Signal regulatory protein (SIRPalpha), a cellular ligand for CD47, regulates neutrophil transmigration.
Recent studies have demonstrated that CD47 plays an important role in regulating human neutrophil (PMN) chemotaxis. Two ligands for CD47, thrombospondin and SIRPalpha, have been described. However, it is not known if SIRP-CD47 interactions play a role in regulating PMN migration. In this study, we show that SIRPalpha1 directly binds to the immunoglobulin variable domain loop of purified human CD47 and that such SIRP-CD47 interactions regulate PMN transmigration. Specifically, PMN migration across both human epithelial monolayers and collagen-coated filters was partially inhibited by anti-SIRP monoclonal antibodies. Similar kinetics of inhibition were observed for PMN transmigration in the presence of soluble, recombinant CD47 consisting of the SIRP-binding loop. In contrast, anti-CD47 monoclonal antibodies inhibited PMN transmigration by markedly different kinetics. Results of signal transduction experiments suggested differential regulation of PMN migration by SIRP versus CD47 by phosphatidylinositol 3-kinase and tyrosine kinases, respectively. Immunoprecipitation followed by Western blotting after SDS-PAGE under nonreducing conditions suggested that several SIRP protein species may be present in PMN. Stimulation of PMN with fMLP resulted in increased surface expression of these SIRP proteins, consistent with the existence of intracellular pools. Taken together, these results demonstrate that PMN migration is regulated by CD47 through SIRPalpha-dependent and SIRPalpha-independent mechanisms.
Liu
Yuan
Y
Division of Gastrointestinal Pathology, Department of Pathology and Laboratory Medicine, Emory University, Atlanta, Georgia 30322, USA. yliu3@emory.edu
Bühring
Hans-Jörg
HJ
Zen
Ke
K
Burst
Stephanie L
SL
Schnell
Frederick J
FJ
Williams
Ifor R
IR
Parkos
Charles A
CA
D000469:Alkaline Phosphatase / Q000378:metabolism; D000730:Androstadienes / Q000494:pharmacology; D000911:Antibodies, Monoclonal / Q000378:metabolism; D015703:Antigens, CD / Q000378:metabolism*; D000943:Antigens, Differentiation; D015153:Blotting, Western; D018821:CD18 Antigens / Q000378:metabolism; D051928:CD47 Antigen; D002352:Carrier Proteins / Q000378:metabolism*; D002448:Cell Adhesion; D002465:Cell Movement; D002478:Cells, Cultured; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D005434:Flow Cytometry; D019833:Genistein / Q000494:pharmacology; D006801:Humans; D007700:Kinetics; D008024:Ligands; D008562:Membrane Glycoproteins / Q000378:metabolism*; D039842:Neural Cell Adhesion Molecule L1; D019006:Neural Cell Adhesion Molecules / Q000378:metabolism*; D009504:Neutrophils / Q000166:cytology* / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D011971:Receptors, Immunologic; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D013997:Time Factors; D000077191:Wortmannin
D000730:Androstadienes; D000911:Antibodies, Monoclonal; D015703:Antigens, CD; D000943:Antigens, Differentiation; D018821:CD18 Antigens; D051928:CD47 Antigen; C497894:CD47 protein, human; D002352:Carrier Proteins; D008024:Ligands; D008562:Membrane Glycoproteins; D039842:Neural Cell Adhesion Molecule L1; D019006:Neural Cell Adhesion Molecules; D011971:Receptors, Immunologic; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D019833:Genistein; D019869:Phosphatidylinositol 3-Kinases; D011505:Protein-Tyrosine Kinases; D000469:Alkaline Phosphatase; D000077191:Wortmannin