Vascular endothelial growth factor receptor-1 and neuropilin-2 form complexes

The products of the neuropilin-1 (Np-1) and neuropilin-2 (Np-2) genes are r eceptors for factors belonging to the class 3 semaphorin family and partici pate in the guidance of growing axons to their targets. In the presence of heparin-like molecules, both receptors also function as receptors for the h eparin-binding 165-amino acid isoform of vascular endothelial growth factor (VEGF(165)). Both receptors are unable to bind to the 121-amino acid isofo rm of vascular endothelial growth factor (VEGF(121)), which lacks a heparin -binding domain. Interestingly, complexes corresponding in size to I-125-VE GF(121) neuropilin complexes are formed when I-125-VEGF(121) is bound and c ross-linked to porcine aortic endothelial cells co-expressing VEGFR-1 and e ither Np-1 or Np-2. These complexes do not seem to represent complexes of I -125-VEGF With a truncated form of VEGFR-1, presumably formed as a result o f the presence of Np-1 or Np-2 in the cells, because such truncated forms c ould not be detected with anti-VEGFR-1 antibodies. Antibodies directed agai nst VEGFR-1 co-immunoprecipitated the I-125-VEGF(121) Np-2 sized cross-link ed complex along with I-125-VEGF(121). VEGFR-1 complexes from cells express ing both VEGFR-1 and Np-2 but not from control cells, indicating that VEGFR -1 and Np-2 associate with each other. To perform the reciprocal experiment we have expressed in porcine aortic endothelial cells a Np-2 receptor cont aining an in-frame myc epitope at the C terminus. Surprisingly, the myc-tag ged Np-2 receptor lost most of its VEGF(165) binding capacity but not its s emaphorin-3F binding ability. Nevertheless, when Np-2myc was co-expressed i n cells with VEGFR-1, it partially regained its VEGF(165) binding ability. Antibodies directed against the myc epitope co-immunoprecipitated I-125-VEG F(165). Np-2myc and I-125-VEGF(165) VEGFR-1 complexes from cells co-express ing VEGFR-1 and Np-2myc, indicating again that VEGFR-1 associates with Np-2 , Our experiments therefore indicate that Np-2, and possibly also Np-1, ass ociate with VEGFR-1 and that such complexes may be part of a cell membrane- associated signaling complex.