The c‐Cbl/CD2AP complex regulates VEGF‐induced endocytosis and degradation of Flt‐1 (VEGFR‐1)

Abstract

Vascular endothelial growth factor (VEGF) and its receptors are key regulators of angiogenesis and are potential targets in cancer therapy. Here we report the down-regulation of activated VEGF receptor (VEGFR)-1/Flt-1 by endocytosis and proteolytic degradation. VEGF stimulation induced a ternary complex of Flt-1, c-Cbl, and CD2AP. Substitution of tyrosine 1333 in Flt-1 with phenylalanine (Y1333F) impaired its binding to c-Cbl. In a transient expression system, VEGF stimulated colocalization of Flt-1, CD2AP, and c-Cbl in endocytic vesicles. This colocalization was significantly impaired by an inhibitor of VEGFR kinase SU5416, the Y1333F mutation in Flt-1, or by a dominant negative form of CD2AP. In Flt-1-overexpressing NIH3T3 cells, expression of the wild-type CD2AP enhanced VEGF-stimulated internalization as well as ubiquitination of Flt-1 whereas that of a mutated form of either CD2AP or c-Cbl failed to do so. These results suggest that the c-Cbl/CD2AP complex binds to activated Flt-1 and plays a crucial role in its endocytosis and subsequent degradation.