Interaction between the integrin Mac-1 and signal regulatory protein (SIRP) mediates fusion in heterologous cells

Macrophage fusion leading to the formation of multinucleated giant cells is a hallmark of chronic inflammation. Several membrane proteins have been implicated in mediating cell-cell attachment during fusion, but their binding partners remain unknown. Recently, we demonstrated that interleukin-4 (IL-4)-induced fusion of mouse macrophages depends on the integrin macrophage antigen 1 (Mac-1). Surprisingly, the genetic deficiency of intercellular adhesion molecule 1 (ICAM-1), an established ligand of Mac-1, did not impair macrophage fusion, suggesting the involvement of other counter-receptors. Here, using various approaches, including signal regulatory protein (SIRP) knockdown, recombinant proteins, adhesion and fusion assays, biolayer interferometry, and peptide libraries, we show that SIRP, which, similar to ICAM-1, belongs to the Ig superfamily and has previously been implicated in cell fusion, interacts with Mac-1. The following results support the conclusion that SIRP is a ligand of Mac-1: (a) recombinant ectodomain of SIRP supports adhesion of Mac-1-expressing cells; (b) Mac-1-SIRP interaction is mediated through the ligand-binding I-M-domain of Mac-1; (c) recognition of SIRP by the I-M-domain conforms to general principles governing binding of Mac-1 to many of its ligands; (d) SIRP reportedly binds CD47; however, anti-CD47 function-blocking mAb produced only a limited inhibition of macrophage adhesion to SIRP; and (e) co-culturing of SIRP- and Mac-1-expressing HEK293 cells resulted in the formation of multinucleated cells. Taken together, these results identify SIRP as a counter-receptor for Mac-1 and suggest that the Mac-1-SIRP interaction may be involved in macrophage fusion.