The Prodomain-bound Form of Bone Morphogenetic Protein 10 Is Biologically Active on Endothelial Cells

Abstract

BMP10 is highly expressed in the developing heart and plays essential roles in cardiogenesis. BMP10 deletion in mice results in embryonic lethality because of impaired cardiac development. In adults, BMP10 expression is restricted to the right atrium, though ventricular hypertrophy is accompanied by increased BMP10 expression in a rat hypertension model. However, reports of BMP10 activity in the circulation are inconclusive. In particular, it is not known whether in vivo secreted BMP10 is active or whether additional factors are required to achieve its bioactivity. It has been shown that high-affinity binding of the BMP10 prodomain to the mature ligand inhibits BMP10 signaling activity in C2C12 cells, and it was proposed that prodomain-bound BMP10 (pBMP10) complex is latent. In this study, we demonstrated that the BMP10 prodomain did not inhibit BMP10 signaling activity in multiple endothelial cells, and that recombinant human pBMP10 complex, expressed in mammalian cells and purified under native conditions, was fully active. In addition, both BMP10 in human plasma and BMP10 secreted from the mouse right atrium were fully active. Finally, we confirmed that active BMP10 secreted from mouse right atrium was in the prodomain-bound form. Our data suggest that circulating BMP10 in adults is fully active and that the reported vascular quiescence function of BMP10 in vivo is due to the direct activity of pBMP10 and does not require an additional activation step. Moreover, being an active ligand, recombinant pBMP10 may have therapeutic potential as an endothelial-selective BMP ligand, in conditions characterized by loss of BMP9/10 signaling.

Rapid Activation of Bone Morphogenic Protein 9 by Receptor-mediated Displacement of Pro-domains

“…Specific activation of latent BMP10⅐pro-domain complexes was achieved by cleavage of the pro-domain by the metalloprotease BMP1 (16). A recent study, however, reveals that the BMP10⅐pro-domain is active in multiple endothelial cells and that the ECD of BMPRII can release BMP10 from the pro-domain complex (33). Questions remain of how these very similar growth factors function differently in a tissue-specific manner, as BMP10 exhibits a unique role during cardiac development that cannot be compensated by BMP9 (6).…”

Regulation of the ALK1 ligands, BMP9 and BMP10

“…Using mammalian cell-expressed full-length human recombinant BMP10 and ex vivo-cultured mouse right atria, we have recently demonstrated that prodomain bound BMP10 is not latent but fully active on multiple ECs. Circulating BMP10 activity can be detected, albeit at much lower levels than BMP9 [38].…”

“…The prodomain remains bound to the mature ligands in circulation [14] (Figure 2A). When BMPs expressed in mammalian cells were subjected to nonreducing SDS/PAGE, a monomeric form (M-form) is present alongside the canonical disulfide-linked dimeric form (Dform) [36][37][38]. In the case of BMP9, two high-resolution crystal structures, both crystallized from mammalian cell expressed proteins and purified under native conditions, have shown that the M-form of BMP9 is in fact a non-covalently linked dimer, almost identical to the D-form ( Figures 2B and 2C) [37,39].…”

ALK1 signaling in development and disease: new paradigms

“…The BMP10 procomplex was initially classified as latent based on activity tests using cultured C2C12 (myoblast) or NIH 3T3 (fibroblast) cells [124, 129]. However, the activity of the BMP10 procomplex on a variety of ECs was recently shown to approach that of the BMP10 growth factor homodimer [130]. These data suggest that surface proteins on ECs in addition to ALK1 might be important for either stabilizing the binding of the BMP10 prodomain or displacing it from the growth factor homodimer.…”