title: Functional elements on SIRPalpha IgV domain mediate cell surface binding to CD47.
abstract: SIRPalpha and SIRPbeta1, 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 SIRPalpha with CD47, another important cell surface molecule, through the extracellular IgV domain regulates important leukocyte functions including macrophage recognition, leukocyte adhesion and transmigration. Although SIRPbeta1 shares highly homologous extracellular IgV structure with SIRPalpha, it does not bind to CD47. Here, we defined key amino acid residues exclusively expressing in the IgV domain of SIRPalpha, but not SIRPbeta1, which determine the extracellular binding interaction of SIRPalpha 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/Val57 are critical. Mutation of either of these residues abates SIRPalpha directly binding to CD47. Functional cell adhesion and leukocyte transmigration assays further demonstrated central roles of Gln67 and Ala/Val57 in SIRPalpha extracellular binding mediated cell interactions and cell migration. Another SIRPalpha-specific residue, Met102, appears to assist SIRPalpha IgV binding through Gln67 and Ala/Val57. An essential role of these amino acid residues in SIRPalpha binding to CD47 was further confirmed by introducing these residues into the SIRPbeta1 IgV domain, which dramatically converts SIRPbeta1 into a CD47-binding molecule. Our results thus revealed the molecular basis by which SIRPalpha binds to CD47 and shed new light into the structural mechanisms of SIRP isoform mediated distinctive extracellular interactions and cellular responses.
authors.affiliation: Department of Biology, Georgia State University, Atlanta, GA 30302, USA. yliu@gsu.edu
authors.forename: Yuan
authors.initials: Y
authors.lastname: Liu
authors.forename: Qiao
authors.initials: Q
authors.lastname: Tong
authors.forename: Yubin
authors.initials: Y
authors.lastname: Zhou
authors.forename: Hsiau-Wei
authors.initials: HW
authors.lastname: Lee
authors.forename: Jenny J
authors.initials: JJ
authors.lastname: Yang
authors.forename: Hans-Jörg
authors.initials: HJ
authors.lastname: Bühring
authors.forename: Yi-Tien
authors.initials: YT
authors.lastname: Chen
authors.forename: Binh
authors.initials: B
authors.lastname: Ha
authors.forename: Celia X-J
authors.initials: CX
authors.lastname: Chen
authors.forename: Yang
authors.initials: Y
authors.lastname: Yang
authors.forename: Ke
authors.initials: K
authors.lastname: Zen
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000943:Antigens, Differentiation / Q000737:chemistry* / Q000378:metabolism*; D001665:Binding Sites; D051928:CD47 Antigen / Q000737:chemistry / Q000378:metabolism*; D002448:Cell Adhesion; D002465:Cell Movement; D018922:HL-60 Cells; D019073:HT29 Cells; D006801:Humans; D007962:Leukocytes / Q000166:cytology; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation / Q000235:genetics; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011817:Rabbits; D011956:Receptors, Cell Surface / Q000737:chemistry / Q000378:metabolism; D011961:Receptors, Fc / Q000276:immunology; D011971:Receptors, Immunologic / Q000737:chemistry* / Q000378:metabolism*; D040681:Structural Homology, Protein; D013329:Structure-Activity Relationship
chemical_list: D000943:Antigens, Differentiation; D051928:CD47 Antigen; D011956:Receptors, Cell Surface; D011961:Receptors, Fc; D011971:Receptors, Immunologic; C501920:SIRPA protein, human; C501921:SIRPB1 protein, human