The Journal Of Biological Chemistry
Signal regulatory protein (SIRP), a highly glycosylated type-1 transmembrane protein, is composed of three immunoglobulin-like extracellular loops as well as a cytoplasmic tail containing three classical tyrosine-based inhibitory motifs. Previous reports indicate that SIRP binds to humoral pattern recognition molecules in the collectin family, namely surfactant proteins D and A (Sp-D and Sp-A, respectively), which are heavily expressed in the lung and constitute one of the first lines of innate immune defense against pathogens. However, little is known about molecular details of the structural interaction of Sp-D with SIRPs. In the present work, we examined the molecular basis of Sp-D binding to SIRP using domain-deleted mutant proteins. We report that Sp-D binds to the membrane-proximal Ig domain (D3) of SIRP in a calcium- and carbohydrate-dependent manner. Mutation of predicted N-glycosylation sites on SIRP indicates that Sp-D binding is dependent on interactions with specific N-glycosylated residues on the membrane-proximal D3 domain of SIRP. Given the remarkable sequence similarity of SIRP to SIRP and the lack of known ligands for the latter, we examined Sp-D binding to SIRP. Here, we report specific binding of Sp-D to the membrane-proximal D3 domain of SIRP. Further studies confirmed that Sp-D binds to SIRP expressed on human neutrophils and differentiated neutrophil-like cells. Because the other known ligand of SIRP, CD47, binds to the membrane-distal domain D1, these findings indicate that multiple, distinct, functional ligand binding sites are present on SIRP that may afford differential regulation of receptor function.