Etv2 is an immediate-response gene in VEGF-treated Flk-1⁺ mesoderm. (A) ESC-derived Flk-1⁺ cells maintained in serum-free medium were treated with VEGF (20 ng/mL) for the indicated periods of time. Gene expressions were quantified by Q-PCR. (B) VEGF(20 ng/mL) treatment for 24 hours significantly increased the Etv2⁺ cells in differentiating ESCs. In addition to an increase in Flk-1⁺ cell numbers, the number of Etv2⁺ cells among Flk-1⁺ cells was also increased. (C) Mesoderm differentiation and role of Etv2 in promoting VSP formation. Primitive mesoderm generated as the Flk-1⁺PDGFRα⁺ DP-PM population diverges into Flk-1⁺ PDGFRα⁻ VSP and Flk-1⁻ PDGFRα⁺ PSP. Etv2 is required for the progression of DP-PM into VSP. VEGF-Flk-1 signaling promotes VSP induction by up-regulating Etv2 expression. Etv2 triggers VSP induction and subsequent EC and HPC generation by inducing a series of hemato-endothelial genes, Scl, GATA2, Fli1, Lmo2, and Erg.