Aberrant expression of Jagged1 and Notch1 are associated with poor outcome in breast cancer. However, the reason that Jagged1 and/or Notch overexpression portends a poor prognosis is unknown. We identify Slug, a transcriptional repressor, as a novel Notch target and show that elevated levels of Slug correlate with increased expression of Jagged1 in various human cancers. Slug was essential for Notch-mediated repression of E-cadherin, which resulted in β-catenin activation and resistance to anoikis. Inhibition of ligand-induced Notch signaling in xenografted Slug-positive/E-cadherin–negative breast tumors promoted apoptosis and inhibited tumor growth and metastasis. This response was associated with down-regulated Slug expression, reexpression of E-cadherin, and suppression of active β-catenin. Our findings suggest that ligand-induced Notch activation, through the induction of Slug, promotes tumor growth and metastasis characterized by epithelial-to-mesenchymal transition and inhibition of anoikis.

Snail family proteins are key regulators of epithelial-mesenchymal transition, but their role in endothelial-to-mesenchymal transition (EMT) is less well studied. We show that Slug, a Snail family member, is expressed by a subset of endothelial cells as well as mesenchymal cells of the atrioventricular canal and outflow tract during cardiac cushion morphogenesis. Slug deficiency results in impaired cellularization of the cardiac cushion at embryonic day (E)–9.5 but is compensated by increased Snail expression at E10.5, which restores cardiac cushion EMT. We further demonstrate that Slug, but not Snail, is directly up-regulated by Notch in endothelial cells and that Slug expression is required for Notch-mediated repression of the vascular endothelial cadherin promoter and for promoting migration of transformed endothelial cells. In contrast, transforming growth factor β (TGF-β) induces Snail but not Slug. Interestingly, activation of Notch in the context of TGF-β stimulation results in synergistic up-regulation of Snail in endothelial cells. Collectively, our data suggest that combined expression of Slug and Snail is required for EMT in cardiac cushion morphogenesis.

Delta-like 4 (DLL4)-mediated Notch signalling has emerged as an attractive target for cancer therapy. However, the potential side effects of blocking this pathway remain uncertain. Here we show that chronic DLL4 blockade causes pathological activation of endothelial cells, disrupts normal organ homeostasis and induces vascular tumours, raising important safety concerns.

Abstract-Various studies have identified a critical role for Notch signaling in cardiovascular development. In this and other systems, Notch receptors and ligands are expressed in regions that undergo epithelial-to-mesenchymal transformation. However, there is no direct evidence that Notch activation can induce mesenchymal transdifferentiation.In this study we show that Notch activation in endothelial cells results in morphological, phenotypic, and functional changes consistent with mesenchymal transformation. These changes include downregulation of endothelial markers (vascular endothelial [VE]-cadherin, Tie1, Tie2, platelet-endothelial cell adhesion molecule-1, and endothelial NO synthase), upregulation of mesenchymal markers (␣-smooth muscle actin, fibronectin, and platelet-derived growth factor receptors), and migration toward platelet-derived growth factor-BB. Notch-induced endothelial-to-mesenchymal transformation does not seem to require external regulation and is restricted to cells expressing activated Notch. Jagged1 stimulation of endothelial cells induces a similar mesenchymal transformation, and Jagged1, Notch1, and Notch4 are expressed in the ventricular outflow tract during stages of endocardial cushion formation. This is the first evidence that Jagged1-Notch interactions induce endothelial-to-mesenchymal transformation, and our findings suggest that Notch signaling may be required for proper endocardial cushion differentiation and/or vascular smooth muscle cell development. Key Words: endothelial-to-mesenchymal transformation Ⅲ Notch Ⅲ Jagged1 Ⅲ endocardial cushion T he Notch signaling pathway plays a critical role during development. Four mammalian Notch receptors (Notch1 through 4) and 5 Notch ligands (Delta-like [Dll]-1, Dll3, Dll4, Jagged1, and Jagged2) have been identified. Notch receptorligand interaction results in a series of proteolytic cleavages of the Notch receptor, producing a C-terminal intracellular fragment (NotchIC) that translocates to the nucleus. In the nucleus, NotchIC binds to the transcriptional repressor CBF1/ RBP-J, thereby derepressing or coactivating the expression of various lineage-specific genes. 1 Perturbation of the Notch pathway has been implicated in the pathogenesis of various cardiovascular diseases in humans. 2 Of interest, patients with Jagged1 mutations (Alagille syndrome) display congenital cardiovascular anomalies that seem to be secondary to faulty endocardial cushion formation. [3][4][5][6] In the mouse, Notch1-deficient embryos demonstrate severe vascular developmental defects, which are exacerbated in Notch1/Notch4 double-mutant embryos. 7 Constitutive activation of Notch4 also causes defects in vascular remodeling. 8,9 Mice that are rendered null for Jagged1 die from hemorrhage early during embryogenesis, whereas mice that are doubly heterozygous for a Jagged1-null allele and a Notch2 hypomorphic allele exhibit cardiac anomalies similar to those seen in Alagille syndrome. 10,11 Genes that lie downstream of Notch activation, such as the basic helix-loop...

Abstract-TheNotch signaling pathway has been demonstrated to play a critical role during mammalian cardiac development based on recent findings from gene-targeted mice.