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BACKGROUND AND OBJECTIVES: To describe the clinical features of anti-NMDAR encephalitis, emphasizing on late-onset patients and antibody test characteristics in serum and CSF. METHODS: Nationwide observational Dutch cohort study, in patients diagnosed with anti-NMDAR encephalitis between 2007 and 2019. RESULTS: One hundred twenty-six patients with anti-NMDAR encephalitis were included with a median age of 24 years (range 1-86 years). The mean annual incidence was 1.00/million (95% CI 0.62-1.59). Patients ≥45 years of age at onset (19%) had fewer seizures (46% vs 71%, p = 0.021), fewer symptoms during disease course (3 vs 6 symptoms, p = 0.020), and more often undetectable serum antibodies compared with younger patients (p = 0.031). In the late-onset group, outcome was worse, and all tumors were carcinomas (both p < 0.0001). CSF was more accurate than serum to detect anti-NMDAR encephalitis (sensitivity 99% vs 68%, p < 0.0001). Using cell-based assay (CBA), CSF provided an unconfirmed positive test result in 11/2,600 patients (0.4%); 6/11 had a neuroinflammatory disease (other than anti-NMDAR encephalitis). Patients with anti-NMDAR encephalitis, who tested positive in CSF only, had lower CSF antibody titers (p = 0.003), but appeared to have an equally severe disease course. DISCUSSION: Anti-NMDAR encephalitis occurs at all ages and is less rare in the elderly patients than initially anticipated. In older patients, the clinical phenotype is less outspoken, has different tumor association, and a less favorable recovery. Detection of antibodies in CSF is the gold standard, and although the CBA has very good validity, it is not perfect. The clinical phenotype should be leading, and confirmation in a research laboratory is recommended, when in doubt.
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Encefalite Antirreceptor de N-Metil-D-Aspartato/diagnóstico , Autoanticorpos , Neoplasias , Adolescente , Adulto , Idade de Início , Idoso , Idoso de 80 Anos ou mais , Encefalite Antirreceptor de N-Metil-D-Aspartato/sangue , Encefalite Antirreceptor de N-Metil-D-Aspartato/líquido cefalorraquidiano , Encefalite Antirreceptor de N-Metil-D-Aspartato/epidemiologia , Autoanticorpos/sangue , Autoanticorpos/líquido cefalorraquidiano , Criança , Pré-Escolar , Estudos de Coortes , Feminino , Humanos , Lactente , Masculino , Pessoa de Meia-Idade , Neoplasias/epidemiologia , Países Baixos/epidemiologia , Adulto JovemRESUMO
In this study we report the clinical features of 32 patients with gamma aminobutyric acid B receptor (GABABR) antibodies, identify additional autoantibodies in patients with anti-GABABR encephalitis that mark the presence of an underlying small cell lung carcinoma and optimize laboratory methods for the detection of GABABR antibodies. Patients (n = 3225) were tested for the presence of GABABR antibodies using cell-based assay, immunohistochemistry and live hippocampal neurons. Clinical data were obtained retrospectively. Potassium channel tetramerization domain-containing (KCTD)16 antibodies were identified by immunoprecipitation, mass spectrometry analysis and cell-based assays. KCTD16 antibodies were identified in 23/32 patients with anti-GABABR encephalitis, and in 1/26 patients with small cell lung carcinoma and Hu antibodies, but not in 329 healthy subjects and disease controls. Of the anti-GABABR encephalitis patients that were screened sufficiently, 18/19 (95%) patients with KCTD16 antibodies had a tumour versus 3/9 (33%) anti-GABABR encephalitis patients without KCTD16 antibodies (P = 0.001). In most cases this was a small cell lung carcinoma. Patients had cognitive or behavioural changes (97%) and prominent seizures (90%). Thirteen patients developed a refractory status epilepticus with intensive care unit admittance (42%). Strikingly, 4/32 patients had a rapidly progressive dementia. The addition of KCTD16 to the GABABR cell-based assay improved sensitivity of the in-house fixed cell-based assay, without loss of specificity. Twenty-two of 26 patients improved (partially) to immunotherapy or chemotherapy. Anti-GABABR encephalitis is a limbic encephalitis with prominent, severe seizures, but patients can also present with rapidly progressive dementia. The co-occurrence of KCTD16 antibodies points towards a paraneoplastic origin. The addition of KCTD16 improves the sensitivity of the cell-based assay.
Assuntos
Autoanticorpos/imunologia , Encefalite/diagnóstico , Encefalite/genética , Ácido gama-Aminobutírico/imunologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Fatores Imunológicos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/imunologia , Masculino , Pessoa de Meia-Idade , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/imunologia , Neurônios/patologia , Convulsões/diagnóstico , Convulsões/genética , Estado Epiléptico/genética , Estado Epiléptico/imunologia , Ácido gama-Aminobutírico/genéticaRESUMO
Paraneoplastic neurological syndromes (PNS) are often characterized by the presence of antineuronal antibodies in patient serum or cerebrospinal fluid. The detection of antineuronal antibodies has proven to be a useful tool in PNS diagnosis and the search for an underlying tumor. Here, we describe three patients with autoantibodies to several epitopes of the axon initial segment protein tripartite motif 46 (TRIM46). We show that anti-TRIM46 antibodies are easy to detect in routine immunohistochemistry screening and can be confirmed by western blotting and cell-based assay. Anti-TRIM46 antibodies can occur in patients with diverse neurological syndromes and are associated with small-cell lung carcinoma.
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Autoimmune encephalitis (AIE) is a group of disorders in which autoantibodies directed at antigens located on the plasma membrane of neurons induce severe neurological symptoms. In contrast to classical paraneoplastic disorders, AIE patients respond well to immunotherapy. The detection of neuronal surface autoantibodies in patients' serum or CSF therefore has serious consequences for the patients' treatment and follow-up and requires the availability of sensitive and specific diagnostic tests. This mini-review provides a guideline for both diagnostic and research laboratories that work on the detection of known surface autoantibodies and/or the identification of novel surface antigens. We discuss the strengths and pitfalls of different techniques for anti-neuronal antibody detection: (1) Immunohistochemistry (IHC) and immunofluorescence on rat/primate brain sections; (2) Immunocytochemistry (ICC) of living cultured hippocampal neurons; and (3) Cell Based Assay (CBA). In addition, we discuss the use of immunoprecipitation and mass spectrometry analysis for the detection of novel neuronal surface antigens, which is a crucial step in further disease classification and the development of novel CBAs.
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We identified de novo nonsense variants in KIDINS220/ARMS in three unrelated patients with spastic paraplegia, intellectual disability, nystagmus, and obesity (SINO). KIDINS220 is an essential scaffold protein coordinating neurotrophin signal pathways in neurites and is spatially and temporally regulated in the brain. Molecular analysis of patients' variants confirmed expression and translation of truncated transcripts similar to recently characterized alternative terminal exon splice isoforms of KIDINS220 KIDINS220 undergoes extensive alternative splicing in specific neuronal populations and developmental time points, reflecting its complex role in neuronal maturation. In mice and humans, KIDINS220 is alternative spliced in the middle region as well as in the last exon. These full-length and KIDINS220 splice variants occur at precise moments in cortical, hippocampal, and motor neuron development, with splice variants similar to the variants seen in our patients and lacking the last exon of KIDINS220 occurring in adult rather than in embryonic brain. We conducted tissue-specific expression studies in zebrafish that resulted in spasms, confirming a functional link with disruption of the KIDINS220 levels in developing neurites. This work reveals a crucial physiological role of KIDINS220 in development and provides insight into how perturbation of the complex interplay of KIDINS220 isoforms and their relative expression can affect neuron control and human metabolism. Altogether, we here show that de novo protein-truncating KIDINS220 variants cause a new syndrome, SINO. This is the first report of KIDINS220 variants causing a human disease.
Assuntos
Deficiência Intelectual/genética , Proteínas de Membrana/genética , Proteínas do Tecido Nervoso/genética , Nistagmo Congênito/genética , Obesidade/genética , Paraplegia/genética , Proteínas de Peixe-Zebra/genética , Processamento Alternativo/genética , Animais , Códon sem Sentido , Modelos Animais de Doenças , Humanos , Deficiência Intelectual/fisiopatologia , Neuritos/metabolismo , Neuritos/patologia , Neurogênese/genética , Neurônios/metabolismo , Neurônios/patologia , Nistagmo Congênito/fisiopatologia , Obesidade/patologia , Células PC12 , Paraplegia/fisiopatologia , Ligação Proteica/genética , Ratos , Transdução de SinaisRESUMO
OBJECTIVE: To determine sensitivity and specificity of a standardized recombinant cell-based indirect immunofluorescence assay (RC-IFA) for anti-Tr antibodies in comparison to a reference procedure. METHODS: Delta/Notch-like epidermal growth factor-related receptor (DNER) was expressed in HEK293 and used as a substrate for RC-IFA. HEK293 control cells expressing CDR2/Yo and CDR2L as well as mock-transfected HEK293 cells were used as controls. Serum samples from 38 patients with anti-Tr antibodies (33 with paraneoplastic cerebellar degeneration [PCD] and Hodgkin lymphoma), 66 patients with anti-Tr-negative PCD, 53 patients with Hodgkin lymphoma without neurologic symptoms, 40 patients with rheumatic diseases, and 42 healthy blood donors were tested for anti-DNER reactivity in the RC-IFA. In addition, RC-IFA results were compared to those from a commercial tissue-based IFA using monkey cerebellum. RESULTS: Using the RC-IFA, anti-DNER was detected in all anti-Tr-positive patients but in none of the controls (sensitivity 100%, 95% confidence interval [CI] 92.8%-100%; specificity 100%, 95% CI 98.7%-100%). In comparison, anti-Tr was not detected in 4 samples with low-titer autoantibodies using the commercial tissue-based assay. Preadsorption of sera with either recombinant full-length DNER or its extracellular domain selectively abolished anti-Tr reactivity. CONCLUSION: Anti-Tr antibodies bind to the extracellular domain of DNER and can be detected by RC-IFA using HEK293 cells expressing the recombinant receptor. The new method performs better than a frequently used commercial tissue-based indirect immunofluorescence assay (IFA) in samples with low-titer antibodies. CLASSIFICATION OF EVIDENCE: This study provides Class II evidence that RC-IFA accurately detects anti-Tr as compared to conventional IFA.
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OBJECTIVE: Autoimmune encephalitis associated with autoantibodies against the N-methyl-d-aspartate receptor (NMDAR) often presents with behavioural change. Our objective was to describe in detail the psychiatric presentation and pathways to care in order to aid the early diagnosis of NMDAR encephalitis. METHODS: Sera and cerebrospinal fluid (CSF) from patients with suspected NMDAR encephalitis were tested on HEK 293 cells transfected with the NR1 subunit of the NMDAR. Clinical information was obtained from the referring psychiatrists and neurologists and by review of the clinical records. RESULTS: Samples from 15 patients (13 female, 2 male, mean age 24 years, range 5-56 years) tested anti-NMDAR positive. Twelve of the 15 patients (80%) presented with prominent psychiatric symptoms and 8 were initially referred to a psychiatric service. The most prominent initial psychiatric symptoms were anxiety in seven (47%), behavioural change (often bizarre) in six (40%) and agitation in five (33%). All patients developed psychiatric symptoms in the first 6 weeks of illness. Thirteen patients received psychotropic medications: antipsychotics in 12 and benzodiazepines in 11. Treating physicians considered the psychotropic medication not effective in 11 patients resulting in many drug switches. At nadir, all patients were in a very poor condition. However, eight patients (53%) recovered (almost) completely. Outcome tended to be better in patients who had received early immunotherapy or tumour removal. CONCLUSIONS: Autoimmune encephalitis and anti-NMDAR testing in serum and CSF should be considered in patients, especially young females, presenting with atypical psychiatric phenomena. Early diagnosis and treatment will likely improve the prognosis of NMDAR encephalitis.
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OBJECTIVE: Anti-Tr is among the better described autoantibodies in paraneoplastic cerebellar degeneration (PCD) combined with Hodgkin lymphoma (HL); however, the Tr antigen remains unidentified. METHODS: We used immunoprecipitation of total rat brain extract followed by mass spectrometry to identify the antigen recognized by anti-Tr-positive sera. By Western blotting and cell-based assays, we tested a total of 12 anti-Tr-positive and 246 control sera and determined the region of the epitope recognized by the anti-Tr antibodies. Deletion and mutant constructs were generated to further map the antigenic region. RESULTS: Mass spectrometry analysis of immunopurified rat brain extract using 4 different anti-Tr-positive sera led to the identification of Delta/Notch-like epidermal growth factor-related receptor (DNER) as the Tr antigen. All but 1 of 246 control samples were negative in the HeLa cell-based screening assay, whereas 12 of the 12 anti-Tr-positive sera stained hemagglutinin-tagged DNER-expressing cells. Only 1 control subject with HL but no ataxia was found to be both DNER and Tr positive. Using deletion constructs, we pinpointed the main epitope to the extracellular domain. Knockdown of endogenous DNER in hippocampal and N-glycosylation mutations abolished the anti-Tr staining, indicating that glycosylation of DNER is required for it to be recognized by anti-Tr antibodies. INTERPRETATION: DNER is the antigen detected by anti-Tr-positive sera. Presence of anti-Tr antibodies in patients with PCD and HL or HL only can now be screened quickly and reliably by using a cell-based screening assay.
Assuntos
Autoanticorpos/sangue , Proteínas do Tecido Nervoso/imunologia , Degeneração Paraneoplásica Cerebelar/imunologia , Degeneração Paraneoplásica Cerebelar/metabolismo , Receptores de Superfície Celular/metabolismo , Sequências de Repetição em Tandem/imunologia , Adulto , Idoso , Animais , Linhagem Celular Transformada , Criança , Feminino , Glicosilação , Hipocampo/patologia , Doença de Hodgkin/imunologia , Humanos , Masculino , Espectrometria de Massas , Pessoa de Meia-Idade , Proteínas do Tecido Nervoso/metabolismo , Neurônios/metabolismo , Ratos , Receptores de Superfície Celular/imunologia , Adulto Jovem , beta-Galactosidase/metabolismoRESUMO
We describe a syndrome of primary microcephaly with simplified gyral pattern in combination with severe infantile epileptic encephalopathy and early-onset permanent diabetes in two unrelated consanguineous families with at least three affected children. Linkage analysis revealed a region on chromosome 18 with a significant LOD score of 4.3. In this area, two homozygous nonconserved missense mutations in immediate early response 3 interacting protein 1 (IER3IP1) were found in patients from both families. IER3IP1 is highly expressed in the fetal brain cortex and fetal pancreas and is thought to be involved in endoplasmic reticulum stress response. We reported one of these families previously in a paper on Wolcott-Rallison syndrome (WRS). WRS is characterized by increased apoptotic cell death as part of an uncontrolled unfolded protein response. Increased apoptosis has been shown to be a cause of microcephaly in animal models. An autopsy specimen from one patient showed increased apoptosis in the cerebral cortex and pancreas beta cells, implicating premature cell death as the pathogenetic mechanism. Both patient fibroblasts and control fibroblasts treated with siRNA specific for IER3IP1 showed an increased susceptibility to apoptotic cell death under stress conditions in comparison to controls. This directly implicates IER3IP1 in the regulation of cell survival. Identification of IER3IP1 mutations sheds light on the mechanisms of brain development and on the pathogenesis of infantile epilepsy and early-onset permanent diabetes.
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Apoptose , Diabetes Mellitus/patologia , Epilepsia/complicações , Epilepsia/patologia , Microcefalia/complicações , Células-Tronco Neurais/patologia , Sequência de Aminoácidos , Apoptose/efeitos dos fármacos , Encéfalo/efeitos dos fármacos , Encéfalo/metabolismo , Encéfalo/patologia , Proteínas de Transporte/química , Proteínas de Transporte/genética , Pré-Escolar , Biologia Computacional , Família , Evolução Fatal , Feminino , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Fibroblastos/patologia , Ligação Genética/efeitos dos fármacos , Humanos , Lactente , Recém-Nascido , Imageamento por Ressonância Magnética , Masculino , Proteínas de Membrana/química , Proteínas de Membrana/genética , Microcefalia/patologia , Dados de Sequência Molecular , Mutação/genética , Células-Tronco Neurais/efeitos dos fármacos , Células-Tronco Neurais/metabolismo , Linhagem , Fator de Necrose Tumoral alfa/farmacologiaRESUMO
Goldberg-Shprintzen syndrome (GOSHS) is a rare clinical disorder characterized by central and enteric nervous system defects. This syndrome is caused by inactivating mutations in the Kinesin Binding Protein (KBP) gene, which encodes a protein of which the precise function is largely unclear. We show that KBP expression is up-regulated during neuronal development in mouse cortical neurons. Moreover, KBP-depleted PC12 cells were defective in nerve growth factor-induced differentiation and neurite outgrowth, suggesting that KBP is required for cell differentiation and neurite development. To identify KBP interacting proteins, we performed a yeast two-hybrid screen and found that KBP binds almost exclusively to microtubule associated or related proteins, specifically SCG10 and several kinesins. We confirmed these results by validating KBP interaction with one of these proteins: SCG10, a microtubule destabilizing protein. Zebrafish studies further demonstrated an epistatic interaction between KBP and SCG10 in vivo. To investigate the possibility of direct interaction between KBP and microtubules, we undertook co-localization and in vitro binding assays, but found no evidence of direct binding. Thus, our data indicate that KBP is involved in neuronal differentiation and that the central and enteric nervous system defects seen in GOSHS are likely caused by microtubule-related defects.
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Proteínas de Transporte/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Proteínas de Membrana/metabolismo , Microtúbulos/metabolismo , Neurogênese , Serpinas/metabolismo , Estatmina/metabolismo , Proteínas de Peixe-Zebra/metabolismo , Animais , Proteínas de Ligação ao Cálcio , Proteínas de Transporte/genética , Linhagem Celular , Células Cultivadas , Anormalidades Craniofaciais/genética , Anormalidades Craniofaciais/metabolismo , Modelos Animais de Doenças , Células HeLa , Doença de Hirschsprung/genética , Doença de Hirschsprung/metabolismo , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Proteínas de Membrana/genética , Camundongos , Camundongos Endogâmicos C57BL , Proteínas dos Microtúbulos , Células NIH 3T3 , Neurônios/citologia , Neurônios/metabolismo , Células PC12 , Ligação Proteica , Ratos , Serpinas/genética , Estatmina/genética , Proteínas de Peixe-Zebra/genéticaRESUMO
The receptor tyrosine kinase Ret plays a critical role in the development of the mammalian excretory and enteric nervous systems. Differential splicing of the primary Ret transcript results in the generation of two main isoforms, Ret9 and Ret51, whose C-terminal amino acid tails diverge after tyrosine (Y) 1062. Monoisoformic mice expressing only Ret9 develop normally and are healthy and fertile. In contrast, animals expressing only Ret51 have aganglionosis of the distal gut and hypoplastic kidneys. By generating monoisoformic mice in which Y1062 of Ret9 has been mutated to phenylalanine, we demonstrate that this amino acid has a critical role in Ret9 signaling that is necessary for the development of the kidneys and the enteric nervous system. These findings argue that the distinct activities of Ret9 and Ret51 result from the differential regulation of Y1062 by C-terminal flanking sequences. However, a mutation which places Y1062 of Ret51 in a Ret9 context improves only marginally the ability of Ret51 to support renal and enteric nervous system development. Finally, monoisoformic mice expressing a variant of Ret9 in which a C-terminal PDZ-binding motif was mutated develop normally and are healthy. Our studies identify Y1062 as a critical regulator of Ret9 signaling and suggest that Ret51-specific motifs are likely to inhibit the activity of this isoform.
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Sistema Nervoso Entérico/crescimento & desenvolvimento , Rim/crescimento & desenvolvimento , Fosfotirosina/fisiologia , Proteínas Proto-Oncogênicas c-ret/fisiologia , Processamento Alternativo , Motivos de Aminoácidos , Animais , Animais Recém-Nascidos , Humanos , Isoenzimas/biossíntese , Isoenzimas/genética , Isoenzimas/fisiologia , Camundongos , Camundongos Mutantes , Mutação , Proteínas Proto-Oncogênicas c-ret/biossíntese , Proteínas Proto-Oncogênicas c-ret/genética , Transdução de SinaisRESUMO
The enteric nervous system (ENS) in vertebrates is derived mainly from vagal neural crest cells that enter the foregut and colonize the entire wall of the gastrointestinal tract. Failure to completely colonize the gut results in the absence of enteric ganglia (Hirschsprung's disease). Two signaling systems mediated by RET and EDNRB have been identified as critical players in enteric neurogenesis. We demonstrate that interaction between these signaling pathways controls ENS development throughout the intestine. Activation of EDNRB specifically enhances the effect of RET signaling on the proliferation of uncommitted ENS progenitors. In addition, we reveal novel antagonistic roles of these pathways on the migration of ENS progenitors. Protein kinase A is a key component of the molecular mechanisms that integrate signaling by the two receptors. Our data provide strong evidence that the coordinate and balanced interaction between receptor tyrosine kinases and G protein-coupled receptors controls the development of the nervous system in mammals.
Assuntos
Sistema Nervoso Entérico/citologia , Sistema Nervoso Entérico/fisiologia , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Proteínas Proto-Oncogênicas/fisiologia , Receptores Proteína Tirosina Quinases/fisiologia , Receptor de Endotelina B/fisiologia , Receptores Acoplados a Proteínas G/fisiologia , Transdução de Sinais/fisiologia , Células-Tronco/fisiologia , Animais , Células Cultivadas , Sistema Nervoso Entérico/embriologia , Sistema Nervoso Entérico/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Proteínas Proto-Oncogênicas/biossíntese , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas c-ret , Receptores Proteína Tirosina Quinases/biossíntese , Receptores Proteína Tirosina Quinases/genética , Receptor de Endotelina B/biossíntese , Receptor de Endotelina B/genética , Receptores de Endotelina/biossíntese , Receptores de Endotelina/genética , Receptores Acoplados a Proteínas G/biossíntese , Receptores Acoplados a Proteínas G/genética , Células-Tronco/metabolismoRESUMO
The majority of neurones and glia of the enteric nervous system (ENS) are derived from the vagal neural crest. Shortly after emigration from the neural tube, ENS progenitors invade the anterior foregut and, migrating in a rostrocaudal direction, colonise in an orderly fashion the rest of the foregut, the midgut and the hindgut. We provide evidence that activation of the receptor tyrosine kinase RET by glial cell line-derived neurotrophic factor (GDNF) is required for the directional migration of ENS progenitors towards and within the gut wall. We find that neural crest-derived cells present within foetal small intestine explants migrate towards an exogenous source of GDNF in a RET-dependent fashion. Consistent with an in vivo role of GDNF in the migration of ENS progenitors, we demonstrate that Gdnf is expressed at high levels in the gut of mouse embryos in a spatially and temporally regulated manner. Thus, during invasion of the foregut by vagal-derived neural crest cells, expression of Gdnf was restricted to the mesenchyme of the stomach, ahead of the invading NC cells. Twenty-four hours later and as the ENS progenitors were colonising the midgut, Gdnf expression was upregulated in a more posterior region - the caecum anlage. In further support of a role of endogenous GDNF in enteric neural crest cell migration, we find that in explant cultures GDNF produced by caecum is sufficient to attract NC cells residing in more anterior gut segments. In addition, two independently generated loss-of-function alleles of murine Ret, Ret.k- and miRet51, result in characteristic defects of neural crest cell migration within the developing gut. Finally, we identify phosphatidylinositol-3 kinase and the mitogen-activated protein kinase signalling pathways as playing crucial roles in the migratory response of enteric neural crest cells to GDNF.
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Proteínas de Drosophila , Sistema Nervoso Entérico/citologia , Sistema Nervoso Entérico/embriologia , Proteínas Proto-Oncogênicas/metabolismo , Receptores Proteína Tirosina Quinases/metabolismo , Células-Tronco/metabolismo , Animais , Células COS , Ceco/citologia , Ceco/embriologia , Ceco/metabolismo , Movimento Celular/genética , Células Cultivadas , Ativação Enzimática , Fator Neurotrófico Derivado de Linhagem de Célula Glial , Receptores de Fator Neurotrófico Derivado de Linhagem de Célula Glial , Mamíferos/embriologia , Mesoderma/metabolismo , Camundongos , Camundongos Mutantes , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Mutação , Fatores de Crescimento Neural/metabolismo , Crista Neural/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas c-ret , Receptores Proteína Tirosina Quinases/genética , Transdução de Sinais , Estômago/citologia , Estômago/embriologiaRESUMO
The RING domain is a cysteine-rich zinc-binding motif, which is found in a wide variety of proteins, among which are several proto-oncogenes and the gene implicated in autosomal recessive juvenile parkinsonism, Parkin. The domain mediates binding to other proteins, either via their RING domains or other motifs. In several proteins, RING domains are found in combination with other cysteine-rich binding motifs and some proteins contain two RING domains. Recent evidence suggests that RING finger proteins function in the ubiquitin pathway as E3 ligases. A variant of the RING domain is the RING-H2 domain, in which one of the cysteines is replaced by a histidine. We have cloned and characterized a novel gene, RNF32, located on chromosome 7q36. RNF32 is contained in 37 kb of genomic DNA and consists of 9 constitutive and 8 alternatively spliced exons, most of which are alternative first exons. A long and a short transcript of the gene are expressed; the short transcript containing exons 1-4 only. This gene encodes two RING-H2 domains separated by an IQ domain of unknown function. This is the first reported gene with a double RING-H2 domain. In humans, RNF32 overlaps with a processed retroposon located on the opposite strand, C7orf13. RNF32 is specifically expressed in testis and ovary, whereas C7orf13 is testis-specific, suggesting that its expression may be regulated by elements in the RNF32 promoter region. RNF32 is expressed during spermatogenesis, most likely in spermatocytes and/or in spermatids, suggesting a possible role in sperm formation.