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Heterozygous de novo mutations in the Activity-Dependent Neuroprotective Homeobox (ADNP) gene underlie Helsmoortel-Van der Aa syndrome (HVDAS). Most of these mutations are situated in the last exon and we previously demonstrated escape from nonsense-mediated decay by detecting mutant ADNP mRNA in patient blood. In this study, wild-type and ADNP mutants are investigated at the protein level and therefore optimal detection of the protein is required. Detection of ADNP by means of western blotting has been ambiguous with reported antibodies resulting in non-specific bands without unique ADNP signal. Validation of an N-terminal ADNP antibody (Aviva Systems) using a blocking peptide competition assay allowed to differentiate between specific and non-specific signals in different sample materials, resulting in a unique band signal around 150 kDa for ADNP, above its theoretical molecular weight of 124 kDa. Detection with different C-terminal antibodies confirmed the signals at an observed molecular weight of 150 kDa. Our antibody panel was subsequently tested by immunoblotting, comparing parental and homozygous CRISPR/Cas9 endonuclease-mediated Adnp knockout cell lines and showed disappearance of the 150 kDa signal, indicative for intact ADNP. By means of both a GFPSpark and Flag-tag N-terminally fused to a human ADNP expression vector, we detected wild-type ADNP together with mutant forms after introduction of patient mutations in E. coli expression systems by site-directed mutagenesis. Furthermore, we were also able to visualize endogenous ADNP with our C-terminal antibody panel in heterozygous cell lines carrying ADNP patient mutations, while the truncated ADNP mutants could only be detected with epitope-tag-specific antibodies, suggesting that addition of an epitope-tag possibly helps stabilizing the protein. However, western blotting of patient-derived hiPSCs, immortalized lymphoblastoid cell lines and post-mortem patient brain material failed to detect a native mutant ADNP protein. In addition, an N-terminal immunoprecipitation-competent ADNP antibody enriched truncating mutants in overexpression lysates, whereas implementation of the same method failed to enrich a possible native mutant protein in immortalized patient-derived lymphoblastoid cell lines. This study aims to shape awareness for critical assessment of mutant ADNP protein analysis in Helsmoortel-Van der Aa syndrome.
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Proteínas de Homeodomínio , Proteínas do Tecido Nervoso , Humanos , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Mutação , Células HEK293 , Transtorno do Espectro Autista , Cardiopatias , Fácies , Transtornos do NeurodesenvolvimentoRESUMO
BACKGROUND: Helsmoortel-Van der Aa syndrome is a neurodevelopmental disorder in which patients present with autism, intellectual disability, and frequent extra-neurological features such as feeding and gastrointestinal problems, visual impairments, and cardiac abnormalities. All patients exhibit heterozygous de novo nonsense or frameshift stop mutations in the Activity-Dependent Neuroprotective Protein (ADNP) gene, accounting for a prevalence of 0.2% of all autism cases worldwide. ADNP fulfills an essential chromatin remodeling function during brain development. In this study, we investigated the cerebellum of a died 6-year-old male patient with the c.1676dupA/p.His559Glnfs*3 ADNP mutation. RESULTS: The clinical presentation of the patient was representative of the Helsmoortel-Van der Aa syndrome. During his lifespan, he underwent two liver transplantations after which the child died because of multiple organ failure. An autopsy was performed, and various tissue samples were taken for further analysis. We performed a molecular characterization of the cerebellum, a brain region involved in motor coordination, known for its highest ADNP expression and compared it to an age-matched control subject. Importantly, epigenome-wide analysis of the ADNP cerebellum identified CpG methylation differences and expression of multiple pathways causing neurodevelopmental delay. Interestingly, transcription factor motif enrichment analysis of differentially methylated genes showed that the ADNP binding motif was the most significantly enriched. RNA sequencing of the autopsy brain further identified downregulation of the WNT signaling pathway and autophagy defects as possible causes of neurodevelopmental delay. Ultimately, label-free quantification mass spectrometry identified differentially expressed proteins involved in mitochondrial stress and sirtuin signaling pathways amongst others. Protein-protein interaction analysis further revealed a network including chromatin remodelers (ADNP, SMARCC2, HDAC2 and YY1), autophagy-related proteins (LAMP1, BECN1 and LC3) as well as a key histone deacetylating enzyme SIRT1, involved in mitochondrial energy metabolism. The protein interaction of ADNP with SIRT1 was further biochemically validated through the microtubule-end binding proteins EB1/EB3 by direct co-immunoprecipitation in mouse cerebellum, suggesting important mito-epigenetic crosstalk between chromatin remodeling and mitochondrial energy metabolism linked to autophagy stress responses. This is further supported by mitochondrial activity assays and stainings in patient-derived fibroblasts which suggest mitochondrial dysfunctions in the ADNP deficient human brain. CONCLUSION: This study forms the baseline clinical and molecular characterization of an ADNP autopsy cerebellum, providing novel insights in the disease mechanisms of the Helsmoortel-Van der Aa syndrome. By combining multi-omic and biochemical approaches, we identified a novel SIRT1-EB1/EB3-ADNP protein complex which may contribute to autophagic flux alterations and impaired mitochondrial metabolism in the Helsmoortel-Van der Aa syndrome and holds promise as a new therapeutic target.
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Transtorno Autístico , Deficiência Intelectual , Masculino , Criança , Animais , Camundongos , Humanos , Deficiência Intelectual/genética , Transtorno Autístico/genética , Sirtuína 1/genética , Sirtuína 1/metabolismo , Genes Mitocondriais , Proteínas de Homeodomínio/genética , Cerebelo/metabolismo , Autopsia , Metilação , Proteínas do Tecido Nervoso/metabolismo , Proteínas de Ligação a DNA/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
We examined more than 97,000 families from four neurodevelopmental disease cohorts and the UK Biobank to identify phenotypic and genetic patterns in parents contributing to neurodevelopmental disease risk in children. We identified within- and cross-disorder correlations between six phenotypes in parents and children, such as obsessive-compulsive disorder (R = 0.32-0.38, p < 10-126). We also found that measures of sub-clinical autism features in parents are associated with several autism severity measures in children, including biparental mean Social Responsiveness Scale scores and proband Repetitive Behaviors Scale scores (regression coefficient = 0.14, p = 3.38 × 10-4). We further describe patterns of phenotypic similarity between spouses, where spouses show correlations for six neurological and psychiatric phenotypes, including a within-disorder correlation for depression (R = 0.24-0.68, p < 0.001) and a cross-disorder correlation between anxiety and bipolar disorder (R = 0.09-0.22, p < 10-92). Using a simulated population, we also found that assortative mating can lead to increases in disease liability over generations and the appearance of "genetic anticipation" in families carrying rare variants. We identified several families in a neurodevelopmental disease cohort where the proband inherited multiple rare variants in disease-associated genes from each of their affected parents. We further identified parental relatedness as a risk factor for neurodevelopmental disorders through its inverse relationship with variant pathogenicity and propose that parental relatedness modulates disease risk by increasing genome-wide homozygosity in children (R = 0.05-0.26, p < 0.05). Our results highlight the utility of assessing parent phenotypes and genotypes toward predicting features in children who carry rare variably expressive variants and implicate assortative mating as a risk factor for increased disease severity in these families.
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Transtorno Autístico , Transtorno Bipolar , Criança , Humanos , Virulência , Pais , Família , Transtorno Autístico/genética , Transtorno Bipolar/genéticaRESUMO
We examined more than 38,000 spouse pairs from four neurodevelopmental disease cohorts and the UK Biobank to identify phenotypic and genetic patterns in parents associated with neurodevelopmental disease risk in children. We identified correlations between six phenotypes in parents and children, including correlations of clinical diagnoses such as obsessive-compulsive disorder (R=0.31-0.49, p<0.001), and two measures of sub-clinical autism features in parents affecting several autism severity measures in children, such as bi-parental mean Social Responsiveness Scale (SRS) scores affecting proband SRS scores (regression coefficient=0.11, p=0.003). We further describe patterns of phenotypic and genetic similarity between spouses, where spouses show both within- and cross-disorder correlations for seven neurological and psychiatric phenotypes, including a within-disorder correlation for depression (R=0.25-0.72, p<0.001) and a cross-disorder correlation between schizophrenia and personality disorder (R=0.20-0.57, p<0.001). Further, these spouses with similar phenotypes were significantly correlated for rare variant burden (R=0.07-0.57, p<0.0001). We propose that assortative mating on these features may drive the increases in genetic risk over generations and the appearance of "genetic anticipation" associated with many variably expressive variants. We further identified parental relatedness as a risk factor for neurodevelopmental disorders through its inverse correlations with burden and pathogenicity of rare variants and propose that parental relatedness drives disease risk by increasing genome-wide homozygosity in children (R=0.09-0.30, p<0.001). Our results highlight the utility of assessing parent phenotypes and genotypes in predicting features in children carrying variably expressive variants and counseling families carrying these variants.
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BACKGROUND: Individuals affected with autism often suffer additional co-morbidities such as intellectual disability. The genes contributing to autism cluster on a relatively limited number of cellular pathways, including chromatin remodeling. However, limited information is available on how mutations in single genes can result in such pleiotropic clinical features in affected individuals. In this review, we summarize available information on one of the most frequently mutated genes in syndromic autism the Activity-Dependent Neuroprotective Protein (ADNP). RESULTS: Heterozygous and predicted loss-of-function ADNP mutations in individuals inevitably result in the clinical presentation with the Helsmoortel-Van der Aa syndrome, a frequent form of syndromic autism. ADNP, a zinc finger DNA-binding protein has a role in chromatin remodeling: The protein is associated with the pericentromeric protein HP1, the SWI/SNF core complex protein BRG1, and other members of this chromatin remodeling complex and, in murine stem cells, with the chromodomain helicase CHD4 in a ChAHP complex. ADNP has recently been shown to possess R-loop processing activity. In addition, many additional functions, for instance, in association with cytoskeletal proteins have been linked to ADNP. CONCLUSIONS: We here present an integrated evaluation of all current aspects of gene function and evaluate how abnormalities in chromatin remodeling might relate to the pleiotropic clinical presentation in individual"s" with Helsmoortel-Van der Aa syndrome.
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Anormalidades Múltiplas , Transtorno Autístico , Deficiência Intelectual , Humanos , Animais , Camundongos , Transtorno Autístico/genética , Cromatina , Metilação de DNA , Proteínas de Homeodomínio/genética , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Deficiência Intelectual/genética , Anormalidades Múltiplas/genéticaRESUMO
An amendment to this paper has been published and can be accessed via a link at the top of the paper.
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Most genes associated with neurodevelopmental disorders (NDDs) were identified with an excess of de novo mutations (DNMs) but the significance in case-control mutation burden analysis is unestablished. Here, we sequence 63 genes in 16,294 NDD cases and an additional 62 genes in 6,211 NDD cases. By combining these with published data, we assess a total of 125 genes in over 16,000 NDD cases and compare the mutation burden to nonpsychiatric controls from ExAC. We identify 48 genes (25 newly reported) showing significant burden of ultra-rare (MAF < 0.01%) gene-disruptive mutations (FDR 5%), six of which reach family-wise error rate (FWER) significance (p < 1.25E-06). Among these 125 targeted genes, we also reevaluate DNM excess in 17,426 NDD trios with 6,499 new autism trios. We identify 90 genes enriched for DNMs (FDR 5%; e.g., GABRG2 and UIMC1); of which, 61 reach FWER significance (p < 3.64E-07; e.g., CASZ1). In addition to doubling the number of patients for many NDD risk genes, we present phenotype-genotype correlations for seven risk genes (CTCF, HNRNPU, KCNQ3, ZBTB18, TCF12, SPEN, and LEO1) based on this large-scale targeted sequencing effort.
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Predisposição Genética para Doença , Transtornos do Neurodesenvolvimento/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fator de Ligação a CCCTC/genética , Estudos de Casos e Controles , Estudos de Coortes , Análise Mutacional de DNA , Proteínas de Ligação a DNA/genética , Feminino , Estudos de Associação Genética , Ribonucleoproteínas Nucleares Heterogêneas Grupo U/genética , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Canal de Potássio KCNQ3/genética , Masculino , Mutação , Proteínas de Ligação a RNA/genética , Proteínas Repressoras/genética , Fatores de Transcrição/genéticaRESUMO
Upon the discovery of numerous genes involved in the pathogenesis of neurodevelopmental disorders, several studies showed that a significant proportion of these genes converge on common pathways and protein networks. Here, we used a reversed approach, by screening the AnkyrinG protein-protein interaction network for genetic variation in a large cohort of 1009 cases with neurodevelopmental disorders. We identified a significant enrichment of de novo potentially disease-causing variants in this network, confirming that this protein network plays an important role in the emergence of several neurodevelopmental disorders.
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Anquirinas/genética , Redes Reguladoras de Genes , Transtornos do Neurodesenvolvimento/genética , Polimorfismo Genético , Mapas de Interação de Proteínas , Anquirinas/metabolismo , Bases de Dados Genéticas , HumanosRESUMO
Given our recent discovery of somatic mutations in autism spectrum disorder (ASD)/intellectual disability (ID) genes in postmortem aged Alzheimer's disease brains correlating with increasing tauopathy, it is important to decipher if tauopathy is underlying brain imaging results of atrophy in ASD/ID children. We concentrated on activity-dependent neuroprotective protein (ADNP), a prevalent autism gene. The unique availability of multiple postmortem brain sections of a 7-year-old male, heterozygous for ADNP de novo mutation c.2244Adup/p.His559Glnfs*3 allowed exploration of tauopathy, reflecting on a general unexplored mechanism. The tested subject exhibited autism, fine motor delays, severe intellectual disability and seizures. The patient died after multiple organ failure following liver transplantation. To compare to other ADNP syndrome mutations, immortalized lymphoblastoid cell lines from three different patients (including ADNP p.Arg216*, p.Lys408Valfs*31, and p.Tyr719* heterozygous dominant mutations) and a control were subjected to RNA-seq. Immunohistochemistry, high-throughput gene expression profiles in numerous postmortem tissues followed. Comparisons to a control brain and to extensive datasets were used. Live cell imaging investigated Tau-microtubule interaction, protecting against tauopathy. Extensive child brain tauopathy paralleled by multiple gene expression changes was discovered. Tauopathy was explained by direct mutation effects on Tau-microtubule interaction and correction by the ADNP active snippet NAP. Significant pathway changes (empirical P value < 0.05) included over 100 genes encompassing neuroactive ligand-receptor and cytokine-cytokine receptor interaction, MAPK and calcium signaling, axon guidance and Wnt signaling pathways. Changes were also seen in steroid biosynthesis genes, suggesting sex differences. Selecting the most affected genes by the ADNP mutations for gene expression analysis, in multiple postmortem tissues, identified Tau (MAPT)-gene-related expression changes compared with extensive normal gene expression (RNA-seq) databases. ADNP showed relatively reduced expression in the ADNP syndrome cerebellum, which was also observed for 25 additional genes (representing >50% of the tested genes), including NLGN1, NLGN2, PAX6, SMARCA4, and SNAP25, converging on nervous system development and tauopathy. NAP provided protection against mutated ADNP disrupted Tau-microtubule association. In conclusion, tauopathy may explain brain-imaging findings in ADNP syndrome children and may provide a new direction for the development of tauopathy protecting drug candidates like NAP in ASD/ID.
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Transtorno do Espectro Autista , Transtorno Autístico , Tauopatias , Idoso , Transtorno do Espectro Autista/genética , Biomarcadores , Encéfalo/diagnóstico por imagem , Encéfalo/metabolismo , Criança , DNA Helicases , Feminino , Proteínas de Homeodomínio/metabolismo , Humanos , Masculino , Proteínas do Tecido Nervoso , Proteínas Nucleares , Tauopatias/genética , Fatores de TranscriçãoRESUMO
BACKGROUND: Fragile X syndrome (FXS) is the most frequent cause of inherited intellectual disability and the most commonly identified monogenic cause of autism. Recent studies have shown that long-term pathological consequences of FXS are not solely confined to the central nervous system (CNS) but rather extend to other physiological dysfunctions in peripheral organs. To gain insights into possible immune dysfunctions in FXS, we profiled a large panel of immune-related biomarkers in the serum of FXS patients and healthy controls. METHODS: We have used a sensitive and robust Electro Chemi Luminescence (ECL)-based immunoassay to measure the levels of 52 cytokines in the serum of n = 25 FXS patients and n = 29 healthy controls. We then used univariate statistics and multivariate analysis, as well as an advanced unsupervised clustering method, to identify combinations of immune-related biomarkers that could discriminate FXS patients from healthy individuals. RESULTS: While the majority of the tested cytokines were present at similar levels in FXS patients and healthy individuals, nine chemokines, CCL2, CCL3, CCL4, CCL11, CCL13, CCL17, CCL22, CCL26 and CXCL10, were present at much lower levels in FXS patients. Using robust regression, we show that six of these biomarkers (CCL2, CCL3, CCL11, CCL22, CCL26 and CXCL10) were negatively associated with FXS diagnosis. Finally, applying the K-sparse unsupervised clustering method to the biomarker dataset allowed for the identification of two subsets of individuals, which essentially matched the FXS and healthy control categories. CONCLUSIONS: Our data show that FXS patients exhibit reduced serum levels of several chemokines and may therefore exhibit impaired immune responses. The present study also highlights the power of unsupervised clustering methods to identify combinations of biomarkers for diagnosis and prognosis in medicine.
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Quimiocinas/sangue , Citocinas/sangue , Síndrome do Cromossomo X Frágil/sangue , Adolescente , Biomarcadores/sangue , Estudos de Casos e Controles , Criança , Pré-Escolar , Feminino , Humanos , Masculino , Prognóstico , Adulto JovemRESUMO
Histones mediate dynamic packaging of nuclear DNA in chromatin, a process that is precisely controlled to guarantee efficient compaction of the genome and proper chromosomal segregation during cell division and to accomplish DNA replication, transcription, and repair. Due to the important structural and regulatory roles played by histones, it is not surprising that histone functional dysregulation or aberrant levels of histones can have severe consequences for multiple cellular processes and ultimately might affect development or contribute to cell transformation. Recently, germline frameshift mutations involving the C-terminal tail of HIST1H1E, which is a widely expressed member of the linker histone family and facilitates higher-order chromatin folding, have been causally linked to an as-yet poorly defined syndrome that includes intellectual disability. We report that these mutations result in stable proteins that reside in the nucleus, bind to chromatin, disrupt proper compaction of DNA, and are associated with a specific methylation pattern. Cells expressing these mutant proteins have a dramatically reduced proliferation rate and competence, hardly enter into the S phase, and undergo accelerated senescence. Remarkably, clinical assessment of a relatively large cohort of subjects sharing these mutations revealed a premature aging phenotype as a previously unrecognized feature of the disorder. Our findings identify a direct link between aberrant chromatin remodeling, cellular senescence, and accelerated aging.
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Senescência Celular/fisiologia , Histonas/fisiologia , Aneuploidia , Nucléolo Celular/metabolismo , Criança , Cromatina/metabolismo , Metilação de DNA , Feminino , Histonas/química , Humanos , Lactente , Masculino , Pessoa de Meia-IdadeRESUMO
BACKGROUND: The 15q11.2 deletion is frequently identified in the neurodevelopmental clinic. Case-control studies have associated the 15q11.2 deletion with neurodevelopmental disorders, and clinical case series have attempted to delineate a microdeletion syndrome with considerable phenotypic variability. The literature on this deletion is extensive and confusing, which is a challenge for genetic counselling. The aim of this study was to estimate the effect size of the 15q11.2 deletion and quantify its contribution to neurodevelopmental disorders. METHODS: We performed meta-analyses on new and previously published case-control studies and used statistical models trained in unselected populations with cognitive assessments. We used new (n=241) and previously published (n=150) data from a clinically referred group of deletion carriers. 15q11.2 duplications (new n=179 and previously published n=35) were used as a neutral control variant. RESULTS: The deletion decreases IQ by 4.3 points. The estimated ORs and respective frequencies in deletion carriers for intellectual disabilities, schizophrenia and epilepsy are 1.7 (3.4%), 1.5 (2%) and 3.1 (2.1%), respectively. There is no increased risk for heart malformations and autism. In the clinically referred group, the frequency and nature of symptoms in deletions are not different from those observed in carriers of the 15q11.2 duplication suggesting that most of the reported symptoms are due to ascertainment bias. CONCLUSIONS: We recommend that the deletion should be classified as 'pathogenic of mild effect size'. Since it explains only a small proportion of the phenotypic variance in carriers, it is not worth discussing in the developmental clinic or in a prenatal setting.
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Transtorno Autístico/genética , Variações do Número de Cópias de DNA , Epilepsia/genética , Cardiopatias/genética , Deficiência Intelectual/genética , Transtornos do Neurodesenvolvimento/genética , Estudos de Casos e Controles , Estudos de Coortes , Feminino , Cardiopatias/congênito , Humanos , Mutação com Perda de Função , Masculino , Deleção de SequênciaRESUMO
The clinical utility of computational phenotyping for both genetic and rare diseases is increasingly appreciated; however, its true potential is yet to be fully realized. Alongside the growing clinical and research availability of sequencing technologies, precise deep and scalable phenotyping is required to serve unmet need in genetic and rare diseases. To improve the lives of individuals affected with rare diseases through deep phenotyping, global big data interrogation is necessary to aid our understanding of disease biology, assist diagnosis, and develop targeted treatment strategies. This includes the application of cutting-edge machine learning methods to image data. As with most digital tools employed in health care, there are ethical and data governance challenges associated with using identifiable personal image data. There are also risks with failing to deliver on the patient benefits of these new technologies, the biggest of which is posed by data siloing. The Minerva Initiative has been designed to enable the public good of deep phenotyping while mitigating these ethical risks. Its open structure, enabling collaboration and data sharing between individuals, clinicians, researchers and private enterprise, is key for delivering precision public health.
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OBJECTIVES: The Fragile X Mental Retardation Protein (FMRP) is a widely expressed RNA-binding protein involved in translation regulation. Since the absence of FMRP leads to Fragile X Syndrome (FXS) and autism, FMRP has been extensively studied in brain. The functions of FMRP in peripheral organs and on metabolic homeostasis remain elusive; therefore, we sought to investigate the systemic consequences of its absence. METHODS: Using metabolomics, in vivo metabolic phenotyping of the Fmr1-KO FXS mouse model and in vitro approaches, we show that the absence of FMRP induced a metabolic shift towards enhanced glucose tolerance and insulin sensitivity, reduced adiposity, and increased ß-adrenergic-driven lipolysis and lipid utilization. RESULTS: Combining proteomics and cellular assays, we highlight that FMRP loss increased hepatic protein synthesis and impacted pathways notably linked to lipid metabolism. Mapping metabolomic and proteomic phenotypes onto a signaling and metabolic network, we predicted that the coordinated metabolic response to FMRP loss was mediated by dysregulation in the abundances of specific hepatic proteins. We experimentally validated these predictions, demonstrating that the translational regulator FMRP associates with a subset of mRNAs involved in lipid metabolism. Finally, we highlight that FXS patients mirror metabolic variations observed in Fmr1-KO mice with reduced circulating glucose and insulin and increased free fatty acids. CONCLUSIONS: Loss of FMRP results in a widespread coordinated systemic response that notably involves upregulation of protein translation in the liver, increased utilization of lipids, and significant changes in metabolic homeostasis. Our study unravels metabolic phenotypes in FXS and further supports the importance of translational regulation in the homeostatic control of systemic metabolism.
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Proteína do X Frágil da Deficiência Intelectual/metabolismo , Síndrome do Cromossomo X Frágil/metabolismo , Glucose/metabolismo , Lipólise , Adipócitos/metabolismo , Animais , Modelos Animais de Doenças , Ácidos Graxos não Esterificados/metabolismo , Feminino , Proteína do X Frágil da Deficiência Intelectual/genética , Síndrome do Cromossomo X Frágil/sangue , Síndrome do Cromossomo X Frágil/patologia , Técnicas de Inativação de Genes , Glucose/análise , Homeostase , Humanos , Insulina/análise , Insulina/metabolismo , Leptina/metabolismo , Fígado/metabolismo , Masculino , Metabolômica , Camundongos , Camundongos Knockout , Biossíntese de Proteínas , Proteômica , RNA Mensageiro/metabolismoRESUMO
Determining pathogenicity of genomic variation identified by next-generation sequencing techniques can be supported by recurrent disruptive variants in the same gene in phenotypically similar individuals. However, interpretation of novel variants in a specific gene in individuals with mild-moderate intellectual disability (ID) without recognizable syndromic features can be challenging and reverse phenotyping is often required. We describe 24 individuals with a de novo disease-causing variant in, or partial deletion of, the F-box only protein 11 gene (FBXO11, also known as VIT1 and PRMT9). FBXO11 is part of the SCF (SKP1-cullin-F-box) complex, a multi-protein E3 ubiquitin-ligase complex catalyzing the ubiquitination of proteins destined for proteasomal degradation. Twenty-two variants were identified by next-generation sequencing, comprising 2 in-frame deletions, 11 missense variants, 1 canonical splice site variant, and 8 nonsense or frameshift variants leading to a truncated protein or degraded transcript. The remaining two variants were identified by array-comparative genomic hybridization and consisted of a partial deletion of FBXO11. All individuals had borderline to severe ID and behavioral problems (autism spectrum disorder, attention-deficit/hyperactivity disorder, anxiety, aggression) were observed in most of them. The most relevant common facial features included a thin upper lip and a broad prominent space between the paramedian peaks of the upper lip. Other features were hypotonia and hyperlaxity of the joints. We show that de novo variants in FBXO11 cause a syndromic form of ID. The current series show the power of reverse phenotyping in the interpretation of novel genetic variances in individuals who initially did not appear to have a clear recognizable phenotype.
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Anormalidades Múltiplas/genética , Comportamento , Proteínas F-Box/genética , Variação Genética , Deficiência Intelectual/genética , Proteína-Arginina N-Metiltransferases/genética , Deleção de Genes , Humanos , SíndromeRESUMO
Mutations in the X chromosomal tRNA 2'Omethyltransferase FTSJ1 cause intellectual disability (ID). Although the gene is ubiquitously expressed affected individuals present no consistent clinical features beyond ID. In order to study the pathological mechanism involved in the aetiology of FTSJ1 deficiency-related cognitive impairment, we generated and characterized an Ftsj1 deficient mouse line based on the gene trapped stem cell line RRD143. Apart from an impaired learning capacity these mice presented with several statistically significantly altered features related to behaviour, pain sensing, bone and energy metabolism, the immune and the hormone system as well as gene expression. These findings show that Ftsj1 deficiency in mammals is not phenotypically restricted to the brain but affects various organ systems. Re-examination of ID patients with FTSJ1 mutations from two previously reported families showed that several features observed in the mouse model were recapitulated in some of the patients. Though the clinical spectrum related to Ftsj1 deficiency in mouse and man is variable, we suggest that an increased pain threshold may be more common in patients with FTSJ1 deficiency. Our findings demonstrate novel roles for Ftsj1 in maintaining proper cellular and tissue functions in a mammalian organism.
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Modelos Animais de Doenças , Deficiência Intelectual/etiologia , Deficiência Intelectual Ligada ao Cromossomo X/genética , Metiltransferases/fisiologia , Mutação , Proteínas Nucleares/genética , tRNA Metiltransferases/fisiologia , Animais , Comportamento Animal , Transtornos Cognitivos/etiologia , Transtornos Cognitivos/patologia , Família , Feminino , Deficiência Intelectual/patologia , Masculino , Metiltransferases/genética , Metiltransferases/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Dor Nociceptiva/etiologia , Dor Nociceptiva/patologia , Proteínas Nucleares/metabolismo , tRNA Metiltransferases/genética , tRNA Metiltransferases/metabolismoRESUMO
PURPOSE: To assess the contribution of rare variants in the genetic background toward variability of neurodevelopmental phenotypes in individuals with rare copy-number variants (CNVs) and gene-disruptive variants. METHODS: We analyzed quantitative clinical information, exome sequencing, and microarray data from 757 probands and 233 parents and siblings who carry disease-associated variants. RESULTS: The number of rare likely deleterious variants in functionally intolerant genes ("other hits") correlated with expression of neurodevelopmental phenotypes in probands with 16p12.1 deletion (n=23, p=0.004) and in autism probands carrying gene-disruptive variants (n=184, p=0.03) compared with their carrier family members. Probands with 16p12.1 deletion and a strong family history presented more severe clinical features (p=0.04) and higher burden of other hits compared with those with mild/no family history (p=0.001). The number of other hits also correlated with severity of cognitive impairment in probands carrying pathogenic CNVs (n=53) or de novo pathogenic variants in disease genes (n=290), and negatively correlated with head size among 80 probands with 16p11.2 deletion. These co-occurring hits involved known disease-associated genes such as SETD5, AUTS2, and NRXN1, and were enriched for cellular and developmental processes. CONCLUSION: Accurate genetic diagnosis of complex disorders will require complete evaluation of the genetic background even after a candidate disease-associated variant is identified.
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Transtorno Autístico/genética , Moléculas de Adesão Celular Neuronais/genética , Triagem de Portadores Genéticos , Metiltransferases/genética , Proteínas do Tecido Nervoso/genética , Proteínas/genética , Transtorno Autístico/fisiopatologia , Proteínas de Ligação ao Cálcio , Cromossomos Humanos Par 16/genética , Cognição/fisiologia , Proteínas do Citoesqueleto , Variações do Número de Cópias de DNA/genética , Feminino , Regulação da Expressão Gênica/genética , Patrimônio Genético , Humanos , Masculino , Moléculas de Adesão de Célula Nervosa , Pais , Linhagem , Fenótipo , Deleção de Sequência/genética , Irmãos , Fatores de TranscriçãoRESUMO
BACKGROUND: In genome-wide screening studies for de novo mutations underlying autism and intellectual disability, mutations in the ADNP gene are consistently reported among the most frequent. ADNP mutations have been identified in children with autism spectrum disorder comorbid with intellectual disability, distinctive facial features, and deficits in multiple organ systems. However, a comprehensive clinical description of the Helsmoortel-Van der Aa syndrome is lacking. METHODS: We identified a worldwide cohort of 78 individuals with likely disruptive mutations in ADNP from January 2014 to October 2016 through systematic literature search, by contacting collaborators, and through direct interaction with parents. Clinicians filled in a structured questionnaire on genetic and clinical findings to enable correlations between genotype and phenotype. Clinical photographs and specialist reports were gathered. Parents were interviewed to complement the written questionnaires. RESULTS: We report on the detailed clinical characterization of a large cohort of individuals with an ADNP mutation and demonstrate a distinctive combination of clinical features, including mild to severe intellectual disability, autism, severe speech and motor delay, and common facial characteristics. Brain abnormalities, behavioral problems, sleep disturbance, epilepsy, hypotonia, visual problems, congenital heart defects, gastrointestinal problems, short stature, and hormonal deficiencies are common comorbidities. Strikingly, individuals with the recurrent p.Tyr719* mutation were more severely affected. CONCLUSIONS: This overview defines the full clinical spectrum of individuals with ADNP mutations, a specific autism subtype. We show that individuals with mutations in ADNP have many overlapping clinical features that are distinctive from those of other autism and/or intellectual disability syndromes. In addition, our data show preliminary evidence of a correlation between genotype and phenotype.
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Proteínas de Homeodomínio/genética , Proteínas do Tecido Nervoso/genética , Transtornos do Neurodesenvolvimento/genética , Anormalidades Múltiplas/genética , Adolescente , Adulto , Transtorno do Espectro Autista/complicações , Transtorno do Espectro Autista/genética , Criança , Pré-Escolar , Estudos de Coortes , Feminino , Humanos , Lactente , Deficiência Intelectual/complicações , Deficiência Intelectual/genética , Masculino , Mutação , Transtornos do Neurodesenvolvimento/complicações , Síndrome , Adulto JovemRESUMO
N-alpha-acetylation is a common co-translational protein modification that is essential for normal cell function in humans. We previously identified the genetic basis of an X-linked infantile lethal Mendelian disorder involving a c.109T>C (p.Ser37Pro) missense variant in NAA10, which encodes the catalytic subunit of the N-terminal acetyltransferase A (NatA) complex. The auxiliary subunit of the NatA complex, NAA15, is the dimeric binding partner for NAA10. Through a genotype-first approach with whole-exome or genome sequencing (WES/WGS) and targeted sequencing analysis, we identified and phenotypically characterized 38 individuals from 33 unrelated families with 25 different de novo or inherited, dominantly acting likely gene disrupting (LGD) variants in NAA15. Clinical features of affected individuals with LGD variants in NAA15 include variable levels of intellectual disability, delayed speech and motor milestones, and autism spectrum disorder. Additionally, mild craniofacial dysmorphology, congenital cardiac anomalies, and seizures are present in some subjects. RNA analysis in cell lines from two individuals showed degradation of the transcripts with LGD variants, probably as a result of nonsense-mediated decay. Functional assays in yeast confirmed a deleterious effect for two of the LGD variants in NAA15. Further supporting a mechanism of haploinsufficiency, individuals with copy-number variant (CNV) deletions involving NAA15 and surrounding genes can present with mild intellectual disability, mild dysmorphic features, motor delays, and decreased growth. We propose that defects in NatA-mediated N-terminal acetylation (NTA) lead to variable levels of neurodevelopmental disorders in humans, supporting the importance of the NatA complex in normal human development.
Assuntos
Anormalidades Múltiplas/genética , Transtorno do Espectro Autista/genética , Predisposição Genética para Doença , Variação Genética , Deficiência Intelectual/genética , Acetiltransferase N-Terminal A/genética , Acetiltransferase N-Terminal E/genética , Adolescente , Adulto , Linhagem Celular , Criança , Éxons/genética , Feminino , Regulação da Expressão Gênica , Humanos , Masculino , Pessoa de Meia-Idade , Mutação/genética , Acetiltransferase N-Terminal A/metabolismo , Acetiltransferase N-Terminal E/metabolismo , Linhagem , Fenótipo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Saccharomyces cerevisiae/metabolismoRESUMO
BACKGROUND: Gamma-aminobutyric acid (GABA) system deficits are integral to the pathophysiologic development of fragile X syndrome (FXS). Ganaxolone, a GABAA receptor positive allosteric modulator, is hypothesized to improve symptoms such as anxiety, hyperactivity, and attention deficits in children with FXS. METHODS: This study was a randomized, double-blind, placebo-controlled, crossover trial of ganaxolone in children with FXS, aged 6-17 years. RESULTS: Sixty-one participants were assessed for eligibility, and 59 were randomized to the study. Fifty-five participants completed at least the first arm and were included in the intention-to-treat analysis; 51 participants completed both treatment arms. There were no statistically significant improvements observed on the primary outcome measure (Clinical Global Impression-Improvement), the key secondary outcome measure (Pediatric Anxiety Rating Scale-R), or any other secondary outcome measures in the overall study population. However, post-hoc analyses revealed positive trends in areas of anxiety, attention, and hyperactivity in participants with higher baseline anxiety and low full-scale IQ scores. No serious adverse events (AEs) occurred, although there was a significant increase in the frequency and severity of AEs related to ganaxolone compared to placebo. CONCLUSIONS: While ganaxolone was found to be safe, there were no significant improvements in the outcome measures in the overall study population. However, ganaxolone in subgroups of children with FXS, including those with higher anxiety or lower cognitive abilities, might have beneficial effects. TRIAL REGISTRATION: ClinicalTrials.gov, NCT01725152.