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1.
Nat Immunol ; 18(8): 911-920, 2017 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-28628091

RESUMEN

Developing pre-B cells in the bone marrow alternate between proliferation and differentiation phases. We found that protein arginine methyl transferase 1 (PRMT1) and B cell translocation gene 2 (BTG2) are critical components of the pre-B cell differentiation program. The BTG2-PRMT1 module induced a cell-cycle arrest of pre-B cells that was accompanied by re-expression of Rag1 and Rag2 and the onset of immunoglobulin light chain gene rearrangements. We found that PRMT1 methylated cyclin-dependent kinase 4 (CDK4), thereby preventing the formation of a CDK4-Cyclin-D3 complex and cell cycle progression. Moreover, BTG2 in concert with PRMT1 efficiently blocked the proliferation of BCR-ABL1-transformed pre-B cells in vitro and in vivo. Our results identify a key molecular mechanism by which the BTG2-PRMT1 module regulates pre-B cell differentiation and inhibits pre-B cell leukemogenesis.


Asunto(s)
Proliferación Celular/genética , Ciclina D3/metabolismo , Quinasa 4 Dependiente de la Ciclina/metabolismo , Proteínas Inmediatas-Precoces/genética , Linfopoyesis/genética , Células Precursoras de Linfocitos B/metabolismo , Proteína-Arginina N-Metiltransferasas/genética , Proteínas Supresoras de Tumor/genética , Animales , Puntos de Control del Ciclo Celular , Diferenciación Celular/genética , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Citometría de Flujo , Técnicas de Silenciamiento del Gen , Reordenamiento Génico de Linfocito B/genética , Genes abl/genética , Proteínas de Homeodominio/genética , Proteínas de Homeodominio/metabolismo , Proteínas Inmediatas-Precoces/metabolismo , Cadenas Ligeras de Inmunoglobulina/genética , Espectrometría de Masas , Ratones , Células Precursoras de Linfocitos B/citología , Proteína-Arginina N-Metiltransferasas/metabolismo , ARN Interferente Pequeño , Reacción en Cadena en Tiempo Real de la Polimerasa , Proteínas Supresoras de Tumor/metabolismo
2.
Cell ; 158(1): 25-40, 2014 Jul 03.
Artículo en Inglés | MEDLINE | ID: mdl-24995976

RESUMEN

Obesity and diabetes affect more than half a billion individuals worldwide. Interestingly, the two conditions do not always coincide and the molecular determinants of "healthy" versus "unhealthy" obesity remain ill-defined. Chronic metabolic inflammation (metaflammation) is believed to be pivotal. Here, we tested a hypothesized anti-inflammatory role for heme oxygenase-1 (HO-1) in the development of metabolic disease. Surprisingly, in matched biopsies from "healthy" versus insulin-resistant obese subjects we find HO-1 to be among the strongest positive predictors of metabolic disease in humans. We find that hepatocyte and macrophage conditional HO-1 deletion in mice evokes resistance to diet-induced insulin resistance and inflammation, dramatically reducing secondary disease such as steatosis and liver toxicity. Intriguingly, cellular assays show that HO-1 defines prestimulation thresholds for inflammatory skewing and NF-κB amplification in macrophages and for insulin signaling in hepatocytes. These findings identify HO-1 inhibition as a potential therapeutic strategy for metabolic disease.


Asunto(s)
Hemo-Oxigenasa 1/metabolismo , Resistencia a la Insulina , Proteínas de la Membrana/metabolismo , Obesidad/complicaciones , Tejido Adiposo/metabolismo , Animales , Dieta Alta en Grasa , Hepatocitos/metabolismo , Humanos , Inflamación/metabolismo , Hígado/metabolismo , Macrófagos/metabolismo , Enfermedades Metabólicas/metabolismo , Enfermedades Metabólicas/fisiopatología , Ratones , Ratones Noqueados , Obesidad/fisiopatología , Especies Reactivas de Oxígeno/metabolismo
3.
Blood ; 134(18): 1510-1516, 2019 10 31.
Artículo en Inglés | MEDLINE | ID: mdl-31501153

RESUMEN

Dysregulated immune responses are essential underlying causes of a plethora of pathologies including cancer, autoimmunity, and immunodeficiency. We here investigated 4 patients from unrelated families presenting with immunodeficiency, autoimmunity, and malignancy. We identified 4 distinct homozygous mutations in TNFRSF9 encoding the tumor necrosis factor receptor superfamily member CD137/4-1BB, leading to reduced, or loss of, protein expression. Lymphocytic responses crucial for immune surveillance, including activation, proliferation, and differentiation, were impaired. Genetic reconstitution of CD137 reversed these defects. CD137 deficiency is a novel inborn error of human immunity characterized by lymphocytic defects with early-onset Epstein-Barr virus (EBV)-associated lymphoma. Our findings elucidate a functional role and relevance of CD137 in human immune homeostasis and antitumor responses.


Asunto(s)
Enfermedades Autoinmunes/genética , Síndromes de Inmunodeficiencia/genética , Linfoma/genética , Miembro 9 de la Superfamilia de Receptores de Factores de Necrosis Tumoral/genética , Enfermedades Autoinmunes/inmunología , Femenino , Predisposición Genética a la Enfermedad , Humanos , Síndromes de Inmunodeficiencia/inmunología , Linfoma/inmunología , Masculino , Linaje , Miembro 9 de la Superfamilia de Receptores de Factores de Necrosis Tumoral/deficiencia
4.
Haematologica ; 104(3): 609-621, 2019 03.
Artículo en Inglés | MEDLINE | ID: mdl-30309848

RESUMEN

Hyper-IgE syndromes comprise a group of inborn errors of immunity. STAT3-deficient hyper-IgE syndrome is characterized by elevated serum IgE levels, recurrent infections and eczema, and characteristic skeletal anomalies. A loss-of-function biallelic mutation in IL6ST encoding the GP130 receptor subunit (p.N404Y) has very recently been identified in a singleton patient (herein referred to as PN404Y) as a novel etiology of hyper-IgE syndrome. Here, we studied a patient with hyper-IgE syndrome caused by a novel homozygous mutation in IL6ST (p.P498L; patient herein referred to as PP498L) leading to abrogated GP130 signaling after stimulation with IL-6 and IL-27 in peripheral blood mononuclear cells as well as IL-6 and IL-11 in fibroblasts. Extending the initial identification of selective GP130 deficiency, we aimed to dissect the effects of aberrant cytokine signaling on T-helper cell differentiation in both patients. Our results reveal the importance of IL-6 signaling for the development of CCR6-expressing memory CD4+ T cells (including T-helper 17-enriched subsets) and non-conventional CD8+T cells which were reduced in both patients. Downstream functional analysis of the GP130 mutants (p.N404Y and p.P498L) have shown differences in response to IL-27, with the p.P498L mutation having a more severe effect that is reflected by reduced T-helper 1 cells in this patient (PP498L) only. Collectively, our data suggest that characteristic features of GP130-deficient hyper-IgE syndrome phenotype are IL-6 and IL-11 dominated, and indicate selective roles of aberrant IL-6 and IL-27 signaling on the differentiation of T-cell subsets.


Asunto(s)
Receptor gp130 de Citocinas/genética , Síndrome de Job/diagnóstico , Síndrome de Job/etiología , Mutación con Pérdida de Función , Subgrupos de Linfocitos T/inmunología , Subgrupos de Linfocitos T/metabolismo , Biomarcadores , Diferenciación Celular/genética , Niño , Preescolar , Receptor gp130 de Citocinas/química , Análisis Mutacional de ADN , Susceptibilidad a Enfermedades , Estudios de Asociación Genética , Humanos , Inmunofenotipificación , Síndrome de Job/metabolismo , Activación de Linfocitos , Masculino , Modelos Moleculares , Linaje , Fenotipo , Conformación Proteica , Radiografía
5.
J Immunol ; 193(1): 268-76, 2014 Jul 01.
Artículo en Inglés | MEDLINE | ID: mdl-24899508

RESUMEN

Signal transduction from the BCR is regulated by the equilibrium between kinases (e.g., spleen tyrosine kinase [Syk]) and phosphatases (e.g., Shp-1). Previous studies showed that Syk-deficient B cells have a developmental block at the pro/pre-B cell stage, whereas a B cell-specific Shp-1 deficiency promoted B-1a cell development and led to autoimmunity. We generated B cell-specific Shp-1 and Syk double-knockout (DKO) mice and compared them to the single-knockout mice deficient for either Syk or Shp-1. Unlike Syk-deficient mice, the DKO mice can generate mature B cells, albeit at >20-fold reduced B cell numbers. The DKO B-2 cells are all Syk-negative, whereas the peritoneal B1 cells of the DKO mice still express Syk, indicating that they require this kinase for their proper development. The DKO B-2 cells cannot be stimulated via the BCR, whereas they are efficiently activated via TLR or CD40. We also found that in DKO pre-B cells, the kinase Zap70 is associated with the pre-BCR, suggesting that Zap70 is important to promote B cell maturation in the absence of Syk and SHP-1. Together, our data show that a properly balanced kinase/phosphatase equilibrium is crucial for normal B cell development and function.


Asunto(s)
Subgrupos de Linfocitos B/inmunología , Péptidos y Proteínas de Señalización Intracelular/inmunología , Proteína Tirosina Fosfatasa no Receptora Tipo 6/inmunología , Proteínas Tirosina Quinasas/inmunología , Transducción de Señal/inmunología , Animales , Subgrupos de Linfocitos B/citología , Péptidos y Proteínas de Señalización Intracelular/genética , Ratones , Ratones Noqueados , Proteína Tirosina Fosfatasa no Receptora Tipo 6/genética , Proteínas Tirosina Quinasas/genética , Transducción de Señal/genética , Quinasa Syk , Proteína Tirosina Quinasa ZAP-70/genética , Proteína Tirosina Quinasa ZAP-70/inmunología
6.
Int Immunol ; 22(2): 71-80, 2010 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-19951957

RESUMEN

Fcgamma receptors (FcgammaRs) play an essential role in the regulation of immune response due to their ability to bind immune complexes. Activating FcgammaRs may facilitate antigen presentation and dendritic-cell maturation, while in the late phase of the immune response, the inhibitory FcgammaRIIb may down-regulate B-cell activation upon cross-linking with activating receptors. In this study, we investigated the in vivo role of FcgammaRs on the modulation of humoral immune response. In order to get well-defined immune complexes that can bind to both the activating and the inhibitory FcgammaRs, we designed a mono-biotinylated single-chain fragment variable construct from the rat anti-mouse CD16/32 clone 2.4G2, linked to avidin-FITC, and tested its effect on the FITC-hapten-specific T-independent type 2 (TI-2) and T-dependent (TD) immune response. When injected intravenously in mice, the complex bound to a small portion of B220+, CD11b(high) and CD11c(high) cells and was localized in the spleen on marginal zone macrophages 15 min after treatment. When applied as a booster following primary immunization with TI-2 (FITC-dextran) or TD (FITC-keyhole limpet haemocyanin) antigens, the complex elevated the number of hapten-specific IgM/IgG-producing B cells. This effect was diminished in CD16KO mice, suggesting that the activating-type FcgammaRIII might be a key mediator of this mechanism.


Asunto(s)
Células Productoras de Anticuerpos/inmunología , Avidina/inmunología , Fluoresceína-5-Isotiocianato/análogos & derivados , Inmunidad Humoral , Receptores de IgG/inmunología , Anticuerpos de Cadena Única/inmunología , Animales , Células Productoras de Anticuerpos/efectos de los fármacos , Linfocitos B/inmunología , Biotinilación , Citocinas/metabolismo , Dextranos/administración & dosificación , Dextranos/inmunología , Fluoresceína-5-Isotiocianato/administración & dosificación , Hemocianinas/administración & dosificación , Hemocianinas/inmunología , Hibridomas , Inmunidad Humoral/efectos de los fármacos , Inyecciones Intravenosas , Macrófagos/inmunología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Unión Proteica , Ratas , Receptores de IgG/deficiencia , Receptores de IgG/genética , Proteínas Recombinantes de Fusión/inmunología , Anticuerpos de Cadena Única/administración & dosificación , Anticuerpos de Cadena Única/genética , Bazo/inmunología , Factores de Tiempo
7.
Life Sci Alliance ; 4(11)2021 11.
Artículo en Inglés | MEDLINE | ID: mdl-34526379

RESUMEN

B cell antigen receptor (BCR) signaling is initiated by protein kinases and limited by counteracting phosphatases that currently are less well studied in their regulation of BCR signaling. Here, we used the B cell line Ramos to identify and quantify human B cell signaling components. Specifically, a protein tyrosine phosphatase profiling revealed a high expression of the protein tyrosine phosphatase 1B (PTP1B) in Ramos and human naïve B cells. The loss of PTP1B leads to increased B cell activation. Through substrate trapping in combination with quantitative mass spectrometry, we identified 22 putative substrates or interactors of PTP1B. We validated Igα, CD22, PLCγ1/2, CBL, BCAP, and APLP2 as specific substrates of PTP1B in Ramos B cells. The tyrosine kinase BTK and the two adaptor proteins GRB2 and VAV1 were identified as direct binding partners and potential substrates of PTP1B. We showed that PTP1B dephosphorylates the inhibitory receptor protein CD22 at phosphotyrosine 807. We conclude that PTP1B negatively modulates BCR signaling by dephosphorylating distinct phosphotyrosines in B cell-specific receptor proteins and various downstream signaling components.


Asunto(s)
Proteína Tirosina Fosfatasa no Receptora Tipo 1/metabolismo , Receptores de Antígenos de Linfocitos B/metabolismo , Linfocitos B/metabolismo , Línea Celular , Proteína Adaptadora GRB2/metabolismo , Espectrometría de Masas/métodos , Fosforilación , Unión Proteica , Proteína Tirosina Fosfatasa no Receptora Tipo 1/genética , Proteína Tirosina Fosfatasa no Receptora Tipo 1/fisiología , Proteínas Tirosina Quinasas/metabolismo , Proteómica/métodos , Proteínas Proto-Oncogénicas c-vav/metabolismo , Receptores de Antígenos de Linfocitos B/fisiología , Lectina 2 Similar a Ig de Unión al Ácido Siálico/metabolismo , Transducción de Señal/genética
8.
Sci Immunol ; 5(49)2020 07 10.
Artículo en Inglés | MEDLINE | ID: mdl-32646852

RESUMEN

The WAVE regulatory complex (WRC) is crucial for assembly of the peripheral branched actin network constituting one of the main drivers of eukaryotic cell migration. Here, we uncover an essential role of the hematopoietic-specific WRC component HEM1 for immune cell development. Germline-encoded HEM1 deficiency underlies an inborn error of immunity with systemic autoimmunity, at cellular level marked by WRC destabilization, reduced filamentous actin, and failure to assemble lamellipodia. Hem1-/- mice display systemic autoimmunity, phenocopying the human disease. In the absence of Hem1, B cells become deprived of extracellular stimuli necessary to maintain the strength of B cell receptor signaling at a level permissive for survival of non-autoreactive B cells. This shifts the balance of B cell fate choices toward autoreactive B cells and thus autoimmunity.


Asunto(s)
Enfermedades Autoinmunes/inmunología , Autoinmunidad/inmunología , Linfocitos B/inmunología , Proteínas de la Membrana/inmunología , Animales , Enfermedades Autoinmunes/genética , Trasplante de Médula Ósea , Línea Celular , Niño , Citoesqueleto , Femenino , Humanos , Lactante , Proteínas de la Membrana/genética , Ratones Endogámicos C57BL , Ratones Noqueados , Linfocitos T/inmunología
9.
Nat Commun ; 10(1): 3106, 2019 07 15.
Artículo en Inglés | MEDLINE | ID: mdl-31308374

RESUMEN

Immune responses need to be controlled tightly to prevent autoimmune diseases, yet underlying molecular mechanisms remain partially understood. Here, we identify biallelic mutations in three patients from two unrelated families in differentially expressed in FDCP6 homolog (DEF6) as the molecular cause of an inborn error of immunity with systemic autoimmunity. Patient T cells exhibit impaired regulation of CTLA-4 surface trafficking associated with reduced functional CTLA-4 availability, which is replicated in DEF6-knockout Jurkat cells. Mechanistically, we identify the small GTPase RAB11 as an interactor of the guanine nucleotide exchange factor DEF6, and find disrupted binding of mutant DEF6 to RAB11 as well as reduced RAB11+CTLA-4+ vesicles in DEF6-mutated cells. One of the patients has been treated with CTLA-4-Ig and achieved sustained remission. Collectively, we uncover DEF6 as player in immune homeostasis ensuring availability of the checkpoint protein CTLA-4 at T-cell surface, identifying a potential target for autoimmune and/or cancer therapy.


Asunto(s)
Antígeno CTLA-4/metabolismo , Proteínas de Unión al ADN/deficiencia , Factores de Intercambio de Guanina Nucleótido/deficiencia , Enfermedades de Inmunodeficiencia Primaria/genética , Antígeno B7-1/metabolismo , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/inmunología , Técnicas de Inactivación de Genes , Factores de Intercambio de Guanina Nucleótido/genética , Factores de Intercambio de Guanina Nucleótido/inmunología , Homeostasis , Humanos , Células Jurkat , Linfocitos T/metabolismo , Linfocitos T/fisiología , Proteínas de Unión al GTP rab/genética , Proteínas de Unión al GTP rab/metabolismo
11.
Immunol Lett ; 104(1-2): 76-82, 2006 Apr 15.
Artículo en Inglés | MEDLINE | ID: mdl-16386802

RESUMEN

The Grb2 associated binder (Gab) adaptor/scaffolding protein family comprises conserved proteins: mammalian Gab1, Gab2 and Gab3, Drosophila Dos and Caenorhabditis elegans Soc1. Gab adaptors are involved in multiple signaling pathways mediated by receptor- and non-receptor protein tyrosine kinases (PTKs), and become phosphorylated upon stimulation by growth factors-, cytokines-, Ig Fc- and antigen receptors. Through its phosphorylated tyrosine containing motifs, proline-rich sequences and pleckstrin homologue (PH) domain Gab adaptors may generate an interacting platform for proteins with SH2 and SH3 domains and may transfer these molecules to the plasma membrane, thereby contributing to their activation. This review will concentrate on the function of mammalian Gab proteins in the signal transduction triggered by immune receptors.


Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/metabolismo , Linfocitos/inmunología , Transducción de Señal/inmunología , Proteínas Adaptadoras Transductoras de Señales/análisis , Proteínas Adaptadoras Transductoras de Señales/genética , Animales , Membrana Celular/química , Membrana Celular/metabolismo , Humanos , Ratones , Fosforilación , Receptores Inmunológicos/metabolismo
12.
Neurochem Int ; 49(1): 94-103, 2006 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-16515823

RESUMEN

This is the first study to demonstrate that the interaction between beta-adrenoceptor activation, and the production of inflammatory mediators can be modulated in opposite ways by two inflammatory stimuli, namely, protein kinase C (PKC)-activating phorbol myristyl acetate (PMA) and lipopolysaccharide (LPS). We provided evidence that isoproterenol treatment, when combined with phorbol ester increased the production of tumor necrosis factor-alpha, interleukin-12, and nitric oxide in murine macrophages, as well as in human monocytes and differentiated PLB-985 cells, while in agreement with earlier findings, it decreased inflammatory mediator production in combination with LPS stimulation. The contrasting effect on inflammatory mediator production, shown for the PMA and LPS activated cells was accompanied by parallel changes in activation of ERK1/2 and p38 MAPKs. Thus, isoproterenol significantly increased MAPK activation (phosphorylation) in PMA-treated cells and, conversely, it decreased the activation of extracellular signal regulated kinase 1/2 (ERK1/2) and p38 in LPS-stimulated cells. The opposing effects of isoproterenol on LPS-induced versus PMA-induced mediator production and the concurrent changes in MAPK activation highlight the role of this kinase pathway in macrophage activation and provide new insights regarding the flexible ways through which beta-adrenoceptor stimulation can modulate the inflammatory response in macrophages. Our results challenge the dogma that beta-adrenoceptor signaling is only immunosuppressive, and offer potential opportunities for new therapeutic approaches in the treatment of inflammatory and autoimmune diseases.


Asunto(s)
Mediadores de Inflamación/inmunología , Sistema de Señalización de MAP Quinasas/inmunología , Macrófagos/inmunología , Neuroinmunomodulación/inmunología , Receptores Adrenérgicos beta/inmunología , Agonistas Adrenérgicos beta/farmacología , Animales , Carcinógenos/farmacología , Línea Celular , Células Cultivadas , Activación Enzimática/efectos de los fármacos , Activación Enzimática/inmunología , Quinasas MAP Reguladas por Señal Extracelular/efectos de los fármacos , Quinasas MAP Reguladas por Señal Extracelular/inmunología , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Humanos , Mediadores de Inflamación/metabolismo , Interleucina-12/inmunología , Interleucina-12/metabolismo , Isoproterenol/farmacología , Lipopolisacáridos/farmacología , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Macrófagos/efectos de los fármacos , Macrófagos/metabolismo , Ratones , Ratones Endogámicos BALB C , Neuroinmunomodulación/efectos de los fármacos , Óxido Nítrico/metabolismo , Fosforilación/efectos de los fármacos , Receptores Adrenérgicos beta/efectos de los fármacos , Receptores Adrenérgicos beta/metabolismo , Acetato de Tetradecanoilforbol/farmacología , Factor de Necrosis Tumoral alfa/inmunología , Factor de Necrosis Tumoral alfa/metabolismo , Regulación hacia Arriba/efectos de los fármacos , Regulación hacia Arriba/inmunología , Proteínas Quinasas p38 Activadas por Mitógenos/efectos de los fármacos , Proteínas Quinasas p38 Activadas por Mitógenos/inmunología , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismo
13.
Ann N Y Acad Sci ; 1090: 326-31, 2006 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-17384277

RESUMEN

RNA silencing experiments showed that knocking down Gab1 adaptor protein in BL41 human Burkitt lymphoma cells significantly reduced B cell receptor (BCR)-induced Erk phosphorylation, indicating that Gab1 plays a pivotal role in regulating Erk activity in B cells.


Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/fisiología , Linfocitos B/enzimología , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Transducción de Señal , Linfocitos B/metabolismo , Línea Celular Tumoral , Humanos , Fosforilación
14.
J Mol Med (Berl) ; 93(2): 143-58, 2015 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-25627575

RESUMEN

It is becoming more and more accepted that, in addition to producing autoantibodies, B lymphocytes have other important functions that influence the development of autoimmunity. For example, autoreactive B cells are able to produce inflammatory cytokines and activate pathogenic T cells. B lymphocytes can react to extracellular signals with a range of responses from anergy to autoreactivity. The final outcome is determined by the relative contribution of signaling events mediated by activating and inhibitory pathways. Besides the B cell antigen receptor (BCR), several costimulatory receptors expressed on B cells can also induce B cell proliferation and survival, or regulate antibody production. These include CD19, CD40, the B cell activating factor receptor, and Toll-like receptors. Hyperactivity of these receptors clearly contributes to breaking B-cell tolerance in several autoimmune diseases. Inhibitors of these activating signals (including protein tyrosine phosphatases, deubiquitinating enzymes and several adaptor proteins) are crucial to control B-cell activation and maintain B-cell tolerance. In this review, we summarize the inhibitory signaling mechanisms that counteract B-cell activation triggered by the BCR and the coreceptors.


Asunto(s)
Linfocitos B/inmunología , Linfocitos B/metabolismo , Tolerancia Inmunológica/inmunología , Inmunomodulación , Activación de Linfocitos/inmunología , Transducción de Señal , Animales , Enfermedades Autoinmunes/tratamiento farmacológico , Enfermedades Autoinmunes/inmunología , Enfermedades Autoinmunes/metabolismo , Linfocitos B/efectos de los fármacos , Humanos , Inmunomodulación/efectos de los fármacos , Terapia Molecular Dirigida , Receptores de Antígenos de Linfocitos B/metabolismo , Receptores Inmunológicos/metabolismo
15.
Immunol Lett ; 92(1-2): 83-90, 2004 Mar 29.
Artículo en Inglés | MEDLINE | ID: mdl-15081531

RESUMEN

Type IIb Fcgamma receptors (FcgammaRIIb) have a major role in regulating B cell activation. Upon its co-aggregation with the B cell receptors (BCR) via immune complexes FcgammaRIIb become phosphorylated on tyrosine within its immunoreceptor tyrosine based inhibitory motif (ITIM) and in turn recruit protein- and inositol phosphatases, inhibiting thereby signal transduction. The intracellular domain of the human FcgammaRIIb has a membrane proximal motif that is very similar to those of MAPK docking site in MAPK-interacting molecules. Additionally, in contrast to the mouse, a serine residue is located next to this motif that is a potential phosphorylation site for Ser/Thr kinases. Our aim was to study the role of the putative MAPK docking motif on FcgammaRIIb mediated function. We report here that MAPKs bind to FcgammaRIIb affinity purified from the detergent extracts of anti-IgM activated and BCR-FcgammaRIIb co-clustered B cells. We detected extracellular signal regulated kinase (ERK) activity in FcgammaRIIb immunoprecipitates and identified the bound proteins as 85, 44 and 42kDa ERKs by Western blots. Active ERKs bound to the synthetic peptide representing the putative docking site of FcgammaRIIb on a Ser/Thr phosphatase dependent manner. The FcgammaRIIb-associated ERKs may phosphorylate the membrane proximal serine of the receptor. We examined the consequences of serine phosphorylation by comparing the proteins that interact with synthetic peptides comprising the combined sequences of the MAPK docking site and the ITIM either in phosphorylated or in non-phosphorylated forms. The results indicate that phosphorylation on serine modifies the binding of Lyn to FcgammaRIIb, thus might negatively regulate phosphorylation of ITIM.


Asunto(s)
Antígenos CD/inmunología , Proteínas Quinasas Activadas por Mitógenos/inmunología , Receptores de IgG/inmunología , Secuencias de Aminoácidos , Antígenos CD/metabolismo , Humanos , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Fragmentos de Péptidos/inmunología , Fragmentos de Péptidos/metabolismo , Fosforilación , Receptores de IgG/metabolismo , Serina/metabolismo
16.
J Exp Med ; 211(3): 427-40, 2014 Mar 10.
Artículo en Inglés | MEDLINE | ID: mdl-24590766

RESUMEN

Tyrosine phosphorylation of signaling molecules that mediate B cell activation in response to various stimuli is tightly regulated by protein tyrosine phosphatases (PTPs). PTP1B is a ubiquitously expressed tyrosine phosphatase with well-characterized functions in metabolic signaling pathways. We show here that PTP1B negatively regulates CD40, B cell activating factor receptor (BAFF-R), and TLR4 signaling in B cells. Specifically, PTP1B counteracts p38 mitogen-activated protein kinase (MAPK) activation by directly dephosphorylating Tyr(182) of this kinase. Mice with a B cell-specific PTP1B deficiency show increased T cell-dependent immune responses and elevated total serum IgG. Furthermore, aged animals develop systemic autoimmunity with elevated serum anti-dsDNA, spontaneous germinal centers in the spleen, and deposition of IgG immune complexes and C3 in the kidney. In a clinical setting, we observed that B cells of rheumatoid arthritis patients have significantly reduced PTP1B expression. Our data suggest that PTP1B plays an important role in the control of B cell activation and the maintenance of immunological tolerance.


Asunto(s)
Artritis Reumatoide/inmunología , Autoinmunidad/inmunología , Linfocitos B/inmunología , Antígenos CD40/metabolismo , Proteína Tirosina Fosfatasa no Receptora Tipo 1/inmunología , Transducción de Señal/inmunología , Adulto , Anciano , Anciano de 80 o más Años , Animales , Antirreumáticos/farmacología , Receptor del Factor Activador de Células B/metabolismo , Western Blotting , Proliferación Celular , Ensayo de Inmunoadsorción Enzimática , Femenino , Citometría de Flujo , Humanos , Inmunohistoquímica , Masculino , Ratones , Persona de Mediana Edad , Plásmidos/genética , Proteína Tirosina Fosfatasa no Receptora Tipo 1/metabolismo , Reacción en Cadena en Tiempo Real de la Polimerasa , Transfección
17.
J Leukoc Biol ; 93(4): 537-47, 2013 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-23362305

RESUMEN

B cells acquire membrane-bound cognate antigens from the surface of the APCs by forming an IS, similar to that seen in T cells. Recognition of membrane-bound antigens on the APCs initiates adhesion of B lymphocytes to the antigen-tethered surface, which is followed by the formation of radial lamellipodia-like structures, a process known as B cell spreading. The spreading response requires the rearrangement of the submembrane actin cytoskeleton and is regulated mainly via signals transmitted by the BCR. Here, we show that cytoplasmic calcium is a regulator of actin cytoskeleton dynamics in B lymphocytes. We find that BCR-induced calcium mobilization is indispensible for adhesion and spreading of B cells and that PLCγ and CRAC-mediated calcium mobilization are critical regulators of these processes. Measuring calcium and actin dynamics in live cells, we found that a generation of actin-based membrane protrusion is strongly linked to the dynamics of a cytoplasmic-free calcium level. Finally, we demonstrate that PLCγ and CRAC channels regulate the activity of actin-severing protein cofilin, linking BCR-induced calcium signaling to the actin dynamics.


Asunto(s)
Citoesqueleto de Actina/metabolismo , Actinas/metabolismo , Canales de Calcio/metabolismo , Calcio/metabolismo , Receptores de Antígenos de Linfocitos B/metabolismo , Citoesqueleto de Actina/genética , Citoesqueleto de Actina/inmunología , Actinas/genética , Actinas/inmunología , Animales , Presentación de Antígeno , Linfocitos B/citología , Linfocitos B/inmunología , Linfocitos B/metabolismo , Canales de Calcio/genética , Canales de Calcio/inmunología , Adhesión Celular , Línea Celular Tumoral , Movimiento Celular , Cofilina 1/genética , Cofilina 1/inmunología , Cofilina 1/metabolismo , Regulación de la Expresión Génica/inmunología , Vectores Genéticos , Lentivirus/genética , Ratones , Fosfolipasa C gamma/genética , Fosfolipasa C gamma/inmunología , Fosfolipasa C gamma/metabolismo , Seudópodos/inmunología , Seudópodos/metabolismo , Receptores de Antígenos de Linfocitos B/genética , Receptores de Antígenos de Linfocitos B/inmunología , Transducción de Señal , Transducción Genética
18.
Cell Signal ; 21(2): 220-7, 2009 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-18950707

RESUMEN

B-cell fate during maturation and the germinal center reaction is regulated through the strength and the duration of the B-cell receptor signal. Signaling pathways discriminating between apoptosis and survival in B cells are keys in understanding adaptive immunity. Gab2 is a member of the Gab/Dos adaptor protein family. It has been shown in several model systems that Gab/Dos family members may regulate both the anti-apoptotic PI3-K/Akt and the mitogenic Ras/MAPK pathways, still their role in B-cells have not been investigated in detail. Here we studied the role of Gab2 in B-cell receptor mediated signaling. We have shown that BCR crosslinking induces the marked phosphorylation of Gab2 through both Lyn and Syk kinases. Subsequently Gab2 recruits p85 regulatory subunit of PI3-K, and SHP-2. Our results revealed that Ig-alpha/Ig-beta, signal transducing unit of the B-cell receptor, may function as scaffold recruiting Gab2 to the signalosome. Overexpression of Gab2 in A20 cells demonstrated that Gab2 is a regulator of the PI3-K/Akt but not that of the Ras/MAPK pathway in B cells. Accordingly to the elevated Akt phosphorylation, overexpression of wild-type Gab2 in A20 cells suppressed Fas-mediated apoptosis, and enhanced BCR-mediated rescue from Fas-induced cell death. Although PH-domain has only a stabilizing effect on membrane recruitment of Gab2, it is indispensable in mediating its anti-apoptotic effect.


Asunto(s)
Linfocitos B/inmunología , Fosfoproteínas/metabolismo , Receptores de Antígenos de Linfocitos B/metabolismo , Proteínas Adaptadoras Transductoras de Señales , Animales , Anticuerpos Antiidiotipos/farmacología , Apoptosis , Sitios de Unión , Supervivencia Celular , Células Cultivadas , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Fosfatidilinositol 3-Quinasas/inmunología , Fosfatidilinositol 3-Quinasas/metabolismo , Fosfoproteínas/inmunología , Fosforilación , Transducción de Señal , Receptor fas/inmunología , Receptor fas/metabolismo , Familia-src Quinasas/metabolismo
19.
J Mol Recognit ; 17(2): 95-105, 2004.
Artículo en Inglés | MEDLINE | ID: mdl-15027030

RESUMEN

The IgG binding Fcgamma receptors (FcgammaRs) play a key role in defence against pathogens by linking humoral and cell-mediated immune responses. Impaired expression and/or function of FcgammaR may result in the development of pathological autoimmunity. Considering the functions of FcgammaRs, they are potential target molecules for drug design to aim at developing novel anti-inflammatory and immunomodulatory therapies. Previous data mostly obtained by X-ray analysis of ligand-receptor complexes indicate the profound role of the CH2 domain in binding to various FcgammaRs. Our aim was to localize linear segments, which are able to bind and also to modulate the function of the low affinity FcgammaRs, like FcgammaRIIb and FcgammaRIIIa. To this end a set of overlapping octapeptides was prepared corresponding to the 231-298 sequence of IgG1 CH2 domain and tested for binding to human recombinant soluble FcgammaRIIb. Based on these results, a second group of peptides was synthesized and their binding properties to recombinant soluble FcgammaRIIb, as well as to FcgammaRs expressed on the cell surface, was investigated. Here we report that peptide representing the Arg(255)-Ser(267) sequence of IgG1 is implicated in the binding to FcgammaRIIb. In addition we found that peptides corresponding to the Arg(255)-Ser(267), Lys(288)-Ser(298) or Pro(230)-Val(240) when presented in a multimeric form conjugated to branched chain polypeptide in uniformly oriented copies induced the release of TNFalpha, a pro-inflammatory cytokine from MonoMac monocyte cell line. These findings indicate that these conjugated peptides are able to cluster the activating FcgammaRs, and mediate FcgammaR dependent function. Peptide Arg(255)-Ser(267) can also be considered as a lead for further functional studies.


Asunto(s)
Antígenos CD/metabolismo , Inmunoglobulina G/metabolismo , Oligopéptidos/síntesis química , Oligopéptidos/metabolismo , Receptores de IgG/metabolismo , Secuencia de Aminoácidos , Sitios de Unión , Células Cultivadas , Cromatografía Líquida de Alta Presión/métodos , Dicroismo Circular/métodos , Ensayo de Inmunoadsorción Enzimática/métodos , Citometría de Flujo/métodos , Humanos , Espectrometría de Masas/métodos , Datos de Secuencia Molecular , Conformación Proteica
20.
Eur J Immunol ; 34(4): 1127-35, 2004 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-15048724

RESUMEN

Receptors specific for the Fc part of IgG (Fc gamma R) are expressed by several cell types and play diverse roles in immune responses. Impaired function of the activating and inhibitory Fc gamma R may result in autoimmunity. Thus, the modulation of IgG-Fc gamma R interaction can be a target for the development of treatments for some autoimmune and inflammatory diseases. This study addresses the localization and functional characterization of linear sequences in human IgG1 which bind to Fc gamma RII. Peptides with overlapping sequences derived from the CH2 domain of human IgG1 between P(234) and S(298) were synthesized and used in binding and functional experiments. Binding of the peptides to Fc gamma R was assayed in vitro and ex vivo, and peptides found to interact were functionally tested. The shortest effective peptide was T(256)-P(271), which bound to soluble recombinant Fc gamma RIIb with K(d)=6 x 10(6) M(-1). The biotinylated peptides R(255)-P(271) and T(256)-P(271) complexed by avidin exhibited functional activity; they induced Fc gamma RIIb-mediated inhibition of the BCR-triggered Ca(2+) response of human Burkitt lymphoma cells, and inflammatory cytokine production (TNF-alpha and IL-6) by the human monocyte cell line MonoMac. In conclusion, our results suggest that the selected peptides functionally represent the Fc gamma RII-binding part of IgG1.


Asunto(s)
Mapeo Epitopo , Inmunoglobulina G/química , Monocitos/inmunología , Péptidos/inmunología , Péptidos/metabolismo , Receptores de IgG/química , Sitios de Unión/inmunología , Western Blotting , Células Cultivadas , Electroforesis en Gel de Poliacrilamida , Ensayo de Inmunoadsorción Enzimática , Citometría de Flujo , Humanos , Inmunoglobulina G/inmunología , Interleucina-6/metabolismo , Proteínas Quinasas Activadas por Mitógenos/efectos de los fármacos , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Péptidos/farmacología , Fosforilación , Estructura Cuaternaria de Proteína , Receptores de Antígenos de Linfocitos B/inmunología , Receptores de IgG/genética , Factor de Necrosis Tumoral alfa/efectos de los fármacos , Factor de Necrosis Tumoral alfa/metabolismo
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