoforms since it decreases Akt1, the predominant isoform expressed in ECs, and up-regulates Akt2 transcription. Mechanistically, Akt2 induction requires the stimulation of TCF-4/β-catenin transcriptional complex that activates the Akt2 promoter. Active phosphorylated Akt2 translocates to the nucleus in Notch-expressing cells resulting in GSK-3β inactivation in this compartment. In the nucleus Akt2, but not Akt1 co-localizes with Lamin B in the nuclear envelope. Besides promoting GSK-3β inactivation, Notch down-regulates FoxO1, another Akt2 nuclear substrate. As a consequence, Notch protects ECs against oxidative stress-induced apoptosis through an Akt2 and Snail1-dependent mechanism.