Unique functions for Notch4 in murine embryonic lymphangiogenesis

To assess DLL4/Notch LEC signaling, we seeded HdLECs on DLL4FC-coated or control FC-coated plates and collected RNA after 6 hours which was subjected to mRNA sequencing. Relative to the HdLECs seeded on FC-coated plates, DLL4FC significantly altered the expression of 675 genes with a padj <0.05. 69 genes were induced 1.2-fold while 68 genes were suppressed 1.2-fold. Analysis of the top 30 upregulated genes revealed DLL4/Notch signaling upregulated the expression of known direct effectors of Notch signaling, Hey1, EphrinB2, Hes4, Dll4, and Hes1, as well as key lymphangiogenic genes, Gja4 (Cx37), Cxcr4, Gja1 (Cx43), Ccl2, Ackr3, and Sema3g). DLL4/Notch signaling also downregulated lymphangiogenic genes, including Apln and Adm . GO: Biological Pathway (BP) analysis indicates the LEC DLL4/Notch signaling induces genes responsible for pattern specification, neurogenesis and chemotaxis. RNAseq screens from in-vitro activation of Lymphatic Endothelial Cells (LEC) by tethered DLL4 (6h) or Ig-Fc (6h).