RESUMO
Despite major advances in genome technology and analysis, >50% of patients with a neurodevelopmental disorder (NDD) remain undiagnosed after extensive evaluation. A point in case is our clinically heterogeneous cohort of NDD patients that remained undiagnosed after FRAXA testing, chromosomal microarray analysis and trio exome sequencing (ES). In this study, we explored the frequency of non-random X chromosome inactivation (XCI) in the mothers of male patients and affected females, the rationale being that skewed XCI might be masking previously discarded genetic variants found on the X chromosome. A multiplex fluorescent PCR-based assay was used to analyse the pattern of XCI after digestion with HhaI methylation-sensitive restriction enzyme. In families with skewed XCI, we re-evaluated trio-based ES and identified pathogenic variants and a deletion on the X chromosome. Linkage analysis and RT-PCR were used to further study the inactive X chromosome allele, and Xdrop long-DNA technology was used to define chromosome deletion boundaries. We found skewed XCI (>90%) in 16/186 (8.6%) mothers of NDD males and in 12/90 (13.3%) NDD females, far beyond the expected rate of XCI in the normal population (3.6%, OR = 4.10; OR = 2.51). By re-analyzing ES and clinical data, we solved 7/28 cases (25%) with skewed XCI, identifying variants in KDM5C, PDZD4, PHF6, TAF1, OTUD5 and ZMYM3, and a deletion in ATRX. We conclude that XCI profiling is a simple assay that targets a subgroup of patients that can benefit from re-evaluation of X-linked variants, thus improving the diagnostic yield in NDD patients and identifying new X-linked disorders.
Assuntos
Genes Ligados ao Cromossomo X , Inativação do Cromossomo X , Feminino , Humanos , Masculino , Mães , Alelos , Cromossomos , Cromossomos Humanos X/genética , Proteínas de Neoplasias/genéticaRESUMO
Posttranscriptional RNA modifications by adenosine-to-inosine (A-to-I) editing are abundant in the brain, yet elucidating functional sites remains challenging. To bridge this gap, we investigate spatiotemporal and genetically regulated A-to-I editing sites across prenatal and postnatal stages of human brain development. More than 10,000 spatiotemporally regulated A-to-I sites were identified that occur predominately in 3' UTRs and introns, as well as 37 sites that recode amino acids in protein coding regions with precise changes in editing levels across development. Hyper-edited transcripts are also enriched in the aging brain and stabilize RNA secondary structures. These features are conserved in murine and non-human primate models of neurodevelopment. Finally, thousands of cis-editing quantitative trait loci (edQTLs) were identified with unique regulatory effects during prenatal and postnatal development. Collectively, this work offers a resolved atlas linking spatiotemporal variation in editing levels to genetic regulatory effects throughout distinct stages of brain maturation.
Assuntos
Inosina , Edição de RNA , Humanos , Animais , Camundongos , Edição de RNA/genética , Inosina/genética , Adenosina/metabolismo , Primatas , Regiões 3' não Traduzidas , Encéfalo/metabolismo , Adenosina Desaminase/metabolismoRESUMO
Detection of Gram-negative bacterial lipid A by the extracellular sensor, myeloid differentiation 2 (MD2)/Toll-like receptor 4 (TLR4), or the intracellular inflammasome sensors, CASP4 and CASP5, induces robust inflammatory responses. The chemical structure of lipid A, specifically its phosphorylation and acylation state, varies across and within bacterial species, potentially allowing pathogens to evade or suppress host immunity. Currently, it is not clear how distinct alterations in the phosphorylation or acylation state of lipid A affect both human TLR4 and CASP4/5 activation. Using a panel of engineered lipooligosaccharides (LOS) derived from Yersinia pestis with defined lipid A structures that vary in their acylation or phosphorylation state, we identified that differences in phosphorylation state did not affect TLR4 or CASP4/5 activation. However, the acylation state differentially impacted TLR4 and CASP4/5 activation. Specifically, all tetra-, penta-, and hexa-acylated LOS variants examined activated CASP4/5-dependent responses, whereas TLR4 responded to penta- and hexa-acylated LOS but did not respond to tetra-acylated LOS or penta-acylated LOS lacking the secondary acyl chain at the 3' position. As expected, lipid A alone was sufficient for TLR4 activation. In contrast, both core oligosaccharide and lipid A were required for robust CASP4/5 inflammasome activation in human macrophages, whereas core oligosaccharide was not required to activate mouse macrophages expressing CASP4. Our findings show that human TLR4 and CASP4/5 detect both shared and nonoverlapping LOS/lipid A structures, which enables the innate immune system to recognize a wider range of bacterial LOS/lipid A and would thereby be expected to constrain the ability of pathogens to evade innate immune detection.
Assuntos
Lipídeo A , Receptor 4 Toll-Like , Acilação , Animais , Humanos , Inflamassomos , Lipídeo A/química , Lipopolissacarídeos , Macrófagos , Camundongos , Receptor 4 Toll-Like/metabolismoRESUMO
BACKGROUND: Sex differences in incidence and/or presentation of schizophrenia (SCZ), major depressive disorder (MDD), and bipolar disorder (BIP) are pervasive. Previous evidence for shared genetic risk and sex differences in brain abnormalities across disorders suggest possible shared sex-dependent genetic risk. METHODS: We conducted the largest to date genome-wide genotype-by-sex (G×S) interaction of risk for these disorders using 85,735 cases (33,403 SCZ, 19,924 BIP, and 32,408 MDD) and 109,946 controls from the PGC (Psychiatric Genomics Consortium) and iPSYCH. RESULTS: Across disorders, genome-wide significant single nucleotide polymorphism-by-sex interaction was detected for a locus encompassing NKAIN2 (rs117780815, p = 3.2 × 10-8), which interacts with sodium/potassium-transporting ATPase (adenosine triphosphatase) enzymes, implicating neuronal excitability. Three additional loci showed evidence (p < 1 × 10-6) for cross-disorder G×S interaction (rs7302529, p = 1.6 × 10-7; rs73033497, p = 8.8 × 10-7; rs7914279, p = 6.4 × 10-7), implicating various functions. Gene-based analyses identified G×S interaction across disorders (p = 8.97 × 10-7) with transcriptional inhibitor SLTM. Most significant in SCZ was a MOCOS gene locus (rs11665282, p = 1.5 × 10-7), implicating vascular endothelial cells. Secondary analysis of the PGC-SCZ dataset detected an interaction (rs13265509, p = 1.1 × 10-7) in a locus containing IDO2, a kynurenine pathway enzyme with immunoregulatory functions implicated in SCZ, BIP, and MDD. Pathway enrichment analysis detected significant G×S interaction of genes regulating vascular endothelial growth factor receptor signaling in MDD (false discovery rate-corrected p < .05). CONCLUSIONS: In the largest genome-wide G×S analysis of mood and psychotic disorders to date, there was substantial genetic overlap between the sexes. However, significant sex-dependent effects were enriched for genes related to neuronal development and immune and vascular functions across and within SCZ, BIP, and MDD at the variant, gene, and pathway levels.
Assuntos
Transtorno Bipolar/genética , Transtorno Depressivo Maior , Transtornos Psicóticos , Esquizofrenia/genética , Caracteres Sexuais , Transtorno Depressivo Maior/genética , Células Endoteliais , Feminino , Predisposição Genética para Doença , Estudo de Associação Genômica Ampla , Humanos , Masculino , Polimorfismo de Nucleotídeo Único , Transtornos Psicóticos/genética , Receptores de Fatores de Crescimento do Endotélio Vascular , SulfurtransferasesRESUMO
BACKGROUND: Lower urinary tract symptoms (LUTS), such as voiding symptoms, overactive bladder, and interstitial cystitis, and anxiety disorders are often comorbid conditions in patients. However, the existing evidence regarding the rates and nature of the co-occurrence of these conditions has not been systematically evaluated. The aim of this study was to examine these relationships. METHODS: We conducted a systematic review and meta-analysis to examine the relationship between LUTS and anxiety. We searched for articles published from January 1990 to July 2019 in PubMed, CENTRAL, PsycINFO, and Google Scholar. Outcomes were anxiety-related disorders and symptoms (clinically significant anxiety) and LUTS. We performed random-effects meta-analyses, inspected funnel plots, and applied the Egger's test to evaluate publication bias. We followed PRISMA guidelines and recorded our protocol on PROSPERO (ID = CRD42019118607). RESULTS: We identified 814 articles, of which 94 fulfilled inclusion criteria, and 23 had sufficient data for meta-analysis. The odds ratio (OR) for clinically significant anxiety among individuals with LUTS was 2.87 (95% CI: 2.38,3.46, p < .001). The OR for LUTS among individuals with clinically significant anxiety was 2.87 (95% CI: 1.07,7.74, p < .001), although very few studies examined this relationship. A large value of I2 index suggests high heterogeneity between studies. CONCLUSION: The results demonstrate a significant association between clinically significant anxiety and LUTS in both females and males. There were limited studies on younger individuals and on individuals ascertained for clinically significant anxiety, which should motivate further study in these areas. Understanding the co-occurrence of these conditions will lead to better prevention and interventions to ameliorate the progression of the symptoms and improve the quality of life. A thorough assessment of anxiety may provide more optimal care for LUTS patients.
Assuntos
Sintomas do Trato Urinário Inferior , Bexiga Urinária Hiperativa , Ansiedade/epidemiologia , Transtornos de Ansiedade/epidemiologia , Feminino , Humanos , Sintomas do Trato Urinário Inferior/epidemiologia , Masculino , Qualidade de VidaRESUMO
There is great interest in the role epigenetic variation induced by non-genetic exposures may play in the context of health and disease. In particular, DNA methylation has previously been shown to be highly dynamic during the earliest stages of development and is influenced by in utero exposures such as maternal smoking and medication. In this study we sought to identify the specific DNA methylation differences in blood associated with prenatal and birth factors, including birth weight, gestational age and maternal smoking. We quantified neonatal methylomic variation in 1263 infants using DNA isolated from a unique collection of archived blood spots taken shortly after birth (mean = 6.08 days; s.d. = 3.24 days). An epigenome-wide association study (EWAS) of gestational age and birth weight identified 4299 and 18 differentially methylated positions (DMPs) respectively, at an experiment-wide significance threshold of p < 1 × 10-7. Our EWAS of maternal smoking during pregnancy identified 110 DMPs in neonatal blood, replicating previously reported genomic loci, including AHRR. Finally, we tested the hypothesis that DNA methylation mediates the relationship between maternal smoking and lower birth weight, finding evidence that methylomic variation at three DMPs may link exposure to outcome. These findings complement an expanding literature on the epigenomic consequences of prenatal exposures and obstetric factors, confirming a link between the maternal environment and gene regulation in neonates. This article is part of the theme issue 'Developing differences: early-life effects and evolutionary medicine'.
Assuntos
Peso ao Nascer/efeitos dos fármacos , Metilação de DNA , Epigenoma/genética , Genoma Humano/genética , Efeitos Tardios da Exposição Pré-Natal/induzido quimicamente , Fumar/efeitos adversos , Feminino , Estudo de Associação Genômica Ampla , Idade Gestacional , Humanos , Recém-Nascido , GravidezRESUMO
Calsenilin modulates A-type potassium channels, regulates presenilin-mediated γ-secretase activity, and represses prodynorphin and c-fos genes expression. RhoA is involved in various cellular functions including proliferation, differentiation, migration, transcription, and regulation of the actin cytoskeleton. Although recent studies demonstrate that calsenilin can directly interact with RhoA and that RhoA inactivation is essential for neuritogenesis, it is uncertain whether there is a link between calsenilin and RhoA-regulated neuritogenesis. Here, we investigated the role of calsenilin in RhoA-regulated neuritogenesis using in vitro and in vivo systems. We found that calsenilin induced RhoA inactivation, which accompanied RhoA phosphorylation and the reduced phosphorylation levels of LIM kinase (LIMK) and cofilin. Interestingly, PC12 cells overexpressing either full-length (FL) or the caspase 3-derived C-terminal fragment (CTF) of calsenilin significantly inactivated RhoA through its interaction with RhoA and p190 Rho GTPase-activating protein (p190RhoGAP). In addition, cells expressing FL and the CTF of calsenilin had increased neurite outgrowth compared to cells expressing the N-terminal fragment (NTF) of calsenilin or vector alone. Moreover, Tat-C3 and Y27632 treatment significantly increased the percentage of neurite-bearing cells, neurite length, and the number of neurites in cells. Finally, calsenilin deficiency in the brains of calsenilin-knockout mice significantly interfered with RhoA inactivation. These findings suggest that calsenilin contributes to neuritogenesis through RhoA inactivation.
Assuntos
Proteínas Interatuantes com Canais de Kv/genética , Proteínas Interatuantes com Canais de Kv/metabolismo , Crescimento Neuronal , Proteína rhoA de Ligação ao GTP/metabolismo , Animais , Técnicas de Inativação de Genes , Células HEK293 , Humanos , Proteínas Interatuantes com Canais de Kv/química , Camundongos , Células PC12 , Fosforilação , Ratos , Transdução de SinaisRESUMO
INTRODUCTION: African Americans' (AAs) late-onset Alzheimer's disease (LOAD) genetic risk profile is incompletely understood. Including clinical covariates in genetic analyses using informed conditioning might improve study power. METHODS: We conducted a genome-wide association study (GWAS) in AAs employing informed conditioning in 1825 LOAD cases and 3784 cognitively normal controls. We derived a posterior liability conditioned on age, sex, diabetes status, current smoking status, educational attainment, and affection status, with parameters informed by external prevalence information. We assessed association between the posterior liability and a genome-wide set of single-nucleotide polymorphisms (SNPs), controlling for APOE and ABCA7, identified previously in a LOAD GWAS of AAs. RESULTS: Two SNPs at novel loci, rs112404845 (P = 3.8 × 10-8), upstream of COBL, and rs16961023 (P = 4.6 × 10-8), downstream of SLC10A2, obtained genome-wide significant evidence of association with the posterior liability. DISCUSSION: An informed conditioning approach can detect LOAD genetic associations in AAs not identified by traditional GWAS.
Assuntos
Doença de Alzheimer/etnologia , Doença de Alzheimer/genética , Negro ou Afro-Americano/genética , Loci Gênicos , Proteínas dos Microfilamentos/genética , Transportadores de Ânions Orgânicos Dependentes de Sódio/genética , Polimorfismo de Nucleotídeo Único , Simportadores/genética , Transportadores de Cassetes de Ligação de ATP/genética , Idoso , Idoso de 80 Anos ou mais , Apolipoproteínas E/genética , Complicações do Diabetes/etnologia , Complicações do Diabetes/genética , Escolaridade , Feminino , Predisposição Genética para Doença , Estudo de Associação Genômica Ampla , Humanos , Masculino , Prevalência , Fumar/etnologia , Fumar/genéticaRESUMO
OBJECTIVE: We assessed the role of prenatal maternal smoking in risk for Tourette syndrome and chronic tic disorder (TS/CT) and pediatric-onset obsessive-compulsive disorder (OCD). METHOD: In an analysis of 73,073 singleton pregnancies from the Danish National Birth Cohort, we calculated incidence rates (IR) per 1,000 person-year for TS/CT and OCD. We then determined crude and adjusted hazard ratios and 95% CIs associated with prenatal maternal smoking, considering smoking as a dichotomous (yes/no) variable or a stratified variable (no smoking, light smoking, and heavy smoking [≥10 cigarettes/day]). Additional analyses examined the effect of maternal smoking on risk for TS/CT with other comorbid psychiatric conditions. RESULTS: In final adjusted analyses, heavy smoking was associated with a 66% increased risk for TS/CT (adjusted hazard ratio = 1.66, 95% CI = 1.17-2.35). In addition, heavy smoking was associated with a 2-fold increased risk for TS/CT with comorbid attention-deficit/hyperactivity disorder (ADHD), and both light and heavy smoking were associated with a more than 2-fold increased risk for TS/CT with any non-ADHD psychiatric comorbidity. Our parallel analyses of pediatric-onset OCD were likely underpowered but showed similar relationships. CONCLUSION: Prenatal maternal smoking was associated with increased risk for TS/CT as well as TS/CT with comorbid psychiatric conditions, even after adjustment for several important variables, including maternal psychiatric history, socioeconomic status, and partner smoking. Our findings point to a pathway linking prenatal tobacco exposure and altered brain development to TS/CT.
Assuntos
Transtorno Obsessivo-Compulsivo/etiologia , Efeitos Tardios da Exposição Pré-Natal/induzido quimicamente , Fumar/efeitos adversos , Transtornos de Tique/etiologia , Adolescente , Criança , Pré-Escolar , Estudos de Coortes , Dinamarca/epidemiologia , Feminino , Humanos , Lactente , Masculino , Transtorno Obsessivo-Compulsivo/epidemiologia , Gravidez , Efeitos Tardios da Exposição Pré-Natal/epidemiologia , Fumar/epidemiologia , Transtornos de Tique/epidemiologia , Síndrome de Tourette/epidemiologia , Síndrome de Tourette/etiologiaRESUMO
Neuronal ELAV-like (nELAVL) RNA binding proteins have been linked to numerous neurological disorders. We performed crosslinking-immunoprecipitation and RNAseq on human brain, and identified nELAVL binding sites on 8681 transcripts. Using knockout mice and RNAi in human neuroblastoma cells, we showed that nELAVL intronic and 3' UTR binding regulates human RNA splicing and abundance. We validated hundreds of nELAVL targets among which were important neuronal and disease-associated transcripts, including Alzheimer's disease (AD) transcripts. We therefore investigated RNA regulation in AD brain, and observed differential splicing of 150 transcripts, which in some cases correlated with differential nELAVL binding. Unexpectedly, the most significant change of nELAVL binding was evident on non-coding Y RNAs. nELAVL/Y RNA complexes were specifically remodeled in AD and after acute UV stress in neuroblastoma cells. We propose that the increased nELAVL/Y RNA association during stress may lead to nELAVL sequestration, redistribution of nELAVL target binding, and altered neuronal RNA splicing.
Assuntos
Encéfalo/fisiologia , Proteínas ELAV/metabolismo , Regulação da Expressão Gênica , RNA Mensageiro/metabolismo , RNA não Traduzido/metabolismo , Doença de Alzheimer/patologia , Animais , Linhagem Celular Tumoral , Humanos , Camundongos Knockout , Neurônios/fisiologia , Ligação Proteica , Interferência de RNA , Splicing de RNARESUMO
Pinpointing the small number of causal variants among the abundant naturally occurring genetic variation is a difficult challenge, but a crucial one for understanding precise molecular mechanisms of disease and follow-up functional studies. We propose and investigate two complementary statistical approaches for identification of rare causal variants in sequencing studies: a backward elimination procedure based on groupwise association tests, and a hierarchical approach that can integrate sequencing data with diverse functional and evolutionary conservation annotations for individual variants. Using simulations, we show that incorporation of multiple bioinformatic predictors of deleteriousness, such as PolyPhen-2, SIFT and GERP++ scores, can improve the power to discover truly causal variants. As proof of principle, we apply the proposed methods to VPS13B, a gene mutated in the rare neurodevelopmental disorder called Cohen syndrome, and recently reported with recessive variants in autism. We identify a small set of promising candidates for causal variants, including two loss-of-function variants and a rare, homozygous probably-damaging variant that could contribute to autism risk.
Assuntos
Transtorno Autístico/genética , Evolução Molecular , Dedos/anormalidades , Variação Genética , Deficiência Intelectual/genética , Microcefalia/genética , Hipotonia Muscular/genética , Miopia/genética , Obesidade/genética , Proteínas de Transporte Vesicular/genética , Proteína 4 Semelhante a Angiopoietina , Angiopoietinas/genética , Transtorno Autístico/diagnóstico , Biologia Computacional , Simulação por Computador , Deficiências do Desenvolvimento/diagnóstico , Deficiências do Desenvolvimento/genética , Frequência do Gene , Estudo de Associação Genômica Ampla , Humanos , Deficiência Intelectual/diagnóstico , Microcefalia/diagnóstico , Modelos Genéticos , Hipotonia Muscular/diagnóstico , Miopia/diagnóstico , Obesidade/diagnóstico , Degeneração Retiniana , SoftwareRESUMO
Response to endotoxins is an important part of the organismal reaction to Gram-negative bacteria and plays a critical role in sepsis and septic shock, as well as other conditions such as metabolic endotoxemia. Humans are generally more sensitive to endotoxins when compared with experimental animals such as mice. Inflammatory caspases mediate endotoxin-induced IL-1ß secretion and lethality in mice, and caspase-4 is an inflammatory caspase that is found in the human, and not mouse, genome. To test whether caspase-4 is involved in endotoxin sensitivity, we developed a transgenic mouse expressing human caspase-4 in its genomic context. Caspase-4 transgenic mice exhibited significantly higher endotoxin sensitivity, as measured by enhanced cytokine secretion and lethality following LPS challenge. Using bone marrow-derived macrophages, we then observed that caspase-4 can support activation of caspase-1 and secretion of IL-1ß and IL-18 in response to priming signals (LPS or Pam3CSK4) alone, without the need for second signals to stimulate the assembly of the inflammasome. These findings indicate that the regulation of caspase-1 activity by human caspase-4 could represent a unique mechanism in humans, as compared with laboratory rodents, and may partially explain the higher sensitivity to endotoxins observed in humans. Regulation of the expression, activation, or activity of caspase-4 therefore represents targets for systemic inflammatory response syndrome, sepsis, septic shock, and related disorders.
Assuntos
Caspases Iniciadoras/imunologia , Caspases/imunologia , Lipopeptídeos/toxicidade , Lipopolissacarídeos/toxicidade , Macrófagos/imunologia , Animais , Caspases/genética , Caspases Iniciadoras/genética , Linhagem Celular , Indução Enzimática/efeitos dos fármacos , Indução Enzimática/genética , Indução Enzimática/imunologia , Humanos , Inflamassomos/genética , Inflamassomos/imunologia , Interleucina-18/genética , Interleucina-18/imunologia , Interleucina-1beta/genética , Interleucina-1beta/imunologia , Camundongos , Camundongos KnockoutRESUMO
This study aimed to elucidate the genetic causes underlying early-onset Parkinsonism (EOP) in a consanguineous Iranian family. To attain this, homozygosity mapping and whole-exome sequencing were performed. As a result, a homozygous mutation (c.773G>A; p.Arg258Gln) lying within the NH2 -terminal Sac1-like inositol phosphatase domain of polyphosphoinositide phosphatase synaptojanin 1 (SYNJ1), which has been implicated in the regulation of endocytic traffic at synapses, was identified as the disease-segregating mutation. This mutation impaired the phosphatase activity of SYNJ1 against its Sac1 domain substrates in vitro. We concluded that the SYNJ1 mutation identified here is responsible for the EOP phenotype seen in our patients probably due to deficiencies in its phosphatase activity and consequent impairment of its synaptic functions. Our finding not only opens new avenues of investigation in the synaptic dysfunction mechanisms associated with Parkinsonism, but also suggests phosphoinositide metabolism as a novel therapeutic target for Parkinsonism.
Assuntos
Epilepsia Generalizada/genética , Epilepsia Generalizada/metabolismo , Transtornos Parkinsonianos/genética , Transtornos Parkinsonianos/metabolismo , Monoéster Fosfórico Hidrolases/genética , Monoéster Fosfórico Hidrolases/metabolismo , Adulto , Idade de Início , Encéfalo/metabolismo , Consanguinidade , Exoma , Feminino , Genes Recessivos , Genótipo , Células HEK293 , Humanos , Masculino , Monoéster Fosfórico Hidrolases/química , Filogenia , Polimorfismo de Nucleotídeo Único , Alinhamento de Sequência , Análise de Sequência de DNARESUMO
Recent developments in sequencing technologies have made it possible to uncover both rare and common genetic variants. Genome-wide association studies (GWASs) can test for the effect of common variants, whereas sequence-based association studies can evaluate the cumulative effect of both rare and common variants on disease risk. Many groupwise association tests, including burden tests and variance-component tests, have been proposed for this purpose. Although such tests do not exclude common variants from their evaluation, they focus mostly on testing the effect of rare variants by upweighting rare-variant effects and downweighting common-variant effects and can therefore lose substantial power when both rare and common genetic variants in a region influence trait susceptibility. There is increasing evidence that the allelic spectrum of risk variants at a given locus might include novel, rare, low-frequency, and common genetic variants. Here, we introduce several sequence kernel association tests to evaluate the cumulative effect of rare and common variants. The proposed tests are computationally efficient and are applicable to both binary and continuous traits. Furthermore, they can readily combine GWAS and whole-exome-sequencing data on the same individuals, when available, and are also applicable to deep-resequencing data of GWAS loci. We evaluate these tests on data simulated under comprehensive scenarios and show that compared with the most commonly used tests, including the burden and variance-component tests, they can achieve substantial increases in power. We next show applications to sequencing studies for Crohn disease and autism spectrum disorders. The proposed tests have been incorporated into the software package SKAT.
Assuntos
Estudos de Associação Genética/métodos , Software , Algoritmos , Transtornos Globais do Desenvolvimento Infantil/genética , Simulação por Computador , Doença de Crohn/genética , Interpretação Estatística de Dados , Frequência do Gene , Predisposição Genética para Doença , Testes Genéticos , Humanos , Modelos Logísticos , Proteína-2 Relacionada a Receptor de Lipoproteína de Baixa Densidade/genética , Modelos Genéticos , Proteína Adaptadora de Sinalização NOD2/genética , RiscoRESUMO
In this study, a consanguineous family with progressive myoclonus epilepsy (PME) was clinically examined and molecularly investigated to determine the molecular events causing disease. Since exclusion of known genes indicated that novel genes causing PME still remained unidentified, homozygosity mapping, exome sequencing, as well as validation and disease-segregation analyses were subsequently carried out for both loci and gene identification. To further assure our results, a muscle biopsy and gene expression analyses were additionally performed. As a result, a homozygous, disease-segregating COL6A2 mutation, p.Asp215Asn, absent in a large number of control individuals, including control individuals of Iranian ancestry, was identified in both affected siblings. COL6A2 was shown to be expressed in the human cerebral cortex and muscle biopsy revealed no specific histochemical pathology. We conclude that the COL6A2 p.Asp215Asn mutation is likely to be responsible for PME in this family; however, additional studies are warranted to further establish the pathogenic role of both COL6A2 and the extracellular proteolysis system in the pathogenesis of PME.
Assuntos
Córtex Cerebral/metabolismo , Colágeno Tipo VI/genética , Mutação , Epilepsias Mioclônicas Progressivas/genética , Adolescente , Adulto , Asparagina , Ácido Aspártico , Cromossomos Humanos Par 21/genética , Colágeno Tipo VI/metabolismo , Consanguinidade , Eletroencefalografia , Feminino , Homozigoto , Humanos , Irã (Geográfico) , Masculino , Pessoa de Meia-Idade , Epilepsias Mioclônicas Progressivas/fisiopatologia , Linhagem , SíndromeRESUMO
Presenilin 1 (PS1) is a component of the γ-secretase complex that cleaves a variety of type I membrane proteins, including the ß-amyloid precursor protein (ß-APP), Notch, and neuronal (N)- and epithelial (E)-cadherins. N-cadherin is an essential adhesion molecule that forms a complex with, and is cleaved by, PS1/γ-secretase and ß-catenin in the plasma membrane. The purpose of this study was to determine whether calsenilin, a presenilin-interacting protein, has a functional role in PS1/γ-secretase-mediated N-cadherin ε-cleavage using Western blot analysis, RT-PCR, immunoprecipitation, subcellular fractionation, biotinylation, and a luciferase reporter assay in SH-SY5Y neuroblastoma cells. Here, we demonstrate that the expression of calsenilin leads to a disruption of PS1/γ-secretase-mediated ε-cleavage of N-cadherin, which results in the significant accumulation of N-cadherin C-terminal fragment 1 (Ncad/CTF1), the reduction of cytoplasmic Ncad/CTF2 release, and a deceleration of PS1-CTF delivery to the cell surface. Interestingly, we also found that the expression of calsenilin is associated with the redistribution of ß-catenin from the cell surface to a cytoplasmic pool, as well as with the negative regulation of genes that are targets of T-cell factor/ß-catenin nuclear signaling. Taken together, our findings suggest that calsenilin is a novel negative regulator of N-cadherin processing that plays an important role in ß-catenin signaling.
Assuntos
Secretases da Proteína Precursora do Amiloide/metabolismo , Antígenos CD/metabolismo , Caderinas/metabolismo , Proteínas Interatuantes com Canais de Kv/metabolismo , Presenilina-1/metabolismo , Proteínas Repressoras/metabolismo , beta Catenina/metabolismo , Transporte Ativo do Núcleo Celular , Antígenos CD/química , Caderinas/química , Linhagem Celular Tumoral , Membrana Celular/metabolismo , Quinase 3 da Glicogênio Sintase/metabolismo , Glicogênio Sintase Quinase 3 beta , Humanos , Proteínas Interatuantes com Canais de Kv/genética , Modelos Neurológicos , Neuroblastoma/genética , Neuroblastoma/metabolismo , Estrutura Terciária de Proteína , Proteólise , Proteínas Repressoras/genética , Transdução de Sinais , beta Catenina/genéticaRESUMO
Rapid advances in sequencing technologies set the stage for the large-scale medical sequencing efforts to be performed in the near future, with the goal of assessing the importance of rare variants in complex diseases. The discovery of new disease susceptibility genes requires powerful statistical methods for rare variant analysis. The low frequency and the expected large number of such variants pose great difficulties for the analysis of these data. We propose here a robust and powerful testing strategy to study the role rare variants may play in affecting susceptibility to complex traits. The strategy is based on assessing whether rare variants in a genetic region collectively occur at significantly higher frequencies in cases compared with controls (or vice versa). A main feature of the proposed methodology is that, although it is an overall test assessing a possibly large number of rare variants simultaneously, the disease variants can be both protective and risk variants, with moderate decreases in statistical power when both types of variants are present. Using simulations, we show that this approach can be powerful under complex and general disease models, as well as in larger genetic regions where the proportion of disease susceptibility variants may be small. Comparisons with previously published tests on simulated data show that the proposed approach can have better power than the existing methods. An application to a recently published study on Type-1 Diabetes finds rare variants in gene IFIH1 to be protective against Type-1 Diabetes.
Assuntos
Simulação por Computador , Predisposição Genética para Doença , Testes Genéticos/estatística & dados numéricos , Estudo de Associação Genômica Ampla/estatística & dados numéricos , Algoritmos , RNA Helicases DEAD-box/genética , Interpretação Estatística de Dados , Diabetes Mellitus Tipo 1/genética , Variação Genética , Haplótipos/genética , Humanos , Helicase IFIH1 Induzida por Interferon , Fatores de Risco , Análise de Sequência de DNARESUMO
Calsenilin, a neuronal calcium binding protein that has been shown to have multiple functions in the cell, interacts with presenilin 1 (PS1) and presenilin 2 (PS2), represses gene transcription and binds to A-type voltage-gated potassium channels. In addition, increased levels of calsenilin are observed in the brains of Alzheimer's disease and epilepsy patients. The present study was designed to investigate the molecular mechanism of calsenilin degradation pathways in cultured cells. Here, we demonstrate that inhibition of the ubiquitin-proteasomal pathway (UPP) but not lysosomal pathway markedly increased the expression levels of calsenilin. Immunofluorescence analysis revealed that following proteasomal inhibition calsenilin accumulated in the endoplasmic reticulum (ER) and Golgi, while lysosomal inhibition had no effect on calsenilin localization. In addition, we found the change of subcellular localization of PS1 from diffuse pattern to punctuate staining pattern in the ER and perinuclear region in the presence of calsenilin. These findings suggest that calsenilin degradation is primarily mediated by the UPP and that impairment in the UPP may contribute to the involvement of calsenilin in disease-associated neurodegeneration.
Assuntos
Proteínas Interatuantes com Canais de Kv/metabolismo , Complexo de Endopeptidases do Proteassoma/metabolismo , Proteínas Repressoras/metabolismo , Ubiquitina/metabolismo , Ubiquitinação , Linhagem Celular Tumoral , Inibidores de Cisteína Proteinase/farmacologia , Retículo Endoplasmático/metabolismo , Complexo de Golgi/metabolismo , Humanos , Leupeptinas/farmacologia , Lisossomos/efeitos dos fármacos , Lisossomos/metabolismo , Doenças Neurodegenerativas/metabolismo , Inibidores de ProteassomaRESUMO
BACKGROUND: SLC25A12, a susceptibility gene for autism spectrum disorders that is mutated in a neurodevelopmental syndrome, encodes a mitochondrial aspartate-glutamate carrier (aspartate-glutamate carrier isoform 1 [AGC1]). AGC1 is an important component of the malate/aspartate shuttle, a crucial system supporting oxidative phosphorylation and adenosine triphosphate production. METHODS: We characterized mice with a disruption of the Slc25a12 gene, followed by confirmatory in vitro studies. RESULTS: Slc25a12-knockout mice, which showed no AGC1 by immunoblotting, were born normally but displayed delayed development and died around 3 weeks after birth. In postnatal day 13 to 14 knockout brains, the brains were smaller with no obvious alteration in gross structure. However, we found a reduction in myelin basic protein (MBP)-positive fibers, consistent with a previous report. Furthermore, the neocortex of knockout mice contained abnormal neurofilamentous accumulations in neurons, suggesting defective axonal transport and/or neurodegeneration. Slice cultures prepared from knockout mice also showed a myelination defect, and reduction of Slc25a12 in rat primary oligodendrocytes led to a cell-autonomous reduction in MBP expression. Myelin deficits in slice cultures from knockout mice could be reversed by administration of pyruvate, indicating that reduction in AGC1 activity leads to reduced production of aspartate/N-acetylaspartate and/or alterations in the dihydronicotinamide adenine dinucleotide/nicotinamide adenine dinucleotide(+) ratio, resulting in myelin defects. CONCLUSIONS: Our data implicate AGC1 activity in myelination and in neuronal structure and indicate that while loss of AGC1 leads to hypomyelination and neuronal changes, subtle alterations in AGC1 expression could affect brain development, contributing to increased autism susceptibility.
Assuntos
Deficiências do Desenvolvimento/metabolismo , Encefalite/metabolismo , Proteínas de Membrana Transportadoras/metabolismo , Proteínas Mitocondriais/metabolismo , Proteína Básica da Mielina/metabolismo , Agrecanas/metabolismo , Animais , Animais Recém-Nascidos , Encéfalo/citologia , Calbindinas , Células Cultivadas , Cerebelo/citologia , Deficiências do Desenvolvimento/genética , Modelos Animais de Doenças , Embrião de Mamíferos , Encefalite/genética , Encefalite/patologia , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Regulação da Expressão Gênica no Desenvolvimento/genética , Proteínas de Fluorescência Verde/genética , Masculino , Proteínas de Membrana Transportadoras/deficiência , Camundongos , Camundongos Knockout , Proteínas de Transporte da Membrana Mitocondrial , Proteínas Mitocondriais/deficiência , Glicoproteína Associada a Mielina , Neurônios/metabolismo , Neurônios/patologia , Oligodendroglia/efeitos dos fármacos , Oligodendroglia/fisiologia , Técnicas de Cultura de Órgãos , Ácido Pirúvico/uso terapêutico , Receptores de Superfície Celular/genética , Receptores de Superfície Celular/metabolismo , Proteína G de Ligação ao Cálcio S100/metabolismo , Células-Tronco/fisiologiaRESUMO
OBJECTIVE: Alzheimer's disease (AD) is a progressive neurodegenerative disease of the central nervous system (CNS). Recently, an increased interest in the role diet plays in the pathology of AD has resulted in a focus on the detrimental effects of diets high in cholesterol and fat and the beneficial effects of caloric restriction. The current study examines how dietary composition modulates cerebral amyloidosis and neuronal integrity in the TgCRND8 mouse model of AD. METHODS: From 4 wks until 18 wks of age, male and female TgCRND8 mice were maintained on one of four diets: (1) reference (regular) commercial chow; (2) high fat/low carbohydrate custom chow (60 kcal% fat/30 kcal% protein/10 kcal% carbohydrate); (3) high protein/low carbohydrate custom chow (60 kcal% protein/30 kcal% fat/10 kcal% carbohydrate); or (4) high carbohydrate/low fat custom chow (60 kcal% carbohydrate/30 kcal% protein/10 kcal% fat). At age 18 wks, mice were sacrificed, and brains studied for (a) wet weight; (b) solubilizable Abeta content by ELISA; (c) amyloid plaque burden; (d) stereologic analysis of selected hippocampal subregions. RESULTS: Animals receiving a high fat diet showed increased brain levels of solubilizable Abeta, although we detected no effect on plaque burden. Unexpectedly, brains of mice fed a high protein/low carbohydrate diet were 5% lower in weight than brains from all other mice. In an effort to identify regions that might link loss of brain mass to cognitive function, we studied neuronal density and volume in hippocampal subregions. Neuronal density and volume in the hippocampal CA3 region of TgCRND8 mice tended to be lower in TgCRND8 mice receiving the high protein/low carbohydrate diet than in those receiving the regular chow. Neuronal density and volume were preserved in CA1 and in the dentate gyrus. INTERPRETATION: Dissociation of Abeta changes from brain mass changes raises the possibility that diet plays a role not only in modulating amyloidosis but also in modulating neuronal vulnerability. However, in the absence of a study of the effects of a high protein/low carbohydrate diet on nontransgenic mice, one cannot be certain how much, if any, of the loss of brain mass exhibited by high protein/low carbohydrate diet-fed TgCRND8 mice was due to an interaction between cerebral amyloidosis and diet. Given the recent evidence that certain factors favor the maintenance of cognitive function in the face of substantial structural neuropathology, we propose that there might also exist factors that sensitize brain neurons to some forms of neurotoxicity, including, perhaps, amyloid neurotoxicity. Identification of these factors could help reconcile the poor clinicopathological correlation between cognitive status and structural neuropathology, including amyloid pathology.