Functional elements on SIRPα IgV domain mediate cell surface binding to CD47

SIRP alpha and SIRP beta 1, the two major isoforms of the signal regulatory protein (SIRP) family, are co-expressed in human leukocytes but mediate distinct extracellular binding interactions and divergent cell signaling responses. Previous studies have demonstrated that binding of SIRP alpha with CD47, another important cell surface molecule, through the extracellular IgV domain regulates important leukocyte functions including macrophage,, recognition, leukocyte adhesion and transmigration. Although SIRP beta 1 shares highly homologous extracellular IgV structure with SIRPQ, it does not bind to CD47. Here, we defined key amino acid residues exclusively expressing in the IgV domain of SIRP alpha, but not SIRP beta 1, which determine the extracellular binding interaction of SIRP alpha to CD47. These key residues include Gln67, a small hydrophobic amino acid (Ala or Val) at the 57th position and Met102. We found that Gln67 and Ala/Va157 are critical. Mutation of either of these residues abates SIRP alpha directly binding to CD47. Functional cell adhesion and leukocyte transmigration assays further demonstrated central roles of Gln67 and Ala/Val57 in SIRP alpha extracellular binding mediated cell interactions and cell migration. Another SlRPuspecific residue, Met102, appears to assist SIRP alpha IgV binding through Gln67 and Ala/Va157. An essential role of these amino acid residues in SIRP alpha binding to CD47 was further confirmed by introducing these residues into the SIRP l IgV domain, which dramatically converts SIRP beta 1 into a CD47-binding molecule. Our results thus revealed the molecular basis by which SIRP alpha binds to CD47 and shed new light into the structural mechanisms of SIRP isoform mediated distinctive extracellular interactions and cellular responses. (c) 2006 Elsevier Ltd. All rights reserved.