Neuropilin-1 exerts co-receptor function for TGF-beta-1 on the membrane of cancer cells and enhances responses to both latent and active TGF-beta

Neuropilin (Nrp)-1 and Nrp-2 are multifunctional proteins frequently expressed by Cancer cells and contribute to tumor progression by mechanisms that are not well understood. They are co-receptors for vascular endothelial growth factor and class 3 semaphorins, but recently we found that Nrp1 also binds latent and active transforming growth factor (TGF)-β1, and activates the latent form latency-associated peptide (LAP)-TGF-β1. Here, we report that Nrp1 has affinity for TGF-β receptors TβRI and TβRII, the signaling TGF-β receptors, as well as TβRIII (betaglycan), as determined in binding assays, pull down assays and confocal microscopy. Nrp1 had a higher affinity for TβRI than TβRII and could form a complex with these receptors. In breast Cancer cells, Nrp1 and TβRI cointernalized in the presence of TGF-β1. Nrp1 acted as a TGF-β co-receptor by augmenting canonical SMAD2/3 signaling. Importantly, Nrp-positive Cancer cells, unlike negative cells, were able to activate latent TGF-β1 and respond. We examined two other membrane proteins that bind LAP-TGF-β, i.e. an RGD-binding Integrin (αvβ3) and Glycoprotein A repetitions predominant (CLRRC32). RGD-binding integrins are frequently expressed by Cancer cells, and glycoprotein A repetitions predominant is expressed by activated regulatory T cells that appear linked to poor tumor immunity. In vitro, these receptors did not activate LAP-TGF-β1, but subsequent addition of Nrp1 activated the cytokine. Thus, Nrp1 might collaborate with other latent TGF-β receptors in TGF-β capture and activation. We also show that Nrp2 has activities similar to Nrp1. We conclude that Nrp1 is a co-receptor for TGF-β1 and augments responses to latent and active TGF-β. Since TGF-β promotes metastasis this is highly relevant to Cancer biology.