RESUMEN
BACKGROUND: Disabling pansclerotic morphea (DPM) is a rare systemic inflammatory disorder, characterized by poor wound healing, fibrosis, cytopenias, hypogammaglobulinemia, and squamous-cell carcinoma. The cause is unknown, and mortality is high. METHODS: We evaluated four patients from three unrelated families with an autosomal dominant pattern of inheritance of DPM. Genomic sequencing independently identified three heterozygous variants in a specific region of the gene that encodes signal transducer and activator of transcription 4 (STAT4). Primary skin fibroblast and cell-line assays were used to define the functional nature of the genetic defect. We also assayed gene expression using single-cell RNA sequencing of peripheral-blood mononuclear cells to identify inflammatory pathways that may be affected in DPM and that may respond to therapy. RESULTS: Genome sequencing revealed three novel heterozygous missense gain-of-function variants in STAT4. In vitro, primary skin fibroblasts showed enhanced interleukin-6 secretion, with impaired wound healing, contraction of the collagen matrix, and matrix secretion. Inhibition of Janus kinase (JAK)-STAT signaling with ruxolitinib led to improvement in the hyperinflammatory fibroblast phenotype in vitro and resolution of inflammatory markers and clinical symptoms in treated patients, without adverse effects. Single-cell RNA sequencing revealed expression patterns consistent with an immunodysregulatory phenotype that were appropriately modified through JAK inhibition. CONCLUSIONS: Gain-of-function variants in STAT4 caused DPM in the families that we studied. The JAK inhibitor ruxolitinib attenuated the dermatologic and inflammatory phenotype in vitro and in the affected family members. (Funded by the American Academy of Allergy, Asthma, and Immunology Foundation and others.).
Asunto(s)
Enfermedades Autoinmunes , Fármacos Dermatológicos , Quinasas Janus , Esclerodermia Sistémica , Quinasas Janus/antagonistas & inhibidores , Nitrilos , Pirazoles/uso terapéutico , Pirazoles/farmacología , Pirimidinas , Esclerodermia Sistémica/tratamiento farmacológico , Esclerodermia Sistémica/genética , Enfermedades Autoinmunes/tratamiento farmacológico , Enfermedades Autoinmunes/genética , Mutación Missense , Mutación con Ganancia de Función , Fármacos Dermatológicos/uso terapéutico , Antiinflamatorios/uso terapéuticoRESUMEN
The T cell receptor (TCR) signaling pathway is an ensemble of numerous proteins that are crucial for an adequate immune response. Disruption of any protein involved in this pathway leads to severe immunodeficiency and unfavorable clinical outcomes. Here, we describe an infant with severe immunodeficiency who was found to have novel biallelic mutations in SLP76. SLP76 is a key protein involved in TCR signaling and in other hematopoietic pathways. Previous studies of this protein were performed using Jurkat-derived human leukemic T cell lines and SLP76-deficient mice. Our current study links this gene, for the first time, to a human immunodeficiency characterized by early-onset life-threatening infections, combined T and B cell immunodeficiency, severe neutrophil defects, and impaired platelet aggregation. Hereby, we characterized aspects of the patient's immune phenotype, modeled them with an SLP76-deficient Jurkat-derived T cell line, and rescued some consequences using ectopic expression of wild-type SLP76. Understanding human diseases due to SLP76 deficiency is helpful in explaining the mixed T cell and neutrophil defects, providing a guide for exploring human SLP76 biology.
Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/deficiencia , Plaquetas/patología , Neutrófilos/patología , Fosfoproteínas/deficiencia , Inmunodeficiencia Combinada Grave/metabolismo , Inmunodeficiencia Combinada Grave/patología , Proteínas Adaptadoras Transductoras de Señales/química , Proteínas Adaptadoras Transductoras de Señales/genética , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Secuencia de Aminoácidos , Secuencia de Bases , Plaquetas/metabolismo , Resultado Fatal , Humanos , Lactante , Recién Nacido , Células Jurkat , Mutación/genética , Neutrófilos/metabolismo , Fenotipo , Fosfoproteínas/química , Fosfoproteínas/genética , Fosfoproteínas/metabolismo , Receptores de Antígenos de Linfocitos B/metabolismo , Receptores de Antígenos de Linfocitos T/metabolismo , Inmunodeficiencia Combinada Grave/inmunología , Transducción de SeñalRESUMEN
IL-6 excess is central to the pathogenesis of multiple inflammatory conditions and is targeted in clinical practice by immunotherapy that blocks the IL-6 receptor encoded by IL6R We describe two patients with homozygous mutations in IL6R who presented with recurrent infections, abnormal acute-phase responses, elevated IgE, eczema, and eosinophilia. This study identifies a novel primary immunodeficiency, clarifying the contribution of IL-6 to the phenotype of patients with mutations in IL6ST, STAT3, and ZNF341, genes encoding different components of the IL-6 signaling pathway, and alerts us to the potential toxicity of drugs targeting the IL-6R.
Asunto(s)
Síndromes de Inmunodeficiencia/patología , Inflamación/patología , Receptores de Interleucina-6/deficiencia , Adolescente , Adulto , Niño , Preescolar , Femenino , Células HEK293 , Humanos , Recién Nacido , Masculino , Receptores de Interleucina-6/metabolismoRESUMEN
MALT1 (mucosa-associated lymphoid tissue lymphoma-translocation gene 1) is an intracellular signaling protein that activates NFκB and is crucial for both the adaptive and innate immune responses. Only 6 patients with immune deficiencies secondary to inherited mutations in the MALT1 gene have been described. PURPOSE: To provide clinical and immunological insights from 2 patients diagnosed with MALT1 immunodeficiency syndrome due to a novel MALT1 mutation. METHODS: Two cousins with suspected combined immunodeficiency underwent immunological and genetic work-up, including lymphocyte phenotyping, lymphocyte activation by mitogen stimulation, and next-generation sequencing (NGS) of T cell receptor gamma chain (TRG) repertoire. Whole exome sequencing was performed to identify the underlying genetic defect. RESULTS: Clinical findings included recurrent infections, failure to thrive, lymphadenopathy, dermatitis, and autoimmunity. Immune work-up revealed lymphocytosis, low to normal levels of immunoglobulins, absence of regulatory T cells, and low Th17 cells. A normal proliferative response was induced by phytohemagglutinin and IL-2 but was diminished with anti-CD3. TRG repertoire was diverse with a clonal expansion pattern. Genetic analysis identified a novel autosomal recessive homozygous c.1799T>A; p. I600N missense mutation in MALT1. MALT1 protein expression was markedly reduced, and in vitro IL-2 production and NFκB signaling pathway were significantly impaired. CONCLUSIONS: Two patients harboring a novel MALT1 mutation presented with signs of immune deficiency and dysregulation and were found to have an abnormal T cell receptor repertoire. These findings reinforce the link between MALT1 deficiency and combined immunodeficiency. Early diagnosis is crucial, and curative treatment by hematopoietic stem cell transplantation may be warranted.
Asunto(s)
Predisposición Genética a la Enfermedad , Síndromes de Inmunodeficiencia/genética , Síndromes de Inmunodeficiencia/inmunología , Proteína 1 de la Translocación del Linfoma del Tejido Linfático Asociado a Mucosas/genética , Mutación , Receptores de Antígenos de Linfocitos T/genética , Receptores de Antígenos de Linfocitos T/metabolismo , Secuencia de Aminoácidos , Biomarcadores , Consanguinidad , Citocinas/metabolismo , Femenino , Estudios de Asociación Genética , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Síndromes de Inmunodeficiencia/diagnóstico , Síndromes de Inmunodeficiencia/metabolismo , Inmunofenotipificación , Masculino , FN-kappa B/metabolismo , Linaje , Subgrupos de Linfocitos T/inmunología , Subgrupos de Linfocitos T/metabolismoRESUMEN
BACKGROUND: Caspase activation and recruitment domain 11 (CARD11) encodes a scaffold protein in lymphocytes that links antigen receptor engagement with downstream signaling to nuclear factor κB, c-Jun N-terminal kinase, and mechanistic target of rapamycin complex 1. Germline CARD11 mutations cause several distinct primary immune disorders in human subjects, including severe combined immune deficiency (biallelic null mutations), B-cell expansion with nuclear factor κB and T-cell anergy (heterozygous, gain-of-function mutations), and severe atopic disease (loss-of-function, heterozygous, dominant interfering mutations), which has focused attention on CARD11 mutations discovered by using whole-exome sequencing. OBJECTIVES: We sought to determine the molecular actions of an extended allelic series of CARD11 and to characterize the expanding range of clinical phenotypes associated with heterozygous CARD11 loss-of-function alleles. METHODS: Cell transfections and primary T-cell assays were used to evaluate signaling and function of CARD11 variants. RESULTS: Here we report on an expanded cohort of patients harboring novel heterozygous CARD11 mutations that extend beyond atopy to include other immunologic phenotypes not previously associated with CARD11 mutations. In addition to (and sometimes excluding) severe atopy, heterozygous missense and indel mutations in CARD11 presented with immunologic phenotypes similar to those observed in signal transducer and activator of transcription 3 loss of function, dedicator of cytokinesis 8 deficiency, common variable immunodeficiency, neutropenia, and immune dysregulation, polyendocrinopathy, enteropathy, X-linked-like syndrome. Pathogenic variants exhibited dominant negative activity and were largely confined to the CARD or coiled-coil domains of the CARD11 protein. CONCLUSION: These results illuminate a broader phenotypic spectrum associated with CARD11 mutations in human subjects and underscore the need for functional studies to demonstrate that rare gene variants encountered in expected and unexpected phenotypes must nonetheless be validated for pathogenic activity.
Asunto(s)
Proteínas Adaptadoras de Señalización CARD/genética , Proteínas Adaptadoras de Señalización CARD/inmunología , Guanilato Ciclasa/genética , Guanilato Ciclasa/inmunología , Enfermedades del Sistema Inmune/genética , Enfermedades del Sistema Inmune/inmunología , Adulto , Femenino , Humanos , Masculino , Mutación , FenotipoRESUMEN
BACKGROUND: Atopic dermatitis (AD) is a highly prevalent chronic inflammatory skin disease that is known to be, at least in part, genetically determined. Mutations in caspase recruitment domain-containing protein 14 (CARD14) have been shown to result in various forms of psoriasis and related disorders. OBJECTIVE: We aimed to identify rare DNA variants conferring a significant risk for AD through genetic and functional studies in a cohort of patients affected with severe AD. METHODS: Whole-exome and direct gene sequencing, immunohistochemistry, real-time PCR, ELISA, and functional assays in human keratinocytes were used. RESULTS: In a cohort of patients referred with severe AD, DNA sequencing revealed in 4 patients 2 rare heterozygous missense mutations in the gene encoding CARD14, a major regulator of nuclear factor κB (NF-κB). A dual luciferase reporter assay demonstrated that both mutations exert a dominant loss-of-function effect and result in decreased NF-κB signaling. Accordingly, immunohistochemistry staining showed decreased expression of CARD14 in patients' skin, as well as decreased levels of activated p65, a surrogate marker for NF-κB activity. CARD14-deficient or mutant-expressing keratinocytes displayed abnormal secretion of key mediators of innate immunity. CONCLUSIONS: Although dominant gain-of-function mutations in CARD14 are associated with psoriasis and related diseases, loss-of-function mutations in the same gene are associated with a severe variant of AD.
Asunto(s)
Proteínas Adaptadoras de Señalización CARD , Dermatitis Atópica , Guanilato Ciclasa , Queratinocitos , Mutación con Pérdida de Función , Proteínas de la Membrana , Mutación Missense , Transducción de Señal/genética , Adolescente , Proteínas Adaptadoras de Señalización CARD/genética , Proteínas Adaptadoras de Señalización CARD/metabolismo , Dermatitis Atópica/genética , Dermatitis Atópica/metabolismo , Dermatitis Atópica/patología , Femenino , Guanilato Ciclasa/genética , Guanilato Ciclasa/metabolismo , Células HEK293 , Humanos , Queratinocitos/metabolismo , Queratinocitos/patología , Masculino , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Índice de Severidad de la Enfermedad , Factor de Transcripción ReIA/genética , Factor de Transcripción ReIA/metabolismoRESUMEN
This corrects the article DOI: 10.1038/ng.3898.
RESUMEN
Patients with hypomorphic mutations in STAT3 and patients with hypermorphic mutations in STAT1 share several clinical and cellular phenotypes suggesting overlapping pathophysiologic mechanisms. We, therefore, examined cytokine signaling and CD4+ T cell differentiation in these cohorts to characterize common pathways. As expected, differentiation of Th17 cells was impaired in both cohorts. We found that STAT1 was hyperphosphorylated in response to cytokine stimulation in both cohorts and that STAT1-dependent PD-L1 up-regulation-known to inhibit Th17 differentiation in mouse models-was markedly enhanced as well. Overexpression of SOCS3 strongly inhibited phosphorylation of STAT1 and PD-L1 up-regulation, suggesting that diminished SOCS3 expression may lead to the observed effects. Defects in Th17 differentiation could be partially overcome in vitro via PD-L1 inhibition and in a mouse model of STAT3 loss-of-function by crossing them with PD-1 knockout mice. PD-L1 may be a potential therapeutic target in several genetic diseases of immune deficiency affecting cytokine signaling.
Asunto(s)
Antígeno B7-H1/fisiología , Diferenciación Celular/fisiología , Factor de Transcripción STAT1/fisiología , Factor de Transcripción STAT3/fisiología , Células Th17/fisiología , Adolescente , Adulto , Animales , Niño , Citocinas/fisiología , Femenino , Humanos , Síndromes de Inmunodeficiencia/genética , Síndromes de Inmunodeficiencia/inmunología , Síndromes de Inmunodeficiencia/fisiopatología , Interleucinas/fisiología , Masculino , Ratones , Ratones Noqueados , Persona de Mediana Edad , Factor de Transcripción STAT1/genética , Factor de Transcripción STAT3/genética , Transducción de Señal/fisiología , Proteína 3 Supresora de la Señalización de Citocinas/fisiología , Regulación hacia Arriba , Adulto JovenRESUMEN
Few monogenic causes for severe manifestations of common allergic diseases have been identified. Through next-generation sequencing on a cohort of patients with severe atopic dermatitis with and without comorbid infections, we found eight individuals, from four families, with novel heterozygous mutations in CARD11, which encodes a scaffolding protein involved in lymphocyte receptor signaling. Disease improved over time in most patients. Transfection of mutant CARD11 expression constructs into T cell lines demonstrated both loss-of-function and dominant-interfering activity upon antigen receptor-induced activation of nuclear factor-κB and mammalian target of rapamycin complex 1 (mTORC1). Patient T cells had similar defects, as well as low production of the cytokine interferon-γ (IFN-γ). The mTORC1 and IFN-γ production defects were partially rescued by supplementation with glutamine, which requires CARD11 for import into T cells. Our findings indicate that a single hypomorphic mutation in CARD11 can cause potentially correctable cellular defects that lead to atopic dermatitis.
Asunto(s)
Proteínas Adaptadoras de Señalización CARD/genética , Dermatitis Atópica/genética , Mutación de Línea Germinal , Guanilato Ciclasa/genética , Sistema de Transporte de Aminoácidos ASC/metabolismo , Estudios de Cohortes , Análisis Mutacional de ADN , Dermatitis Atópica/inmunología , Femenino , Genes Dominantes , Glutamina/metabolismo , Humanos , Células Jurkat , Activación de Linfocitos , Masculino , Diana Mecanicista del Complejo 1 de la Rapamicina , Antígenos de Histocompatibilidad Menor/metabolismo , Complejos Multiproteicos/metabolismo , FN-kappa B/metabolismo , Linaje , Linfocitos T/inmunología , Linfocitos T/metabolismo , Serina-Treonina Quinasas TOR/metabolismoAsunto(s)
Eosinofilia/genética , Mutación , Factor de Transcripción STAT5/genética , Urticaria/genética , Edad de Inicio , Preescolar , Dermatitis/genética , Dermatitis/inmunología , Diarrea/genética , Eosinofilia/sangre , Eosinofilia/inmunología , Femenino , Humanos , Polimorfismo de Nucleótido Simple , Factor de Transcripción STAT5/inmunología , Síndrome , Linfocitos T/inmunología , Urticaria/inmunología , Urticaria/patologíaRESUMEN
NF-κB essential modulator (NEMO) and cylindromatosis protein (CYLD) are intracellular proteins that regulate the NF-κB signaling pathway. Although mice with either CYLD deficiency or an alteration in the zinc finger domain of NEMO (K392R) are born healthy, we found that the combination of these two gene defects in double mutant (DM) mice is early embryonic lethal but can be rescued by the absence of TNF receptor 1 (TNFR1). Notably, NEMO was not recruited into the TNFR1 complex of DM cells, and consequently NF-κB induction by TNF was severely impaired and DM cells were sensitized to TNF-induced cell death. Interestingly, the TNF signaling defects can be fully rescued by reconstitution of DM cells with CYLD lacking ubiquitin hydrolase activity but not with CYLD mutated in TNF receptor-associated factor 2 (TRAF2) or NEMO binding sites. Therefore, our data demonstrate an unexpected non-catalytic function for CYLD as an adapter protein between TRAF2 and the NEMO zinc finger that is important for TNF-induced NF-κB signaling during embryogenesis.
Asunto(s)
Desarrollo Embrionario/genética , Péptidos y Proteínas de Señalización Intracelular/genética , Receptores Tipo I de Factores de Necrosis Tumoral/genética , Proteínas Supresoras de Tumor/genética , Animales , Apoptosis/efectos de los fármacos , Apoptosis/genética , Western Blotting , Células Cultivadas , Embrión de Mamíferos/citología , Fibroblastos/citología , Fibroblastos/efectos de los fármacos , Fibroblastos/metabolismo , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Ratones Noqueados , FN-kappa B/genética , FN-kappa B/metabolismo , Unión Proteica , Receptores Tipo I de Factores de Necrosis Tumoral/metabolismo , Transducción de Señal/efectos de los fármacos , Transducción de Señal/genética , Factor 2 Asociado a Receptor de TNF/genética , Factor 2 Asociado a Receptor de TNF/metabolismo , Factor de Necrosis Tumoral alfa/farmacología , Proteínas Supresoras de Tumor/metabolismo , Dedos de Zinc/genéticaRESUMEN
Germline loss-of-function mutations in the transcription factor signal transducer and activator of transcription 3 (STAT3) cause immunodeficiency, whereas somatic gain-of-function mutations in STAT3 are associated with large granular lymphocytic leukemic, myelodysplastic syndrome, and aplastic anemia. Recently, germline mutations in STAT3 have also been associated with autoimmune disease. Here, we report on 13 individuals from 10 families with lymphoproliferation and early-onset solid-organ autoimmunity associated with 9 different germline heterozygous mutations in STAT3. Patients exhibited a variety of clinical features, with most having lymphadenopathy, autoimmune cytopenias, multiorgan autoimmunity (lung, gastrointestinal, hepatic, and/or endocrine dysfunction), infections, and short stature. Functional analyses demonstrate that these mutations confer a gain-of-function in STAT3 leading to secondary defects in STAT5 and STAT1 phosphorylation and the regulatory T-cell compartment. Treatment targeting a cytokine pathway that signals through STAT3 led to clinical improvement in 1 patient, suggesting a potential therapeutic option for such patients. These results suggest that there is a broad range of autoimmunity caused by germline STAT3 gain-of-function mutations, and that hematologic autoimmunity is a major component of this newly described disorder. Some patients for this study were enrolled in a trial registered at www.clinicaltrials.gov as #NCT00001350.
Asunto(s)
Enfermedades Autoinmunes/genética , Enfermedades Genéticas Congénitas/genética , Trastornos Linfoproliferativos/genética , Factor de Transcripción STAT3/genética , Adolescente , Adulto , Enfermedades Autoinmunes/inmunología , Enfermedades Autoinmunes/patología , Niño , Preescolar , Femenino , Enfermedades Genéticas Congénitas/inmunología , Enfermedades Genéticas Congénitas/patología , Humanos , Lactante , Trastornos Linfoproliferativos/inmunología , Trastornos Linfoproliferativos/patología , Masculino , Mutación , Fosforilación/genética , Fosforilación/inmunología , Factor de Transcripción STAT1/genética , Factor de Transcripción STAT1/inmunología , Factor de Transcripción STAT3/inmunología , Factor de Transcripción STAT5/genética , Factor de Transcripción STAT5/inmunología , Linfocitos T Reguladores/inmunología , Linfocitos T Reguladores/patologíaRESUMEN
Novel inhibitor of histone acetyltransferase repressor (NIR) is a transcriptional corepressor with inhibitor of histone acetyltransferase activity and is a potent suppressor of p53. Although NIR deficiency in mice leads to early embryonic lethality, lymphoid-restricted deletion resulted in the absence of double-positive CD4(+)CD8(+) thymocytes, whereas bone-marrow-derived B cells were arrested at the B220(+)CD19(-) pro-B-cell stage. V(D)J recombination was preserved in NIR-deficient DN3 double-negative thymocytes, suggesting that NIR does not affect p53 function in response to physiologic DNA breaks. Nevertheless, the combined deficiency of NIR and p53 provided rescue of DN3L double-negative thymocytes and their further differentiation to double- and single-positive thymocytes, whereas B cells in the marrow further developed to the B220(+)CD19(+) pro-B-cell stage. Our results show that NIR cooperate with p53 to impose checkpoint for the generation of mature B and T lymphocytes.
Asunto(s)
Diferenciación Celular/inmunología , Proteínas Represoras/inmunología , Timocitos/inmunología , Animales , Antígenos de Diferenciación/genética , Antígenos de Diferenciación/inmunología , Células de la Médula Ósea/citología , Células de la Médula Ósea/inmunología , Diferenciación Celular/genética , Roturas del ADN , Ratones , Células Precursoras de Linfocitos B/citología , Proteínas Represoras/genética , Timocitos/citología , Proteína p53 Supresora de Tumor/genética , Proteína p53 Supresora de Tumor/inmunologíaRESUMEN
Mutations in the TNF family of proteins have been associated with inherited forms of immune deficiency. Using an array-based sequencing assay, we identified an autosomal-dominant deficiency in TNF-like weak inducer of apoptosis (TWEAK; TNFSF12) in a kindred with recurrent infection and impaired antibody responses to protein and polysaccharide vaccines. This mutation occurs in the sixth exon of TWEAK and results in the amino acid substitution R145C within the conserved TNF-homology domain of the full-length protein. TWEAK mutant protein formed high molecular weight aggregates under nonreducing conditions, suggesting an increased propensity for intermolecular interactions. As a result, mutant TWEAK associated with B-cell-activating factor (BAFF) protein and down-regulated the BAFF-mediated activation of the noncanonical NF-κB pathway through inhibition of p100 processing to p52, resulting in inhibition of BAFF-dependent B-cell survival and proliferation. As BAFF mediates T-cell-independent isotype switching and B-cell survival, our data implicate TWEAK as a disease-susceptibility gene for a humoral immunodeficiency.
Asunto(s)
Linfocitos B/inmunología , Enfermedades Genéticas Congénitas/inmunología , Predisposición Genética a la Enfermedad , Síndromes de Inmunodeficiencia/inmunología , Mutación Missense , Factores de Necrosis Tumoral/inmunología , Adulto , Sustitución de Aminoácidos , Factor Activador de Células B/genética , Factor Activador de Células B/inmunología , Linfocitos B/patología , Proliferación Celular , Supervivencia Celular/genética , Supervivencia Celular/inmunología , Niño , Preescolar , Citocina TWEAK , Regulación hacia Abajo/genética , Regulación hacia Abajo/inmunología , Femenino , Enfermedades Genéticas Congénitas/genética , Enfermedades Genéticas Congénitas/patología , Humanos , Síndromes de Inmunodeficiencia/genética , Síndromes de Inmunodeficiencia/patología , Masculino , Subunidad p52 de NF-kappa B/genética , Subunidad p52 de NF-kappa B/inmunología , Factores de Necrosis Tumoral/genéticaRESUMEN
X-linked hyper-IgM syndrome (XHM) is a combined immune deficiency disorder caused by mutations in CD40 ligand. We tested CP-870,893, a human CD40 agonist monoclonal antibody, in the treatment of two XHM patients with biliary Cryptosporidiosis. CP-870,893 activated B cells and APCs in vitro, restoring class switch recombination in XHM B cells and inducing cytokine secretion by monocytes. CP-870,893 infusions were well tolerated and showed significant activity in vivo, decreasing leukocyte concentration in peripheral blood. Although specific antibody responses were lacking, frequent dosing in one subject primed T cells to secrete IFN-g and suppressed oocyst shedding in the stool. Nevertheless, relapse occurred after discontinuation of therapy. The CD40 receptor was rapidly internalized following binding with CP-870,893, potentially explaining the limited capacity of CP-870,893 to mediate immune reconstitution. This study demonstrates that CP-870,893 suppressed oocysts shedding in XHM patients with biliary cryptosporidiosis. The continued study of CD40 agonists in XHM is warranted.
Asunto(s)
Anticuerpos Monoclonales/uso terapéutico , Ligando de CD40/agonistas , Criptosporidiosis/tratamiento farmacológico , Síndrome de Inmunodeficiencia con Hiper-IgM Tipo 1/tratamiento farmacológico , Adolescente , Anticuerpos Monoclonales Humanizados , Ligando de CD40/inmunología , Criptosporidiosis/inmunología , Criptosporidiosis/microbiología , Cryptosporidium/aislamiento & purificación , Cryptosporidium/fisiología , Citocinas/inmunología , Heces/microbiología , Humanos , Síndrome de Inmunodeficiencia con Hiper-IgM Tipo 1/inmunología , Síndrome de Inmunodeficiencia con Hiper-IgM Tipo 1/microbiología , Recuento de Leucocitos , Leucocitos Mononucleares/efectos de los fármacos , Leucocitos Mononucleares/inmunología , Masculino , Linfocitos T/efectos de los fármacos , Linfocitos T/inmunologíaRESUMEN
Ectodermal dysplasia with immune deficiency (EDI) is an immunological and developmental disorder caused by alterations in the gene encoding NF-κB essential modulator (NEMO; also known as IκB kinase γ subunit [IKKγ]). Missense mutations in the gene encoding NEMO are associated with reduced signal-induced nuclear translocation of NF-κB proteins, resulting in defective expression of NF-κB target genes. Here, we report 2 unrelated male patients with EDI, both of whom have normal NEMO coding sequences, but exhibit a marked reduction in expression of full-length NEMO protein. TLR4 stimulation of APCs from these patients induced normal cytoplasmic activation and nuclear translocation of NF-κB. However, cells deficient in full-length NEMO were defective in expression of NF-κB-regulated cytokines, such as IL-12, suggesting a downstream defect in chromatin accessibility for NF-κB transcription factors. TLR4-stimulated APCs from the patients were defective in IKKα-dependent H3 histone phosphorylation at the IL-12 promoter and recruitment of NF-κB heterodimers RelA and cRel to the promoter. Expression of a super-active form of IKKα restored IL-12 production in a NEMO knockdown human monocytic cell line following LPS treatment. Our findings suggest that NEMO regulates the nuclear function of IKKα and offer new insights into the mechanisms underlying diminished NF-κB signaling in patients with EDI.
Asunto(s)
Displasia Ectodérmica/inmunología , Displasia Ectodérmica/metabolismo , Enfermedades Genéticas Ligadas al Cromosoma X/inmunología , Enfermedades Genéticas Ligadas al Cromosoma X/metabolismo , Quinasa I-kappa B/metabolismo , Síndromes de Inmunodeficiencia/inmunología , Síndromes de Inmunodeficiencia/metabolismo , Adolescente , Línea Celular , Núcleo Celular/metabolismo , Preescolar , Citocinas/genética , Citocinas/metabolismo , Proteínas de Unión al ADN/metabolismo , Displasia Ectodérmica/genética , Expresión Génica , Técnicas de Silenciamiento del Gen , Reordenamiento Génico , Enfermedades Genéticas Ligadas al Cromosoma X/genética , Humanos , Quinasa I-kappa B/antagonistas & inhibidores , Quinasa I-kappa B/genética , Síndromes de Inmunodeficiencia/genética , Interleucina-12/genética , Interleucina-12/metabolismo , Sistema de Señalización de MAP Quinasas , Masculino , Monocitos/inmunología , Monocitos/metabolismo , FN-kappa B/metabolismo , Proteínas Nucleares/metabolismo , Enfermedades de Inmunodeficiencia Primaria , Regiones Promotoras Genéticas , Proteínas Proto-Oncogénicas c-rel , Transducción de Señal , Receptor Toll-Like 4/metabolismo , Factor de Transcripción ReIA/metabolismoRESUMEN
CYLD is a lysine 63-deubiquitinating enzyme that inhibits NF-κB and JNK signaling. Here, we show that CYLD knock-out mice have markedly increased numbers of regulatory T cells (Tregs) in peripheral lymphoid organs but not in the thymus. In vitro stimulation of CYLD-deficient naive T cells with anti-CD3/28 in the presence of TGF-ß led to a marked increase in the number of Foxp3-expressing T cells when compared with stimulated naive control CD4(+) cells. Under endogenous conditions, CYLD formed a complex with Smad7 that facilitated CYLD deubiquitination of Smad7 at lysine 360 and 374 residues. Moreover, this site-specific ubiquitination of Smad7 was required for activation of TAK1 and p38 kinases. Finally, knockdown of Smad7 or inhibition of p38 activity in primary T cells impaired Treg differentiation. Together, our results show that CYLD regulates TGF-ß signaling function in T cells and the development of Tregs through deubiquitination of Smad7.
Asunto(s)
Cisteína Endopeptidasas/metabolismo , Transducción de Señal , Proteína smad7/metabolismo , Linfocitos T Reguladores/citología , Factor de Crecimiento Transformador beta/metabolismo , Animales , Linfocitos T CD4-Positivos/citología , Linfocitos T CD4-Positivos/efectos de los fármacos , Linfocitos T CD4-Positivos/metabolismo , Cisteína Endopeptidasas/deficiencia , Cisteína Endopeptidasas/genética , Enzima Desubiquitinante CYLD , Factores de Transcripción Forkhead/genética , Técnicas de Inactivación de Genes , Células HeLa , Humanos , Subunidad alfa del Receptor de Interleucina-2/metabolismo , Ganglios Linfáticos/inmunología , Quinasas Quinasa Quinasa PAM/metabolismo , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Ratones , Regiones Promotoras Genéticas , Unión Proteica , Transducción de Señal/efectos de los fármacos , Linfocitos T Reguladores/efectos de los fármacos , Linfocitos T Reguladores/metabolismo , Factor de Crecimiento Transformador beta/farmacología , Ubiquitinación/efectos de los fármacos , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismoRESUMEN
The covalent attachment of lysine 63-linked polyubiquitin to the zinc-finger domain of IKBKG/NEMO (also known as IKKγ) is necessary for full activation of NF-κB. Impairments of this biochemical mechanism explain the deleterious effects of hypomorphic NEMO mutations on NF-κB signaling function in humans suffering from X-linked ectodermal dysplasia and immunodeficiency. Nevertheless, the biological function of the NEMO zinc-finger domain in the regulation of mitogen-activated protein kinase (MAPK) activity is poorly understood. Here we show that dendritic cells from patients with EDI caused by a C-terminal E391X deletion of the zinc finger of NEMO exhibit impaired MAPK activation in response to lipopolysaccharide (LPS) stimulation. Interestingly, DCs from patients with a C417R missense mutation within the zinc finger domain of NEMO in which ubiquitination of NEMO is preserved are also defective in JNK and ERK activity following LPS stimulation. Our findings indicate that the structural integrity of the NEMO ZF domain is more important than its polyubiquitination for full activation of the MAPK. Furthermore, phosphorylation and polyubiquitination of upstream TAK1 were significantly reduced in the E391X zinc-finger deleted patients, indicating that the NEMO zinc finger may play an important role in assembling the proximal signaling complex for MAPK activation.
Asunto(s)
Células Dendríticas/metabolismo , Displasia Ectodérmica/enzimología , Displasia Ectodérmica/genética , Quinasa I-kappa B/genética , Síndromes de Inmunodeficiencia/enzimología , Síndromes de Inmunodeficiencia/genética , Sistema de Señalización de MAP Quinasas/genética , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Células Cultivadas , Citocinas/biosíntesis , Displasia Ectodérmica/inmunología , Enfermedades Genéticas Ligadas al Cromosoma X , Humanos , Quinasa I-kappa B/química , Quinasa I-kappa B/metabolismo , Síndromes de Inmunodeficiencia/inmunología , Lipopolisacáridos/inmunología , Quinasas Quinasa Quinasa PAM/metabolismo , Proteínas Quinasas Activadas por Mitógenos/genética , Mutación , FN-kappa B/metabolismo , Enfermedades de Inmunodeficiencia Primaria , Eliminación de Secuencia , Ubiquitinación , Dedos de Zinc/genéticaAsunto(s)
Proteínas Serina-Treonina Quinasas/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , Proteínas de la Ataxia Telangiectasia Mutada , Aurora Quinasa B , Aurora Quinasas , Proteínas de Ciclo Celular/metabolismo , Proteínas de Unión al ADN/metabolismo , Humanos , Fosforilación , Estructura Terciaria de Proteína , Proteínas Represoras/metabolismo , Proteínas Supresoras de Tumor/metabolismoRESUMEN
NIR (novel INHAT repressor) is a transcriptional co-repressor with inhibitor of histone acetyltransferase (INHAT) activity and has previously been shown to physically interact with and suppress p53 transcriptional activity and function. However, the mechanism by which NIR suppresses p53 is not completely understood. Using a proteomic approach, we have identified the Aurora kinase B as a novel binding partner of NIR. We show that Aurora B, NIR and p53 exist in a protein complex in which Aurora B binds to NIR, thus also indirectly associates with p53. Functionally, overexpression of Aurora B or NIR suppresses p53 transcriptional activity, and depletion of Aurora B or NIR causes p53-dependent apoptosis and cell growth arrest, due to the up-regulation of p21 and Bax. We then demonstrate that Aurora B phosphorylates multiple sites in the p53 DNA-binding domain in vitro, and this phosphorylation probably also occurs in cells. Importantly, the Aurora B-mediated phosphorylation on Ser(269) or Thr(284) significantly compromises p53 transcriptional activity. Taken together, these results provide novel insight into NIR-mediated p53 suppression and also suggest an additional way for p53 regulation.