Regulation of multiple functions of SHPS-1, a transmembrane glycoprotein, by its cytoplasmic region

Sakakibara, Ryuji; Ohnishi, Hiroshi; Kobayashi, Hill Hiroki; Kiuchi, Daisuke; Hayashi, Akiko; Kaneko, Yuka; Honma, Nakayuki; Okazawa, Hideki; Hirata, Yukio; Matozaki, Takashi

doi:10.1016/j.bbrc.2003.08.031

Cited by 19 publications

(21 citation statements)

References 35 publications

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“…We have now further shown that forced expression in RAW264.7 cells of mSHPS-1-4F, which does not bind SHP-1 or SHP-2 (25,26), markedly promoted the phagocytosis of IgG-opsonized mouse RBCs. Given that binding of CD47 to SHPS-1 negatively regulates phagocytosis by RAW264.7 cells, expression of mSHPS-1-4F appears to prevent the inhibition of Fc␥R-mediated phagocytosis by endogenous SHPS-1 in a dominant-negative manner.…”

Section: Discussionmentioning

confidence: 64%

“…To generate an expression plasmid for the mSHPS-1-4F mutant of mouse SHPS-1, in which all four tyrosine residues (Tyr 436 , Tyr 460 , Tyr 477 , Tyr 501 ) in the cytoplasmic region had been changed to phenylalanine by site-directed mutagenesis, we subcloned the corresponding full-length cDNA (provided by T. Noguchi, Kobe University, Hyogo, Japan) into pTracer-CMV (Invitrogen Life Technologies), yielding pTracer-CMVmSHPS-1-4F, as described (25). A cDNA that encodes the mSHPS-1-⌬Cyto mutant of mouse SHPS-1, which lacks almost the entire cytoplasmic region (aa 405-509), was generated by PCR, as described (25).…”

Section: Cells Cell Culture and Generation Of Raw2647 Cells Expresmentioning

confidence: 99%

“…A cDNA that encodes the mSHPS-1-⌬Cyto mutant of mouse SHPS-1, which lacks almost the entire cytoplasmic region (aa 405-509), was generated by PCR, as described (25). The PCR product was digested with EcoRI and NotI and then subcloned into the corresponding sites of pTracer-CMV.…”

Section: Cells Cell Culture and Generation Of Raw2647 Cells Expresmentioning

confidence: 99%

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Mutational Analysis of the Mechanism of Negative Regulation by Src Homology 2 Domain-Containing Protein Tyrosine Phosphatase Substrate-1 of Phagocytosis in Macrophages

Ikeda

Okazawa

Ohnishi

et al. 2006

The Journal of Immunology

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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 FcγR-mediated phagocytosis of mouse RBCs or SRBCs. Forced expression of another mutant form (SHPS-1-ΔCyto) 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-ΔCyto, in RAW264.7 cells enhanced FcγR-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.

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Section: Discussionmentioning

confidence: 64%

Section: Cells Cell Culture and Generation Of Raw2647 Cells Expresmentioning

confidence: 99%

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Mutational Analysis of the Mechanism of Negative Regulation by Src Homology 2 Domain-Containing Protein Tyrosine Phosphatase Substrate-1 of Phagocytosis in Macrophages

Ikeda

Okazawa

Ohnishi

et al. 2006

The Journal of Immunology

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“…Proteins-An SHPS-1-Fc fusion protein, which contained the extracellular region of mouse SHPS-1 (amino acids 1-371) fused to the Fc portion of human IgG, and a CD47-Fc fusion protein, which contained the extracellular region of mouse CD47 (amino acids 1-161) fused to the Fc portion of human IgG, were produced and purified as described previously (27).…”

Section: Preparation Of Shps-1-fc and Cd47-fc Fusionmentioning

confidence: 99%

“…The binding of SHPS-1-Fc to CHO-Ras cells stably expressing CD47 was determined essentially as described previously (27). Briefly, confluent CHO-Ras cells stably expressing CD47 in 96-well plates were incubated for 30 min at 37°C with various concentrations of SHPS-1-Fc in the culture medium described above, after which the cells were washed with ice-cold phosphate-buffered saline (PBS) and incubated for 30 min at 4°C with horseradish peroxidase-conjugated goat pAbs to the Fc fragment of human IgG (Jackson ImmunoResearch).…”

Section: Analysis For the Binding Of Fc Fusion Proteins To Cho-ras Cementioning

confidence: 99%

Ectodomain Shedding of SHPS-1 and Its Role in Regulation of Cell Migration

Ohnishi

Kobayashi

Okazawa

et al. 2004

Journal of Biological Chemistry

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SHPS-1 is a transmembrane protein whose cytoplasmic region undergoes tyrosine phosphorylation and then binds the protein-tyrosine phosphatase SHP-2. Formation of the SHPS-1-SHP-2 complex is implicated in regulation of cell migration. In addition, SHPS-1 and its ligand CD47 constitute an intercellular recognition system that contributes to inhibition of cell migration by cell-cell contact. The ectodomain of SHPS-1 has now been shown to be shed from cells in a reaction likely mediated by a metalloproteinase. This process was promoted by activation of protein kinase C or of Ras, and the released ectodomain exhibited minimal CD47-binding activity. Metalloproteinases catalyzed the cleavage of a recombinant SHPS-1-Fc fusion protein in vitro, and the primary cleavage site was localized to the juxtamembrane region of SHPS-1. Forced expression of an SHPS-1 mutant resistant to ectodomain shedding impaired cell migration, cell spreading, and reorganization of the actin cytoskeleton. It also increased the tyrosine phosphorylation of paxillin and FAK triggered by cell adhesion. These results suggest that shedding of the ectodomain of SHPS-1 plays an important role in regulation of cell migration and spreading by this protein.The extracellular region of various transmembrane proteins is cleaved by proteases, such as metalloproteinases, and released as a soluble protein fragment (1, 2). This process, known as "ectodomain shedding," often influences the mode of action or biological activity of the affected protein. For instance, ectodomain shedding of heparin-binding epidermal growth factor (EGF) 1 -like growth factor (HB-EGF), a membrane-anchored EGF-related protein (3), results in its activation, in that the released fragment binds to EGF receptors and thereby stimulates cell proliferation (3, 4). In contrast, the membraneanchored forms of c-kit ligand (5) and of ephrins, the latter of which are ligands of Eph receptor tyrosine kinases (6), are fully functional, whereas their soluble forms generated by ectodomain shedding exhibit little or no biological activity. Ectodomain shedding is thus an important regulator of the function of certain membrane proteins that contribute to cellcell communication.SHPS-1 (SHP substrate-1) (7), also known as SIRP␣ (8), BIT (9), and p84 neural adhesion molecule (10), is a receptor-like transmembrane protein that is particularly abundant in neurons and macrophages (10, 11), although other cell types, such as fibroblasts, also express this protein (7). The putative extracellular region of SHPS-1 comprises three immunoglobulin (Ig)-like domains with multiple N-linked glycosylation sites, whereas the cytoplasmic region of the protein contains four YXX(L/V/I) motifs, which are putative tyrosine phosphorylation sites and binding sites for the Src homology 2 (SH2) domains of the protein-tyrosine phosphatases SHP-2 and SHP-1 (7, 8). Tyrosine phosphorylation of SHPS-1 is regulated by various growth factors, including insulin and EGF, as well as by cell adhesion to extracellular matrix proteins (12). SH...

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Macrocyclic Peptide-Mediated Blockade of the CD47-SIRPα Interaction as a Potential Cancer Immunotherapy

Hazama

Yin

Murata

et al. 2020

Cell Chemical Biology

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Regulation of multiple functions of SHPS-1, a transmembrane glycoprotein, by its cytoplasmic region

Cited by 19 publications

References 35 publications

Mutational Analysis of the Mechanism of Negative Regulation by Src Homology 2 Domain-Containing Protein Tyrosine Phosphatase Substrate-1 of Phagocytosis in Macrophages

Mutational Analysis of the Mechanism of Negative Regulation by Src Homology 2 Domain-Containing Protein Tyrosine Phosphatase Substrate-1 of Phagocytosis in Macrophages

Ectodomain Shedding of SHPS-1 and Its Role in Regulation of Cell Migration

Macrocyclic Peptide-Mediated Blockade of the CD47-SIRPα Interaction as a Potential Cancer Immunotherapy

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