Activation of diacylglycerol kinase α is required for VEGF-induced angiogenic signaling in vitro

Vascular endothelial growth factor-A (VEGF-A) promotes angiogenesis by stimulating migration, proliferation and organization of endothelium, through the activation of signaling pathways involving Src tyrosine kinase. As we had previously shown that Src-mediated activation of diacylglycerol kinase- α (Dgk- α ) is required for hepatocytes growth factor-stimulated cell migration, we asked whether Dgk- α is involved in the transduction of angiogenic signaling. In PAE-KDR cells, an endothelial-derived cell line expressing VEGFR-2, VEGF-A 165 , stimulates the enzymatic activity of Dgk- α : activation is inhibited by R59949, an isoform-specific Dgk inhibitor, and is dependent on Src tyrosine kinase, with which Dgk- α forms a complex. Conversely in HUVEC, VEGF-A 165 -induced activation of Dgk is only partially sensitive to R59949, suggesting that also other isoforms may be activated, albeit still dependent on Src tyrosine kinase. Specific inhibition of Dgk- α , obtained in both cells by R59949 and in PAE-KDR by expression of Dgk- α dominant-negative mutant, impairs VEGF-A 165 -dependent chemotaxis, proliferation and in vitro angiogenesis. In addition, in HUVEC, specific downregulation of Dgk- α by siRNA impairs in vitro angiogenesis on matrigel, further suggesting the requirement for Dgk- α in angiogenic signaling in HUVEC. Thus, we propose that activation of Dgk- α generates a signal essential for both proliferative and migratory response to VEGF-A 165 , suggesting that it may constitute a novel pharmacological target for angiogenesis control.