The interaction between Lyn and FcεRIβ is indispensable for FcεRI-mediated human mast cell activation.
ABSTRACT FcεRIβ reportedly functions as an amplifier of the FcεRIγ-mediated activation signal using a reconstitution system. However, the amplification mechanisms in human mast cells (MCs) are poorly understood. We previously reported the hyperexpression of FcεRIβ of MCs in giant papillae from vernal keratoconjunctivitis patients, compared with that in conjunctivae from nonallergic conjunctivitis patients. Elucidation of the molecular mechanisms of the amplification induced by FcεRIβ should provide new targets for novel therapeutic interventions. The aim is to understand in greater details the function of FcεRIβ in human MC FcεRI expression and signaling.
FcεRIβ and Lyn expression was reduced using a lentiviral shRNA silencing technique. Localization of Lyn and FcεRIβ in cultured MCs was examined by confocal microscopic analysis. Mediators were measured by ELISAs.
The diminution of FcεRIβ significantly downregulated cell surface FcεRI expression and FcεRI-mediated mediator release/production. The downregulation of FcεRI-mediated degranulation was not only due to the decrease in FcεRI expression. The diminution of FcεRIβ inhibited the redistribution of Lyn within the cell membrane following IgE sensitization. The diminution of Lyn in MCs significantly downregulated FcεRI-mediated degranulation. The recombinant cell-penetrating forms of phosphorylated FcεRIβ immunoreceptor tyrosine-based activation motif (ITAM) for intracellular delivery disturbed the interaction between Lyn and phosphorylated endogenous FcεRIβ ITAM, resulted in inhibiting IgE-dependent histamine release from MCs in vitro and from giant papillae specimens ex vivo.
The interaction between Lyn and FcεRIβ is indispensable for FcεRI-mediated human MC activation, and specific inhibition of the interaction may represent a new therapeutic strategy for the treatment of human allergic diseases.
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ABSTRACT: We recently reported that the interaction between Lyn and FcεRIβ is indispensable for FcεRI-mediated human mast cells (MC) activation and that FcεRIβ functions as an amplifier of FcεRI-mediated activation signal. Some of FcεRIβ in cytoplasm appeared not to be co-localized with FcεRIα. The function of FcεRIβ in the cytoplasm remains unknown. The localization of FcεRIβ and FcεRIα in giant papillae specimens from patients with allergic keratoconjunctivitis and of FcεRIβ, FcεRIα, and Lyn in cultured human MCs was examined using confocal microscopy. FcεRIβ was overexpressed using an adenovirus vector system. Mediators were measured by enzyme immunoassays or enzyme-linked immunosorbent assays. In the subepithelial region, FcεRIβ was mainly localized in the cell membrane of MCs. In the perivascular region, FcεRIβ expression was scattered throughout the cytoplasm and in the cell membrane of MCs. Overexpression of FcεRIβ in MCs mainly increased its cytoplasmic expression and slightly upregulated cell surface FcεRI expression. However, overexpression of FcεRIβ in MCs resulted in downregulation of the tyrosine phosphorylation levels of FcεRIβ and Syk and downregulation of the Ca(2+) influx soon after FcεRI aggregation, and then resulted in downregulation of degranulation, PGD2 synthesis, and production of a set of cytokines. This negative regulatory effect may be due to inhibition of the redistribution of Lyn to small patches within the plasma membrane. Cytoplasmic FcεRIβ, which is not co-localized with FcεRIα, may function as a negative regulator, since it can capture important signaling molecules such as Lyn. This article is protected by copyright. All rights reserved.Clinical & Experimental Allergy 10/2013; 44(2). DOI:10.1111/cea.12210 · 4.32 Impact Factor
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ABSTRACT: The risk to develop anaphylaxis depends on the sensitization pattern, the proportion of the involved immunoglobulin classes, the avidity and affinity of immunoglobulins to bind an allergen, characteristics of the allergen, the route of allergen application, and, last but not least, the presence of cofactors of anaphylaxis. To be able to calculate the risk to develop anaphylaxis and to anticipate the severity of the reactions under certain conditions, it is necessary to understand how all these factors interact with each other. Important progress for risk assessment in anaphylaxis is based on component-resolved stratified diagnostics, which allow to (i) determine a patient's sensitization pattern on a molecular basis, (ii) correlate clinical responses to defined sensitization patterns, and (iii) better identify cross-reactive allergens. Together with the increasing knowledge regarding the role and mode of action of cofactors of anaphylaxis, these data pave the way to unscramble the complex interactions determining the clinical relevance of sensitizations, the risk of anaphylaxis, and the severity of reactions. As a consequence, this understanding allows to better determine the individual risk in response to an identified allergen and results in more specific advices and education for our patients to prevent further life-threatening anaphylactic reactions.Allergy 11/2013; 68(12). DOI:10.1111/all.12322 · 6.00 Impact Factor
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ABSTRACT: BACKGROUND: Wheal reactions to intradermally injected neuropeptides, such as substance P (SP) and vasoactive intestinal peptide, are significantly larger and longer lasting in patients with chronic urticaria (CU) than in nonatopic control (NC) subjects. Mas-related gene X2 (MrgX2) has been identified as a receptor for basic neuropeptides, such as SP and vasoactive intestinal peptide. Mast cell (MC) responsiveness to eosinophil mediators contributes to the late-phase reaction of allergy. OBJECTIVE: We sought to compare the frequency of MrgX2 expression in skin MCs from patients with CU and NC subjects and to identify the receptor for basic eosinophil granule proteins on human skin MCs. METHODS: MrgX2 expression was investigated by using immunofluorescence in skin tissues from NC subjects and patients with severe CU and on skin-derived cultured MCs. MrgX2 expression in human MCs was reduced by using a lentiviral small hairpin RNA silencing technique. Ca(2+) influx was measured in CHO cells transfected with MrgX2 in response to eosinophil granule proteins. Histamine and prostaglandin D2 levels were measured by using enzyme immunoassays. RESULTS: The number of MrgX2(+) skin MCs and the percentage of MrgX2(+) MCs in all MCs in patients with CU were significantly greater than those in NC subjects. Eosinophil infiltration in urticarial lesions was observed in 7 of 9 patients with CU. SP, major basic protein, and eosinophil peroxidase, but not eosinophil-derived neurotoxin, induced histamine release from human skin MCs through MrgX2. CONCLUSION: MrgX2 might be a new target molecule for the treatment of wheal reactions in patients with severe CU.Journal of Allergy and Clinical Immunology 06/2014; 134(3):622-633. DOI:10.1016/j.jaci.2014.05.004 · 11.25 Impact Factor