Mechanisms of VEGFR2 activation by VEGF, neuropilin, and heparin

Vascular endothelial growth factor (VEGF) and its receptor VEGFR are master regulators of vasculogenesis and angiogenesis. VEGF activates VEGFR by inducing its dimerization and trans-autophosphorylation. The coreceptor neuropilin (Nrp1 and Nrp2) and heparan sulfate proteoglycan (HSPG) modulate VEGF-VEGFR signaling, but the underlying mechanisms remain incompletely understood. Here we report a cryo-EM structure of the dimeric mouse VEGF₁₆₄-VEGFR2-Nrp1 ectodomain complex with a 2:2:2 stoichiometry, revealing direct Nrp1-VEGFR2 interactions that stabilize the VEGFR2 dimer. We also determined two cryo-EM structures of the VEGF₁₆₄-VEGFR2-Nrp1 complex in the presence of short- or long-chain heparin, which bridges all three proteins and promotes the formation of two distinct tetrameric complexes. Long-chain heparin induces a cis tetrameric complex consistent with receptor clustering on the same cell surface, whereas short-chain heparin promotes a trans tetrameric assembly which might be formed by two dimeric complexes from opposing cells. Our structure-based mutational analyses support the model that both the Nrp1-VEGFR2 interface and the heparin-mediated clustering enhance VEGFR2 signaling.