Mutational analysis of the mechanism of negative regulation by SRC homology 2 domain-containing protein tyrosine phosphatase substrate-1 of phagocytosis in macrophages.
Src homology 2 domain-containing protein tyrosine phosphatase substrate-1 (SHPS-1) is a transmembrane protein predominantly expressed in macrophages. The binding of CD47 on RBCs to SHPS-1 on macrophages is implicated in inhibition of phagocytosis of the former cells by the latter. We have now shown that forced expression in mouse RAW264.7 macrophages of a mutant version (SHPS-1-4F) of mouse SHPS-1, in which four tyrosine phosphorylation sites are replaced by phenylalanine, markedly promoted Fc gammaR-mediated phagocytosis of mouse RBCs or SRBCs. Forced expression of another mutant form (SHPS-1-deltaCyto) of mouse SHPS-1, which lacks most of the cytoplasmic region, did not promote such phagocytosis. Similarly, forced expression of a rat version of SHPS-1-4F, but not that of rat wild-type SHPS-1 or SHPS-1-deltaCyto, in RAW264.7 cells enhanced Fc gammaR-mediated phagocytosis of RBCs. Tyrosine phosphorylation of endogenous SHPS-1 as well as its association with Src homology 2 domain-containing protein tyrosine phosphatase-1 were not markedly inhibited by expression of SHPS-1-4F. Furthermore, the attachment of IgG-opsonized RBCs to RAW264.7 cells was markedly increased by expression of SHPS-1-4F, and this effect did not appear to be mediated by the interaction between CD47 and SHPS-1. These data suggest that inhibition by SHPS-1 of phagocytosis in macrophages is mediated, at least in part, in a manner independent of the transinteraction between CD47 and SHPS-1. In addition, the cytoplasmic region as well as tyrosine phosphorylation sites in this region of SHPS-1 appear indispensable for this inhibitory action of SHPS-1. Moreover, SHPS-1 may regulate the attachment of RBCs to macrophages by an as yet unidentified mechanism.
Ikeda
Hiroshi
H
Laboratory of Biosignal Sciences, Institute for Molecular and Cellular Regulation, Gunma University, Gunma, Japan.
Okazawa
Hideki
H
Ohnishi
Hiroshi
H
Murata
Yoji
Y
Oldenborg
Per-Arne
PA
Matozaki
Takashi
T
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D051928:CD47 Antigen / Q000235:genetics / Q000378:metabolism; D016827:CD8 Antigens / Q000235:genetics / Q000378:metabolism; D004912:Erythrocytes / Q000378:metabolism; D005786:Gene Expression Regulation; D047908:Intracellular Signaling Peptides and Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D008264:Macrophages / Q000276:immunology / Q000378:metabolism*; D051379:Mice; D018345:Mice, Knockout; D009154:Mutation / Q000235:genetics; D010587:Phagocytosis; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D000081082:Phosphoinositide-3 Kinase Inhibitors; D019000:Phosphotyrosine / Q000378:metabolism; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D051381:Rats; D017452:Receptors, IgG / Q000276:immunology / Q000378:metabolism; D011971:Receptors, Immunologic / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D000072377:Syk Kinase; D018909:src Homology Domains
D000911:Antibodies, Monoclonal; D051928:CD47 Antigen; D016827:CD8 Antigens; D047908:Intracellular Signaling Peptides and Proteins; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D019000:Phosphotyrosine; D047428:Protein Kinase Inhibitors; D011505:Protein-Tyrosine Kinases; D017452:Receptors, IgG; D011971:Receptors, Immunologic; D000072377:Syk Kinase; C501922:Sirpa protein, mouse; C000606897:Syk protein, mouse; C000606898:Syk protein, rat