title: Spatial raft coalescence represents an initial step in Fc gamma R signaling.
abstract: Characterization of lipid rafts as separated membrane microdomains consist of heterogeneous proteins suggesting that lateral assembly of rafts after Ag receptor cross-linking represents the earliest signal generating process. In line with the concept, cross-linked Ag receptors have been shown to associate with detergent-insoluble raft fraction without the aid of Src family kinases. However, it has not been established whether spatial raft coalescence could also precede Src family kinase activation. In this study, we showed that spatial raft coalescence after low-affinity FcgammaR cross-linking in RAW264.7 macrophages is independent of Src family kinase activity. The lateral raft assembly was found to be ascribed to the action of ligand-binding subunits, rather than to immunoreceptor tyrosine-based activation motif-bearing signal subunits, because monomeric murine FcgammaRIIb expressed in rat basophilic leukemia cells successfully induced spatial raft reorganization after cross-linking. We also showed that extracellular and transmembrane region of FcgammaRIIb is sufficient for raft stabilization. Moreover, this receptor fragment triggers rapid calcium mobilization and linker for activation of T cells phosphorylation, in a manner sensitive to Src family kinase inhibition and to cholesterol depletion. Presence of immunoreceptor tyrosine-based inhibitory motif and addition of immunoreceptor tyrosine-based activation motif to the receptor fragment abolished and enhanced the responses, respectively, but did not affect raft stabilization. These findings support the concept that ligand-binding subunit is responsible for raft coalescence, and that this event triggers initial biochemical signaling.
authors.affiliation: Department of Allergy and Rheumatology, Graduate School of Medicine and Faculty of Medicine, University of Tokyo, Tokyo, Japan.
authors.forename: Hajime
authors.initials: H
authors.lastname: Kono
authors.forename: Takeshi
authors.initials: T
authors.lastname: Suzuki
authors.forename: Kazuhiko
authors.initials: K
authors.lastname: Yamamoto
authors.forename: Masato
authors.initials: M
authors.lastname: Okada
authors.forename: Tadashi
authors.initials: T
authors.lastname: Yamamoto
authors.forename: Zen-ichiro
authors.initials: Z
authors.lastname: Honda
mesh_terms: D000818:Animals; D001854:Bone Marrow Cells / Q000201:enzymology / Q000276:immunology / Q000378:metabolism; D002460:Cell Line; D002478:Cells, Cultured; D003902:Detergents / Q000378:metabolism; D005110:Extracellular Space / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D007424:Intracellular Fluid / Q000276:immunology / Q000378:metabolism; D008264:Macrophages / Q000201:enzymology / Q000276:immunology / Q000378:metabolism; D021962:Membrane Microdomains / Q000201:enzymology / Q000276:immunology / Q000378:metabolism* / Q000502:physiology; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D017830:Octoxynol / Q000378:metabolism; D017434:Protein Structure, Tertiary / Q000235:genetics; D051381:Rats; D017452:Receptors, IgG / Q000172:deficiency / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D015398:Signal Transduction / Q000235:genetics / Q000276:immunology*; D012995:Solubility; D014407:Tumor Cells, Cultured; D019061:src-Family Kinases / Q000378:metabolism
chemical_list: D003902:Detergents; D017452:Receptors, IgG; D017830:Octoxynol; D019061:src-Family Kinases