Erythroferrone antagonism of BMPs is governed by a composite heparin-binding motif

Iron homeostasis is regulated by bone morphogenetic protein (BMP) signaling, which induces hepcidin to negatively regulate serum iron levels and iron import. After a major blood loss event, developing erythroblasts produce erythroferrone (ERFE), which inhibits hepatocyte BMP signaling to increase serum iron levels and drive their maturation to erythrocytes. While ERFE was recently shown to contain a heparin-binding motif (HBM), its mechanistic significance remains poorly understood. Here, we establish that the ERFE HBM is essential for BMP inhibition and uncover a novel ternary mechanism for ligand antagonism. Using biophysical assays and molecular dynamics (MD) simulations, we show that heparin/heparan sulfate (HS) simultaneously engages with the HBM of both ERFE and BMP6 to stabilize a high-order inhibitory complex. This complex exerts far greater affinity for HS in the extracellular matrix than ERFE alone, supporting potent, ligand-dependent localization to the cell surface. Importantly, we demonstrate that ERFE preferentially engages the BMP6:HS complex over other BMP ligands, suggesting modes of ligand-HS interactions are key determinants for selectivity. Together, these findings reframe ERFE as a matrix-assisted antagonist that exploits HS as a structural co-factor for BMP antagonism and gives insight into the mechanism for tissue-restricted BMP antagonism of ERFE.