title: The ubiquitin ligase c-Cbl down-regulates FcgammaRIIa activation in human neutrophils.
abstract: Little is known about the mechanisms that arrest FcgammaRIIa signaling in human neutrophils once engaged by immune complexes or opsonized pathogens. In our previous studies, we observed a loss of immunoreactivity of Abs directed against FcgammaRIIa following its cross-linking. In this study, we report on the mechanisms involved in this event. A stimulated internalization of FcgammaRIIa leading to the down-regulation of its surface expression was observed by flow cytometry and confocal microscopy. Immunoprecipitation of the receptor showed that FcgammaRIIa is ubiquitinated after stimulation. MG132 and clasto-lactacystin beta-lactone inhibited the loss of immunoreactivity of FcgammaRIIa, suggesting that this receptor was down-regulated via the proteasomal pathway. The E3 ubiquitin ligase c-Cbl was found to translocate from the cytosol to the plasma membrane following receptor cross-linking. Furthermore, c-Cbl was recruited to the same subset of high-density, detergent-resistant membrane fractions as stimulated FcgammaRIIa itself. Silencing the expression of c-Cbl by small interfering RNA decreased FcgammaRIIa ubiquitination and prevented its degradation without affecting the internalisation process. It also prolonged the stimulation of the tyrosine phosphorylation response to the cross-linking of the receptor. We conclude that c-Cbl mediates the ubiquitination of stimulated FcgammaRIIa and thereby contributes to the termination of FcgammaRIIa signaling via its proteasomal degradation, thus leading to the down-regulation of neutrophil signalisation and function (phagocytosis) through this receptor.
authors.affiliation: Centre de recherche en rhumatologie et immunologie, Centre de recherche du Centre hospitalier universitaire de Québec, Department of Medicine, Faculty of Medicine, Laval University, Quebec, Canada.
authors.forename: Louis
authors.initials: L
authors.lastname: Marois
authors.forename: Myriam
authors.initials: M
authors.lastname: Vaillancourt
authors.forename: Sébastien
authors.initials: S
authors.lastname: Marois
authors.forename: Sophie
authors.initials: S
authors.lastname: Proulx
authors.forename: Guillaume
authors.initials: G
authors.lastname: Paré
authors.forename: Emmanuelle
authors.initials: E
authors.lastname: Rollet-Labelle
authors.forename: Paul H
authors.initials: PH
authors.lastname: Naccache
mesh_terms: D015153:Blotting, Western; D015536:Down-Regulation; D005434:Flow Cytometry; D006801:Humans; D047468:Immunoprecipitation; D018613:Microscopy, Confocal; D009504:Neutrophils / Q000276:immunology* / Q000378:metabolism; D050721:Proto-Oncogene Proteins c-cbl / Q000276:immunology* / Q000378:metabolism; D034741:RNA, Small Interfering; D017452:Receptors, IgG / Q000276:immunology* / Q000378:metabolism; D015398:Signal Transduction / Q000276:immunology*; D014162:Transfection; D054875:Ubiquitination
chemical_list: C445323:FCGR2A protein, human; D034741:RNA, Small Interfering; D017452:Receptors, IgG; D050721:Proto-Oncogene Proteins c-cbl