Functional association of CD9 with the Fc gamma receptors in macrophages.
CD9, a member of the tetraspan family of proteins, is highly expressed on macrophages. Although a clear function for the molecule has yet to be described, we have found that the anti-CD9 mAb activates mouse macrophages. The rat anti-CD9 mAb, KMC8.8, but not the F(ab')(2), induced tyrosine phosphorylation of proteins including syk and cbl and induced cell aggregation in the mouse macrophage cell line, J774, suggesting that co-cross-linking of CD9 and Fc gamma R was required for the signal. Co-cross-linking of CD9-Fc gamma R with KMC8.8 on macrophages from three different FcR-deficient mice, FcR gamma-chain(-/-), Fc gamma RIIB(-/-), and Fc gamma RIII(-/-), revealed that Fc gamma RIII is specific and crucial for syk phosphorylation. Although both KMC8.8 and the anti-Fc gamma RIIB/III mAb, 2.4G2, evoked similar phosphorylation patterns, only KMC8.8 induced cell aggregation. Additionally, KMC8.8 treatment led to reduce levels of TNF-alpha production and p42/44 extracellular signal-related kinase phosphorylation relative to 2.4G2 stimulation. Immunofluorescence staining showed that co-cross-linking of CD9-Fc gamma R with KMC8.8 induced filopodium extension before cell aggregation, which was followed by simultaneous colocalization of CD9, Fc gamma RIIB/III, Mac-1, ICAM-1, and F-actin at the cell-cell adhesion site. Moreover, KMC8.8 treatment of Fc gamma R-deficient macrophages revealed that the colocalization of CD9, Fc gamma RIII, Mac-1, and F-actin requires co-cross-linking of CD9-Fc gamma RIII, whereas co-cross-linking of CD9-Fc gamma RIIB induced the colocalization of only CD9 and Fc gamma RIIB. Our results demonstrate that co-cross-linking of CD9 and Fc gamma Rs activates macrophages; therefore, CD9 may collaborate with FcRs functioning in infection and inflammation on macrophages.
Kaji
K
K
Department of Life Science, Tokyo Institute of Technology, Yokohama, Japan.
Takeshita
S
S
Miyake
K
K
Takai
T
T
Kudo
A
A
D000199:Actins / Q000378:metabolism; D000818:Animals; D000911:Antibodies, Monoclonal / Q000378:metabolism / Q000494:pharmacology; D015703:Antigens, CD / Q000276:immunology / Q000378:metabolism / Q000502:physiology*; D002448:Cell Adhesion / Q000276:immunology; D002449:Cell Aggregation / Q000276:immunology; D002460:Cell Line; D048429:Cell Size / Q000276:immunology; D002784:Cholesterol / Q000378:metabolism; D003902:Detergents; D006017:Glycolipids / Q000378:metabolism; D007106:Immune Sera / Q000378:metabolism; D018799:Intercellular Adhesion Molecule-1 / Q000378:metabolism; D016177:Macrophage-1 Antigen / Q000378:metabolism; D008264:Macrophages / Q000166:cytology / Q000276:immunology* / Q000378:metabolism*; D008562:Membrane Glycoproteins; D008563:Membrane Lipids / Q000378:metabolism; D051379:Mice; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000378:metabolism; D011554:Pseudopodia / Q000276:immunology; D017452:Receptors, IgG / Q000276:immunology / Q000378:metabolism* / Q000502:physiology; D015398:Signal Transduction / Q000276:immunology; D012995:Solubility; D060245:Tetraspanin 29; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis
D000199:Actins; D000911:Antibodies, Monoclonal; D015703:Antigens, CD; C554870:Cd9 protein, mouse; D003902:Detergents; D006017:Glycolipids; D007106:Immune Sera; D016177:Macrophage-1 Antigen; D008562:Membrane Glycoproteins; D008563:Membrane Lipids; D017452:Receptors, IgG; D060245:Tetraspanin 29; D014409:Tumor Necrosis Factor-alpha; D018799:Intercellular Adhesion Molecule-1; D019000:Phosphotyrosine; D002784:Cholesterol