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1.
Gastroenterology ; 2019 Nov 16.
Artigo em Inglês | MEDLINE | ID: mdl-31743734

RESUMO

BACKGROUND & AIMS: Mutations in the tetratricopeptide repeat domain 7A gene (TTC7A) cause intestinal epithelial and immune defects. Patients can become immune deficient and develop apoptotic enterocolitis, multiple intestinal atresia, and recurrent intestinal stenosis. The intestinal disease in patients with TTC7A deficiency is severe, untreatable, and recurs despite resection or allogeneic hematopoietic stem cell transplant. We screened drugs for those that prevent apoptosis of in cells with TTC7A deficiency and tested their effects in an animal model of the disease. METHODS: We developed a high-throughput screen to identify compounds approved by the Food and Drug Administration that reduce activity of caspases 3 and 7 in TTC7A-knockout HAP1 (human haploid) cells and reduce the susceptibility to apoptosis. We validated the effects of identified agents in HeLa cells that stably express TTC7A with point mutations found in patients. Signaling pathways in cells were analyzed by immunoblots. We tested the effects of identified agents in zebrafish with disruption of ttc7a, which develop intestinal defects, and colonoids derived from biopsies of patients with and without mutations in TTC7A. We performed real-time imaging of intestinal peristalsis in zebrafish and histologic analyses of intestinal tissues from patients and zebrafish. Colonoids were analyzed by immunofluorescence and for ion transport. RESULTS: TTC7A-knockout HAP1 cells have abnormal morphology and undergo apoptosis, due to increased levels of active caspases 3 and 7. We identified drugs that increased cell viability; leflunomide (used to treat patients with inflammatory conditions) reduced caspase 3 and 7 activity in cells by 96%. TTC7A-knockout cells contained cleaved caspase 3 and had reduced levels of phosphorylated AKT and XIAP; incubation of these cells with leflunomide increased levels of phosphorylated AKT and XIAP and reduced levels of cleaved caspase 3. Administration of leflunomide to ttc7a-/- zebrafish increased gut motility, reduced intestinal tract narrowing, increased intestinal cell survival, increased sizes of intestinal luminal spaces, and restored villi and goblet cell morphology. Exposure of patient-derived colonoids to leflunomide increased cell survival, polarity, and transport function. CONCLUSIONS: In a drug screen, we identified leflunomide as an agent that reduces apoptosis and levels of caspase 3 and activates AKT signaling and in TTC7A-knockout cells. In zebrafish with disruption of ttc7a, leflunomide restores gut motility, reduces intestinal tract narrowing, and increases intestinal cell survival. This drug might be repurposed for treatment of TTC7A deficiency.

2.
Science ; 366(6464): 460-467, 2019 10 25.
Artigo em Inglês | MEDLINE | ID: mdl-31649195

RESUMO

The nucleotide oligomerization domain (NOD)-like receptors 1 and 2 (NOD1/2) are intracellular pattern-recognition proteins that activate immune signaling pathways in response to peptidoglycans associated with microorganisms. Recruitment to bacteria-containing endosomes and other intracellular membranes is required for NOD1/2 signaling, and NOD1/2 mutations that disrupt membrane localization are associated with inflammatory bowel disease and other inflammatory conditions. However, little is known about this recruitment process. We found that NOD1/2 S-palmitoylation is required for membrane recruitment and immune signaling. ZDHHC5 was identified as the palmitoyltransferase responsible for this critical posttranslational modification, and several disease-associated mutations in NOD2 were found to be associated with defective S-palmitoylation. Thus, ZDHHC5-mediated S-palmitoylation of NOD1/2 is critical for their ability to respond to peptidoglycans and to mount an effective immune response.

4.
Proc Natl Acad Sci U S A ; 116(3): 970-975, 2019 01 15.
Artigo em Inglês | MEDLINE | ID: mdl-30591564

RESUMO

Receptor-interacting serine/threonine-protein kinase 1 (RIPK1) is a critical regulator of cell death and inflammation, but its relevance for human disease pathogenesis remains elusive. Studies of monogenic disorders might provide critical insights into disease mechanisms and therapeutic targeting of RIPK1 for common diseases. Here, we report on eight patients from six unrelated pedigrees with biallelic loss-of-function mutations in RIPK1 presenting with primary immunodeficiency and/or intestinal inflammation. Mutations in RIPK1 were associated with reduced NF-κB activity, defective differentiation of T and B cells, increased inflammasome activity, and impaired response to TNFR1-mediated cell death in intestinal epithelial cells. The characterization of RIPK1-deficient patients highlights the essential role of RIPK1 in controlling human immune and intestinal homeostasis, and might have critical implications for therapies targeting RIPK1.


Assuntos
Diferenciação Celular , Imunidade nas Mucosas/genética , Doenças Inflamatórias Intestinais , Mucosa Intestinal , Proteína Serina-Treonina Quinases de Interação com Receptores , Imunodeficiência Combinada Severa , Linfócitos B/imunologia , Linfócitos B/patologia , Diferenciação Celular/genética , Diferenciação Celular/imunologia , Células Epiteliais/imunologia , Células Epiteliais/patologia , Feminino , Células HCT116 , Células HEK293 , Humanos , Doenças Inflamatórias Intestinais/genética , Doenças Inflamatórias Intestinais/imunologia , Doenças Inflamatórias Intestinais/patologia , Mucosa Intestinal/imunologia , Mucosa Intestinal/patologia , Masculino , Mutação , NF-kappa B/genética , NF-kappa B/imunologia , Proteína Serina-Treonina Quinases de Interação com Receptores/deficiência , Proteína Serina-Treonina Quinases de Interação com Receptores/imunologia , Imunodeficiência Combinada Severa/genética , Imunodeficiência Combinada Severa/imunologia , Imunodeficiência Combinada Severa/patologia , Linfócitos T/imunologia , Linfócitos T/patologia
5.
Gastroenterology ; 2018 Sep 26.
Artigo em Inglês | MEDLINE | ID: mdl-30267714

RESUMO

Caspase-8 (CASP8) is a protease that initiates apoptosis and regulates inflammation and immune responses. We identified germline mutations in CASP8 in 3 unrelated patients with infant-onset inflammatory bowel disease: 2 patients were homozygous for the mutation 710A>G, p.Q237R, which resulted in reduced protein expression, and 1 patient carried the mutation 793C>T, p.R265W. We isolated peripheral blood mononuclear cells from our index patient and observed defects in T- and B-cell maturation, proliferation, and/or activation. Macrophages from 1 patient with CASP8 deficiency and monocytic BLaER1 cells with knockout of CASP8 or overexpression of CASP8 with the 710A>G mutation had altered inflammasome activity on stimulation with lipopolysaccharide. Patient-derived intestinal organoids and colon carcinoma cells with knockout of CASP8 had defects in cell death processes that involved loss of TRAIL signaling and increased necroptosis. These findings indicate that CASP8 controls inflammation, innate and adaptive immunity, and intestinal barrier integrity in humans.

6.
J Clin Invest ; 128(9): 3957-3975, 2018 Aug 31.
Artigo em Inglês | MEDLINE | ID: mdl-29969437

RESUMO

Biallelic loss-of-function (LOF) mutations of the NCF4 gene, encoding the p40phox subunit of the phagocyte NADPH oxidase, have been described in only 1 patient. We report on 24 p40phox-deficient patients from 12 additional families in 8 countries. These patients display 8 different in-frame or out-of-frame mutations of NCF4 that are homozygous in 11 of the families and compound heterozygous in another. When overexpressed in NB4 neutrophil-like cells and EBV-transformed B cells in vitro, the mutant alleles were found to be LOF, with the exception of the p.R58C and c.120_134del alleles, which were hypomorphic. Particle-induced NADPH oxidase activity was severely impaired in the patients' neutrophils, whereas PMA-induced dihydrorhodamine-1,2,3 (DHR) oxidation, which is widely used as a diagnostic test for chronic granulomatous disease (CGD), was normal or mildly impaired in the patients. Moreover, the NADPH oxidase activity of EBV-transformed B cells was also severely impaired, whereas that of mononuclear phagocytes was normal. Finally, the killing of Candida albicans and Aspergillus fumigatus hyphae by neutrophils was conserved in these patients, unlike in patients with CGD. The patients suffer from hyperinflammation and peripheral infections, but they do not have any of the invasive bacterial or fungal infections seen in CGD. Inherited p40phox deficiency underlies a distinctive condition, resembling a mild, atypical form of CGD.

7.
EMBO Mol Med ; 10(4)2018 04.
Artigo em Inglês | MEDLINE | ID: mdl-29567797

RESUMO

Herein, we report the first identification of biallelic-inherited mutations in ALPI as a Mendelian cause of inflammatory bowel disease in two unrelated patients. ALPI encodes for intestinal phosphatase alkaline, a brush border metalloenzyme that hydrolyses phosphate from the lipid A moiety of lipopolysaccharides and thereby drastically reduces Toll-like receptor 4 agonist activity. Prediction tools and structural modelling indicate that all mutations affect critical residues or inter-subunit interactions, and heterologous expression in HEK293T cells demonstrated that all ALPI mutations were loss of function. ALPI mutations impaired either stability or catalytic activity of ALPI and rendered it unable to detoxify lipopolysaccharide-dependent signalling. Furthermore, ALPI expression was reduced in patients' biopsies, and ALPI activity was undetectable in ALPI-deficient patient's stool. Our findings highlight the crucial role of ALPI in regulating host-microbiota interactions and restraining host inflammatory responses. These results indicate that ALPI mutations should be included in screening for monogenic causes of inflammatory bowel diseases and lay the groundwork for ALPI-based treatments in intestinal inflammatory disorders.

8.
Nat Commun ; 8: 14816, 2017 04 03.
Artigo em Inglês | MEDLINE | ID: mdl-28368018

RESUMO

Human actin-related protein 2/3 complex (Arp2/3), required for actin filament branching, has two ARPC1 component isoforms, with ARPC1B prominently expressed in blood cells. Here we show in a child with microthrombocytopenia, eosinophilia and inflammatory disease, a homozygous frameshift mutation in ARPC1B (p.Val91Trpfs*30). Platelet lysates reveal no ARPC1B protein and greatly reduced Arp2/3 complex. Missense ARPC1B mutations are identified in an unrelated patient with similar symptoms and ARPC1B deficiency. ARPC1B-deficient platelets are microthrombocytes similar to those seen in Wiskott-Aldrich syndrome that show aberrant spreading consistent with loss of Arp2/3 function. Knockout of ARPC1B in megakaryocytic cells results in decreased proplatelet formation, and as observed in platelets from patients, increased ARPC1A expression. Thus loss of ARPC1B produces a unique set of platelet abnormalities, and is associated with haematopoietic/immune symptoms affecting cell lineages where this isoform predominates. In agreement with recent experimental studies, our findings suggest that ARPC1 isoforms are not functionally interchangeable.


Assuntos
Complexo 2-3 de Proteínas Relacionadas à Actina/deficiência , Transtornos Plaquetários/metabolismo , Plaquetas/metabolismo , Inflamação/patologia , Complexo 2-3 de Proteínas Relacionadas à Actina/metabolismo , Actinas/metabolismo , Plaquetas/efeitos dos fármacos , Plaquetas/patologia , Plaquetas/ultraestrutura , Células da Medula Óssea/efeitos dos fármacos , Células da Medula Óssea/metabolismo , Células da Medula Óssea/patologia , Forma Celular , Suscetibilidade a Doenças , Fibrinogênio/farmacologia , Técnicas de Inativação de Genes , Humanos , Megacariócitos/efeitos dos fármacos , Megacariócitos/metabolismo , Megacariócitos/patologia , Mutação/genética , Vasculite/patologia , Síndrome de Wiskott-Aldrich/patologia
9.
Hum Mol Genet ; 24(23): 6614-23, 2015 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-26358773

RESUMO

Congenital sodium diarrhea (CSD) refers to an intractable diarrhea of intrauterine onset with high fecal sodium loss. CSD is clinically and genetically heterogeneous. Syndromic CSD is caused by SPINT2 mutations. While we recently described four cases of the non-syndromic form of CSD that were caused by dominant activating mutations in intestinal receptor guanylate cyclase C (GC-C), the genetic cause for the majority of CSD is still unknown. Therefore, we aimed to determine the genetic cause for non-GC-C non-syndromic CSD in 18 patients from 16 unrelated families applying whole-exome sequencing and/or chromosomal microarray analyses and/or direct Sanger sequencing. SLC9A3 missense, splicing and truncation mutations, including an instance of uniparental disomy, and whole-gene deletion were identified in nine patients from eight families with CSD. Two of these nine patients developed inflammatory bowel disease (IBD) at 4 and 16 years of age. SLC9A3 encodes Na(+)/H(+) antiporter 3 (NHE3), which is the major intestinal brush-border Na(+)/H(+) exchanger. All mutations were in the NHE3 N-terminal transport domain, and all missense mutations were in the putative membrane-spanning domains. Identified SLC9A3 missense mutations were functionally characterized in plasma membrane NHE null fibroblasts. SLC9A3 missense mutations compromised NHE3 activity by reducing basal surface expression and/or loss of basal transport function of NHE3 molecules, whereas acute regulation was normal. This study identifies recessive mutations in NHE3, a downstream target of GC-C, as a cause of CSD and implies primary basal NHE3 malfunction as a predisposition for IBD in a subset of patients.


Assuntos
Anormalidades Múltiplas/genética , Diarreia/congênito , Erros Inatos do Metabolismo/genética , Mutação , Trocadores de Sódio-Hidrogênio/genética , Anormalidades Múltiplas/metabolismo , Anormalidades Múltiplas/fisiopatologia , Adolescente , Adulto , Criança , Pré-Escolar , Análise Mutacional de DNA , Diarreia/genética , Diarreia/metabolismo , Diarreia/fisiopatologia , Feminino , Genes Recessivos , Humanos , Lactente , Recém-Nascido , Doenças Inflamatórias Intestinais/genética , Doenças Inflamatórias Intestinais/metabolismo , Doenças Inflamatórias Intestinais/fisiopatologia , Mucosa Intestinal/metabolismo , Intestinos/fisiopatologia , Masculino , Erros Inatos do Metabolismo/metabolismo , Erros Inatos do Metabolismo/fisiopatologia , Microvilosidades/metabolismo , Análise de Sequência com Séries de Oligonucleotídeos , Trocador 3 de Sódio-Hidrogênio , Adulto Jovem
10.
Immunity ; 41(6): 898-908, 2014 Dec 18.
Artigo em Inglês | MEDLINE | ID: mdl-25526305

RESUMO

The nucleotide-binding oligomerization domain (NOD) proteins NOD1 and NOD2, the founding members of the intracellular NOD-like receptor family, sense conserved motifs in bacterial peptidoglycan and induce proinflammatory and antimicrobial responses. Here, we discuss recent developments about the mechanisms by which NOD1 and NOD2 are activated by bacterial ligands, the regulation of their signaling pathways, and their role in host defense and inflammatory disease. Several routes for the entry of peptidoglycan ligands to the host cytosol to trigger activation of NOD1 and NOD2 have been elucidated. Furthermore, genetic screens and biochemical analyses have revealed mechanisms that regulate NOD1 and NOD2 signaling. Finally, recent studies have suggested several mechanisms to account for the link between NOD2 variants and susceptibility to Crohn's disease. Further understanding of NOD1 and NOD2 should provide new insight into the pathogenesis of disease and the development of new strategies to treat inflammatory and infectious disorders.


Assuntos
Infecções Bacterianas/imunologia , Inflamação/imunologia , Proteína Adaptadora de Sinalização NOD1/imunologia , Proteína Adaptadora de Sinalização NOD2/imunologia , Receptores de Reconhecimento de Padrão/imunologia , Animais , Antígenos de Bactérias/imunologia , Interações Hospedeiro-Patógeno , Humanos , Imunidade Inata , Peptidoglicano/imunologia , Transdução de Sinais
11.
J Biol Chem ; 289(41): 28213-24, 2014 Oct 10.
Artigo em Inglês | MEDLINE | ID: mdl-25170077

RESUMO

NOD2 encodes an intracellular multidomain pattern recognition receptor that is the strongest known genetic risk factor in the pathogenesis of Crohn disease (CD), a chronic relapsing inflammatory disorder of the intestinal tract. NOD2 functions as a sensor for bacterial cell wall components and activates proinflammatory and antimicrobial signaling pathways. Here, using a genome-wide small interfering RNA (siRNA) screen, we identify numerous genes that regulate secretion of the proinflammatory cytokine IL-8 in response to NOD2 activation. Moreover, many of the identified IL-8 regulators are linked by protein-protein interactions, revealing subnetworks of highly connected IL-8 regulators implicated in processes such as vesicle formation, mRNA stability, and protein ubiquitination and trafficking. A TNFα counterscreen to induce IL-8 secretion in an NOD2-independent manner reveals that the majority of the identified regulators affect IL-8 secretion irrespective of the initiating stimuli. Using immortalized macrophages, we validate the ubiquitin protease, USP8, and the endosomal sorting protein, VPS28, as negative regulators of NOD2-induced cytokine secretion. Interestingly, several genes that affect NOD2-induced IL-8 secretion are present in loci associated with CD risk by genome-wide association studies, supporting a role for the NOD2/IL-8 pathway, and not just NOD2, in the pathogenesis of CD. Overall, this screen provides a valuable resource in the advancement of our understanding of the genes that regulate the secretion of IL-8.


Assuntos
Doença de Crohn/genética , Endopeptidases/genética , Complexos Endossomais de Distribuição Requeridos para Transporte/genética , Interleucina-8/genética , Macrófagos/metabolismo , Proteína Adaptadora de Sinalização NOD2/genética , RNA Interferente Pequeno/genética , Ubiquitina Tiolesterase/genética , Animais , Células da Medula Óssea/metabolismo , Células da Medula Óssea/patologia , Linhagem Celular Transformada , Doença de Crohn/metabolismo , Doença de Crohn/patologia , Endopeptidases/metabolismo , Complexos Endossomais de Distribuição Requeridos para Transporte/metabolismo , Loci Gênicos , Estudo de Associação Genômica Ampla , Células HEK293 , Ensaios de Triagem em Larga Escala , Humanos , Interleucina-8/agonistas , Interleucina-8/antagonistas & inibidores , Interleucina-8/metabolismo , Macrófagos/patologia , Camundongos , NF-kappa B/genética , NF-kappa B/metabolismo , Proteína Adaptadora de Sinalização NOD2/antagonistas & inibidores , Proteína Adaptadora de Sinalização NOD2/metabolismo , Mapeamento de Interação de Proteínas , Transporte Proteico , Estabilidade de RNA , RNA Interferente Pequeno/metabolismo , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/metabolismo , Ubiquitina Tiolesterase/metabolismo , Ubiquitinação
12.
Sci Signal ; 6(258): rs3, 2013 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-23322906

RESUMO

The cytoplasmic receptor NOD2 (nucleotide-binding oligomerization domain 2) senses peptidoglycan fragments and triggers host defense pathways, including activation of nuclear factor κB (NF-κB) signaling, which lead to inflammatory immune responses. Dysregulation of NOD2 signaling is associated with inflammatory diseases, such as Crohn's disease and Blau syndrome. We used a genome-wide small interfering RNA screen to identify regulators of the NOD2 signaling pathway. Several genes associated with Crohn's disease risk were identified in the screen. A comparison of candidates from this screen with other "omics" data sets revealed interconnected networks of genes implicated in NF-κB signaling, thus supporting a role for NOD2 and NF-κB pathways in the pathogenesis of Crohn's disease. Many of these regulators were validated in secondary assays, such as measurement of interleukin-8 secretion, which is partially dependent on NF-κB. Knockdown of putative regulators in human embryonic kidney 293 cells followed by stimulation with tumor necrosis factor-α revealed that most of the genes identified were general regulators of NF-κB signaling. Overall, the genes identified here provide a resource to facilitate the elucidation of the molecular mechanisms that regulate NOD2- and NF-κB-mediated inflammation.


Assuntos
Genoma Humano/genética , NF-kappa B/genética , Proteína Adaptadora de Sinalização NOD2/genética , RNA Interferente Pequeno/genética , Transdução de Sinais/genética , Acetilmuramil-Alanil-Isoglutamina/farmacologia , Animais , Western Blotting , Células da Medula Óssea/citologia , Células da Medula Óssea/efeitos dos fármacos , Células da Medula Óssea/metabolismo , Sobrevivência Celular/genética , Células Cultivadas , Doença de Crohn/genética , Regulação da Expressão Gênica/efeitos dos fármacos , Redes Reguladoras de Genes/efeitos dos fármacos , Células HEK293 , Humanos , Proteínas I-kappa B/metabolismo , Macrófagos/citologia , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Camundongos , Camundongos Knockout , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Modelos Genéticos , NADPH Oxidases/deficiência , NADPH Oxidases/genética , NF-kappa B/metabolismo , Proteína Adaptadora de Sinalização NOD2/metabolismo , Interferência de RNA , Transdução de Sinais/efeitos dos fármacos , Fator de Necrose Tumoral alfa/farmacologia
14.
J Immunol ; 187(6): 2849-52, 2011 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-21849681

RESUMO

Nucleotide-binding oligomerization domain 2 (Nod2) mutations including L1007fsinsC are associated with the development of Crohn's disease (CD). These CD-associated Nod2 mutations are common in healthy white populations, suggesting that they may confer some protective function, but experimental evidence is lacking. Using a mouse strain that expresses Nod2(2939iCstop), the equivalent of the L1007fsinsC mutation, we found that macrophages homozygous for Nod2(2939iCstop) are impaired in the recognition of muramyl dipeptide and Enterococcus faecalis, a commensal bacterium that is a common cause of sepsis-associated lethality in humans. Notably, Nod2 deficiency and homozygocity for Nod2(2939iCstop) were associated with reduced production of TNF-α and IL-6 and lethality after systemic infection with E. faecalis despite normal bacteria loads. Consistently, inhibition of TNF-α signaling protected wild-type mice from E. faecalis-induced lethality. These results suggest that the same Nod2 mutation can increase the susceptibility to CD, but also protect the host from systemic infection by a common enteric bacterium.


Assuntos
Doença de Crohn/genética , Enterococcus faecalis/imunologia , Infecções por Bactérias Gram-Positivas/genética , Macrófagos/imunologia , Mutação , Proteína Adaptadora de Sinalização NOD2/genética , Animais , Doença de Crohn/imunologia , Citocinas/biossíntese , Citocinas/imunologia , Técnicas de Introdução de Genes , Infecções por Bactérias Gram-Positivas/imunologia , Immunoblotting , Inflamação/genética , Inflamação/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Mutantes , Proteína Adaptadora de Sinalização NOD2/imunologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa
15.
Immunity ; 34(5): 769-80, 2011 May 27.
Artigo em Inglês | MEDLINE | ID: mdl-21565531

RESUMO

The intracellular sensor Nod2 is activated in response to bacteria, and the impairment of this response is linked to Crohn's disease. However, the function of Nod2 in host defense remains poorly understood. We found that Nod2-/- mice exhibited impaired intestinal clearance of Citrobacter rodentium, an enteric bacterium that models human infection by pathogenic Escherichia coli. The increased bacterial burden was preceded by reduced CCL2 chemokine production, inflammatory monocyte recruitment, and Th1 cell responses in the intestine. Colonic stromal cells, but not epithelial cells or resident CD11b+ phagocytic cells, produced CCL2 in response to C. rodentium in a Nod2-dependent manner. Unlike resident phagocytic cells, inflammatory monocytes produced IL-12, a cytokine that induces adaptive immunity required for pathogen clearance. Adoptive transfer of Ly6C(hi) monocytes restored the clearance of the pathogen in infected Ccr2-/- mice. Thus, Nod2 mediates CCL2-CCR2-dependent recruitment of inflammatory monocytes, which is important in promoting bacterial eradication in the intestine.


Assuntos
Quimiocina CCL2/imunologia , Citrobacter rodentium/imunologia , Colite/imunologia , Infecções por Enterobacteriaceae/imunologia , Monócitos/imunologia , Proteína Adaptadora de Sinalização NOD2/imunologia , Animais , Colite/microbiologia , Colite/patologia , Modelos Animais de Doenças , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteína Adaptadora de Sinalização NOD2/biossíntese , Proteína Adaptadora de Sinalização NOD2/deficiência , Células Estromais/imunologia
16.
Trends Immunol ; 32(2): 73-9, 2011 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-21251876

RESUMO

The identification of several families of innate pattern recognition receptors has greatly enhanced our understanding of the host innate immune response against a variety of pathogens. One such family of innate receptors is the nucleotide-binding domain and leucine rich repeat containing receptors (NLRs). NOD2 has been characterized as a cytosolic sensor of bacteria peptidoglycan (PGN). For almost 10 years, NOD2 was assigned with the function of mediating the RICK- and nuclear factor-κB induced proinflammatory response triggered by PGN. Recent studies have extended the biological activity of NOD2 to include the induction of autophagy and antiviral responses, as well as mediating direct T cell activation. Here, we highlight and discuss these new findings in the context of immune activation and pathogen detection.


Assuntos
Autofagia , Ativação Linfocitária , Proteína Adaptadora de Sinalização NOD2/imunologia , Linfócitos T/imunologia , Linfócitos T/virologia , Imunidade Adaptativa , Animais , Humanos , Proteína Adaptadora de Sinalização NOD2/genética
17.
Nat Immunol ; 10(12): 1267-74, 2009 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-19881508

RESUMO

Nod2 belongs to the nucleotide-binding oligomerization domain receptor (NLR) family of proteins, which function as intracellular pathogen sensors in innate immune cells. Nod2 deficiency results in an impaired immune response to bacterial pathogens. However, how this protein promotes host defense against intracellular parasites is unknown. Here we found that Nod2(-/-) mice had less clearance of Toxoplasma gondii and lower interferon-gamma (IFN-gamma) production. Reconstitution of T cell-deficient mice with Nod2(-/-) T cells followed by T. gondii infection demonstrated a T cell-intrinsic defect. Nod2(-/-) CD4(+) T cells had poor helper T cell differentiation, which was associated with impaired production of interleukin 2 (IL-2) and nuclear accumulation of the transcription factor subunit c-Rel. Our data demonstrate a T cell-intrinsic role for Nod2 signaling that is critical for host defense against T. gondii.


Assuntos
Proteína Adaptadora de Sinalização NOD2/imunologia , Linfócitos T/imunologia , Toxoplasma/imunologia , Toxoplasmose/imunologia , Animais , Diferenciação Celular , Colite/imunologia , Colite/patologia , Interferon gama/biossíntese , Interferon gama/imunologia , Interleucina-2/biossíntese , Interleucina-2/imunologia , Camundongos , Camundongos Knockout , Proteína Adaptadora de Sinalização NOD2/deficiência , Transdução de Sinais , Taxa de Sobrevida , Linfócitos T/citologia , Toxoplasmose/metabolismo
18.
Immunol Rev ; 227(1): 106-28, 2009 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-19120480

RESUMO

Nucleotide oligomerization domain (NOD)-like receptors (NLRs) are a specialized group of intracellular proteins that play a critical role in the regulation of the host innate immune response. NLRs act as scaffolding proteins that assemble signaling platforms that trigger nuclear factor-kappaB and mitogen-activated protein kinase signaling pathways and control the activation of inflammatory caspases. Importantly, mutations in several members of the NLR family have been linked to a variety of inflammatory diseases consistent with these molecules playing an important role in host-pathogen interactions and the inflammatory response. In this review, we focus on the role of Nod1 and Nod2 in host defense and in particular discuss recent finding regarding the role of Nlrc4, Nlpr1, and Nlrp3 inflammasomes in caspase-1 activation and subsequent release of proinflammatory cytokines such as interleukin-1 beta.


Assuntos
Infecções Bacterianas/imunologia , Caspase 1/metabolismo , Interações Hospedeiro-Patógeno/imunologia , Proteínas Adaptadoras de Sinalização NOD/imunologia , Proteínas Adaptadoras de Sinalização NOD/metabolismo , Fagocitose/imunologia , Animais , Apoptose/imunologia , Doenças Autoimunes/genética , Doenças Autoimunes/imunologia , Infecções Bacterianas/prevenção & controle , Caspase 1/imunologia , Predisposição Genética para Doença , Humanos , Imunidade Inata , Interleucina-1beta/metabolismo , Camundongos , Mutação , Proteínas Adaptadoras de Sinalização NOD/genética , Transdução de Sinais/imunologia
19.
FEBS J ; 275(10): 2561-73, 2008 May.
Artigo em Inglês | MEDLINE | ID: mdl-18422655

RESUMO

Eph receptor tyrosine kinases regulate many important biological processes. In the present study, we explored the substrate specificity of the EphA4 receptor tyrosine kinase using peptide arrays. We define a consensus substrate motif for EphA4 and go on to identify and test a number of potential EphA4 substrates and map their putative site(s) of phosphorylation. Cotransfection studies validate two of the predicted substrates: Nck2 and Dok1. Our findings identify several potential EphA4 substrates and demonstrate the general utility of using peptide arrays to rapidly identify and map protein kinase phosphorylation sites.


Assuntos
Peptídeos/metabolismo , Análise Serial de Proteínas , Receptor EphA4/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Motivos de Aminoácidos , Animais , Células Cultivadas , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Humanos , Camundongos , Dados de Sequência Molecular , Proteínas Oncogênicas/genética , Proteínas Oncogênicas/metabolismo , Peptídeos/genética , Fosfoproteínas/genética , Fosfoproteínas/metabolismo , Fosforilação , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Receptor EphA4/genética , Reprodutibilidade dos Testes , Especificidade por Substrato , Tirosina/metabolismo
20.
Proc Natl Acad Sci U S A ; 104(52): 20973-8, 2007 Dec 26.
Artigo em Inglês | MEDLINE | ID: mdl-18093944

RESUMO

The intracellular signaling targets used by mammalian axon guidance receptors to organize the nervous system in vivo are unclear. The Nck1 and Nck2 SH2/SH3 adaptors (collectively Nck) can couple phosphotyrosine (pTyr) signals to reorganization of the actin cytoskeleton and are therefore candidates for linking guidance cues to the regulatory machinery of the cytoskeleton. We find that selective inactivation of Nck in the murine nervous system causes a hopping gait and a defect in the spinal central pattern generator, which is characterized by synchronous firing of bilateral ventral motor neurons. Nck-deficient mice also show abnormal projections of corticospinal tract axons and defective development of the posterior tract of the anterior commissure. These phenotypes are consistent with a role for Nck in signaling initiated by different classes of guidance receptors, including the EphA4 receptor tyrosine kinase. Our data indicate that Nck adaptors couple pTyr guidance signals to cytoskeletal events required for the ipsilateral projections of spinal cord neurons and thus for normal limb movement.


Assuntos
Proteínas Oncogênicas/fisiologia , Caminhada , Actinas/química , Proteínas Adaptadoras de Transdução de Sinal , Animais , Axônios/metabolismo , Quimerina 1/metabolismo , Citoesqueleto/metabolismo , Fibroblastos/metabolismo , Locomoção , Camundongos , Modelos Biológicos , Neurônios Motores/metabolismo , Proteínas Oncogênicas/metabolismo , Fenótipo , Receptor EphA4/química , Transdução de Sinais , Medula Espinal/metabolismo , Domínios de Homologia de src
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