Hypoxic induction of human vascular endothelial growth factor expression through c-Src activation

ANGIOGENESIS, the formation of new microvasculature by capillary sprouting, is crucial for tumour development¹. Hypoxic regions of solid tumours produce the powerful and directly acting angiogenic protein VEGF/VPF (vascular endothelial growth factor/vascular permeability factor)^2-6. We now investigate the signal transduction pathway involved in hypoxic induction of VEGF expression. Hypoxia is known to induce a tyrosine kinase cascade that results in the activation of nitrogen-fixation genes in Rhizobium meliloti ⁷, and activation of tyrosine kinases is critical in signalling triggered by growth factors and ultraviolet light. We show here that genistein, an inhibitor of protein tyrosine kinase⁸, blocks VEGF induction. Hypoxia increases the kinase activity of pp60^c-src and its phosphorylation on tyrosine 416 but does not activate Fyn or Yes. Expression of either a dominant-negative mutant form of c-Src or of Raf-1 markedly reduces VEGF induction. VEGF induction by hypoxia in c-src(-) cells is impaired, although there is a compensatory activation of Fyn. Our results provide an insight into hypoxia-triggered intracellular signalling, define VEGF as a new downstream target for c-Src, and suggest a role for c-Src in promoting angiogenesis.