Histidine-Rich Glycoprotein and Stanniocalcin-2 High Affinity Interactions with Inflammatory Cells

Abstract

The plasma protein Histidine-rich glycoprotein (HRG), is implicated in macrophage polarization to an M1 antitumoral phenotype. The broadly expressed, secreted protein Stanniocalcin 2 (STC2), also implicated in tumor inflammation, is an HRG interaction partner. With the aim to biochemically characterize the HRG and STC2 complex, binding of recombinant HRG and STC2 preparations to each other and to cells was explored using quartz crystal microbalance (QCM) methodology. Protein functionality was tested in a phagocytosis assay, where HRG increased phagocytosis by monocytic U937 cells while STC2 suppressed HRG-induced phagocytosis. Binding of HRG to STC2 measured using QCM showed an affinity between the proteins in the nanomolar range, which occurred in a conformation-dependent manner. Both HRG and STC2 bound individually and in combination to vitamin D3-treated, differentiated U937 monocytes. HRG, but not STC2, also bound to formaldehyde-fixed U937 cells irrespective of their differentiation stage, involving both a high affinity interaction and binding to heparan sulfate. These data show that binding of HRG to STC2 occurs with high affinity and that HRG and STC2 bind to separate sites on U937 monocytes, suggesting that they exert their effects through distinct cell surface entities.