Angiogenesis, new capillary blood vessel formation, is essential for embryonic development, wound healing, and cancer growth. Vascular endothelial growth factor (VEGF) induces angiogenesis by activating endothelial cell migration and proliferation. Serum response factor (SRF) is a transcription factor important for embryonic development and activation of immediate early gene expression. The roles of SRF in endothelial cell biology and angiogenesis have not been explored. Here we demonstrate that SRF is a downstream mediator of VEGF signaling in endothelial cells and a critical requirement for VEGF‐induced angiogenesis. Knockdown of SRF protein levels in human and rat endothelial cells abolished VEGF‐induced in vitro angiogenesis, impaired endothelial cell migration and proliferation, and inhibited VEGF‐induced actin polymerization and immediate early gene expression. Injection of SRF antisense expression plasmid into gastric ulcers in rats significantly inhibited in vivo angiogenesis in granulation tissue. Mechanistically, this study also revealed that VEGF promotes SRF expression and nuclear translocation and increases SRF binding activity to DNA in endothelial cells through both Rho‐actin and MEK‐ERK dependent signaling pathways. These findings have potential therapeutic implications, e.g., local anti‐SRF treatment may inhibit angiogenesis crucial for tumor growth.