RESUMO
Prion diseases (transmissible spongiform encephalopathies) are fatal neurodegenerative diseases, including Creutzfeldt-Jakob disease in humans, scrapie in sheep and bovine spongiform encephalopathy in cattle. While genome-wide association studies in human and quantitative trait loci mapping in mice have provided evidence for multiple susceptibility genes, few of these have been confirmed functionally. Phenotyping mouse models is generally the method of choice. However, this is not a feasible option where many novel genes, without pre-existing models, would need to be tested. We have therefore developed and applied an in-vitro screen to triage and prioritize candidate modifier genes for more detailed future studies which is faster, far more cost effective and ethical relative to mouse bioassay models. An in vitro prion bioassay, the scrapie cell assay, uses a neuroblastoma-derived cell line (PK1) that is susceptible to RML prions and able to propagate prions at high levels. In this study, we have generated stable gene silencing and/or overexpressing PK1-derived cell lines to test whether perturbation of 14 candidate genes affects prion susceptibility. While no consistent differences were determined for seven genes, highly significant changes were detected for Zbtb38, Sorcs1, Stmn2, Hspa13, Fkbp9, Actr10 and Plg, suggesting that they play key roles in the fundamental processes of prion propagation or clearance. Many neurodegenerative diseases involve the accumulation of misfolded protein aggregates and 'prion-like' seeding and spread has been implicated in their pathogenesis. It is therefore expected that some of these prion-modifier genes may be of wider relevance in neurodegeneration.
Assuntos
Predisposição Genética para Doença , Doenças Priônicas/genética , Animais , Linhagem Celular , Expressão Gênica , Técnicas de Inativação de Genes , Estudo de Associação Genômica Ampla , Humanos , Técnicas In Vitro , Camundongos , Locos de Características Quantitativas , Interferência de RNA , ScrapieRESUMO
Hexanucleotide repeat expansions in C9orf72 are a major cause of frontotemporal lobar degeneration (FTLD) and amyotrophic lateral sclerosis (ALS). Understanding the disease mechanisms and a method for clinical diagnostic genotyping have been hindered because of the difficulty in estimating the expansion size. We found 96 repeat-primed PCR expansions: 85/2,974 in six neurodegenerative diseases cohorts (FTLD, ALS, Alzheimer disease, sporadic Creutzfeldt-Jakob disease, Huntington disease-like syndrome, and other nonspecific neurodegenerative disease syndromes) and 11/7,579 (0.15%) in UK 1958 birth cohort (58BC) controls. With the use of a modified Southern blot method, the estimated expansion range (smear maxima) in cases was 800-4,400. Similarly, large expansions were detected in the population controls. Differences in expansion size and morphology were detected between DNA samples from tissue and cell lines. Of those in whom repeat-primed PCR detected expansions, 68/69 were confirmed by blotting, which was specific for greater than 275 repeats. We found that morphology in the expansion smear varied among different individuals and among different brain regions in the same individual. Expansion size correlated with age at clinical onset but did not differ between diagnostic groups. Evidence of instability of repeat size in control families, as well as neighboring SNP and microsatellite analyses, support multiple expansion events on the same haplotype background. Our method of estimating the size of large expansions has potential clinical utility. C9orf72-related disease might mimic several neurodegenerative disorders and, with potentially 90,000 carriers in the United Kingdom, is more common than previously realized.
Assuntos
Esclerose Lateral Amiotrófica/genética , Expansão das Repetições de DNA , Degeneração Lobar Frontotemporal/genética , Degeneração Neural/genética , Proteínas/genética , Esclerose Lateral Amiotrófica/patologia , Encéfalo/patologia , Proteína C9orf72 , Estudos de Coortes , DNA/genética , Degeneração Lobar Frontotemporal/patologia , Predisposição Genética para Doença , Haplótipos , Humanos , Repetições de Microssatélites , Degeneração Neural/patologia , Polimorfismo de Nucleotídeo Único , Reino UnidoRESUMO
Prion diseases are fatal neurodegenerative disorders that include bovine spongiform encephalopathy (BSE) and scrapie in animals and Creutzfeldt-Jakob disease (CJD) in humans. They are characterized by long incubation periods, variation in which is determined by many factors including genetic background. In some cases it is possible that incubation time may be directly correlated to the level of gene expression. To test this hypothesis, we combined incubation time data from five different inbred lines of mice with quantitative gene expression profiling in normal brains and identified five genes with expression levels that correlate with incubation time. One of these genes, Hspa13 (Stch), is a member of the Hsp70 family of ATPase heat shock proteins, which have been previously implicated in prion propagation. To test whether Hspa13 plays a causal role in determining the incubation period, we tested two overexpressing mouse models. The Tc1 human chromosome 21 (Hsa21) transchromosomic mouse model of Down syndrome is trisomic for many Hsa21 genes including Hspa13 and following Chandler/Rocky Mountain Laboratory (RML) prion inoculation, shows a 4% reduction in incubation time. Furthermore, a transgenic model with eightfold overexpression of mouse Hspa13 exhibited highly significant reductions in incubation time of 16, 15, and 7% following infection with Chandler/RML, ME7, and MRC2 prion strains, respectively. These data further implicate Hsp70-like molecular chaperones in protein misfolding disorders such as prion disease.
Assuntos
Regulação da Expressão Gênica , Proteínas de Choque Térmico HSP70/biossíntese , Proteínas de Choque Térmico HSP70/fisiologia , Doenças Priônicas/genética , Adenosina Trifosfatases/química , Animais , Proteínas de Choque Térmico HSP70/genética , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos CBA , Camundongos Transgênicos , Modelos Genéticos , Neurônios/metabolismo , Análise de Sequência com Séries de Oligonucleotídeos , Fenótipo , Príons/metabolismo , RNA Complementar/metabolismoRESUMO
Prion diseases are fatal neurodegenerative diseases of humans and animals caused by the misfolding and aggregation of prion protein (PrP). Mammalian prion diseases are under strong genetic control but few risk factors are known aside from the PrP gene locus (PRNP). No genome-wide association study (GWAS) has been done aside from a small sample of variant Creutzfeldt-Jakob disease (CJD). We conducted GWAS of sporadic CJD (sCJD), variant CJD (vCJD), iatrogenic CJD, inherited prion disease, kuru and resistance to kuru despite attendance at mortuary feasts. After quality control, we analysed 2000 samples and 6015 control individuals (provided by the Wellcome Trust Case Control Consortium and KORA-gen) for 491032-511862 SNPs in the European study. Association studies were done in each geographical and aetiological group followed by several combined analyses. The PRNP locus was highly associated with risk in all geographical and aetiological groups. This association was driven by the known coding variation at rs1799990 (PRNP codon 129). No non-PRNP loci achieved genome-wide significance in the meta-analysis of all human prion disease. SNPs at the ZBTB38-RASA2 locus were associated with CJD in the UK (rs295301, P = 3.13 × 10(-8); OR, 0.70) but these SNPs showed no replication evidence of association in German sCJD or in Papua New Guinea-based tests. A SNP in the CHN2 gene was associated with vCJD [P = 1.5 × 10(-7); odds ratio (OR), 2.36], but not in UK sCJD (P = 0.049; OR, 1.24), in German sCJD or in PNG groups. In the overall meta-analysis of CJD, 14 SNPs were associated (P < 10(-5); two at PRNP, three at ZBTB38-RASA2, nine at nine other independent non-PRNP loci), more than would be expected by chance. None of the loci recently identified as genome-wide significant in studies of other neurodegenerative diseases showed any clear evidence of association in prion diseases. Concerning common genetic variation, it is likely that the PRNP locus contains the only strong risk factors that act universally across human prion diseases. Our data are most consistent with several other risk loci of modest overall effects which will require further genetic association studies to provide definitive evidence.
Assuntos
Predisposição Genética para Doença , Estudo de Associação Genômica Ampla , Polimorfismo de Nucleotídeo Único , Doenças Priônicas/genética , Príons/genética , Estudos de Casos e Controles , Síndrome de Creutzfeldt-Jakob/genética , Resistência à Doença , Encefalopatia Espongiforme Bovina/genética , Feminino , Humanos , Kuru/genética , Proteínas de Neoplasias/genética , Proteínas Priônicas , Fatores de Risco , Proteínas Ativadoras de ras GTPase/genéticaRESUMO
An expanded hexanucleotide repeat in the C9orf72 gene is the most common genetic cause of frontotemporal dementia and amyotrophic lateral sclerosis (c9FTD/ALS). We now report the first description of a homozygous patient and compare it to a series of heterozygous cases. The patient developed early-onset frontotemporal dementia without additional features. Neuropathological analysis showed c9FTD/ALS characteristics, with abundant p62-positive inclusions in the frontal and temporal cortices, hippocampus and cerebellum, as well as less abundant TDP-43-positive inclusions. Overall, the clinical and pathological features were severe, but did not fall outside the usual disease spectrum. Quantification of C9orf72 transcript levels in post-mortem brain demonstrated expression of all known C9orf72 transcript variants, but at a reduced level. The pathogenic mechanisms by which the hexanucleotide repeat expansion causes disease are unclear and both gain- and loss-of-function mechanisms may play a role. Our data support a gain-of-function mechanism as pure homozygous loss of function would be expected to lead to a more severe, or completely different clinical phenotype to the one described here, which falls within the usual range. Our findings have implications for genetic counselling, highlighting the need to use genetic tests that distinguish C9orf72 homozygosity.
Assuntos
Esclerose Lateral Amiotrófica/genética , Encéfalo/patologia , Expansão das Repetições de DNA/genética , Demência Frontotemporal/genética , Mutação/genética , Proteínas/genética , Idade de Início , Esclerose Lateral Amiotrófica/patologia , Proteína C9orf72 , Proteínas de Ligação a DNA/metabolismo , Demência Frontotemporal/patologia , Homozigoto , Humanos , Masculino , Pessoa de Meia-Idade , Testes Neuropsicológicos , Linhagem , Proteínas/metabolismoRESUMO
BACKGROUND: Kuru is a devastating epidemic prion disease that affected a highly restricted geographic area of the Papua New Guinea highlands; at its peak, it predominantly affected adult women and children of both sexes. Its incidence has steadily declined since the cessation of its route of transmission, endocannibalism. METHODS: We performed genetic and selected clinical and genealogic assessments of more than 3000 persons from Eastern Highland populations, including 709 who participated in cannibalistic mortuary feasts, 152 of whom subsequently died of kuru. RESULTS: Persons who were exposed to kuru and survived the epidemic in Papua New Guinea are predominantly heterozygotes at the known resistance factor at codon 129 of the prion protein gene (PRNP). We now report a novel PRNP variant--G127V--that was found exclusively in people who lived in the region in which kuru was prevalent and that was present in half of the otherwise susceptible women from the region of highest exposure who were homozygous for methionine at PRNP codon 129. Although this allele is common in the area with the highest incidence of kuru, it is not found in patients with kuru and in unexposed population groups worldwide. Genealogic analysis reveals a significantly lower incidence of kuru in pedigrees that harbor the protective allele than in geographically matched control families. CONCLUSIONS: The 127V polymorphism is an acquired prion disease resistance factor selected during the kuru epidemic, rather than a pathogenic mutation that could have triggered the kuru epidemic. Variants at codons 127 and 129 of PRNP demonstrate the population genetic response to an epidemic of prion disease and represent a powerful episode of recent selection in humans.
Assuntos
Predisposição Genética para Doença , Kuru/genética , Polimorfismo Genético , Príons/genética , Adolescente , Adulto , Idoso , Canibalismo , Surtos de Doenças , Feminino , Frequência do Gene , Aptidão Genética , Genótipo , Haplótipos , Humanos , Kuru/epidemiologia , Masculino , Pessoa de Meia-Idade , Papua Nova Guiné/epidemiologia , Proteínas Priônicas , Adulto JovemRESUMO
Genetic factors are implicated in the aetiology of sporadic late-onset neurodegenerative diseases. Whether these genetic variants are predominantly common or rare, and how multiple genetic factors interact with each other to cause disease is poorly understood. Inherited prion diseases are highly heterogeneous and may be clinically mistaken for sporadic Creutzfeldt-Jakob disease because of a negative family history. Here we report our investigation of patients from the UK with four extra octapeptide repeats, which suggest that the risk of clinical disease is increased by a combination of the mutation and a susceptibility haplotype on the wild-type chromosome. The predominant clinical syndrome is a progressive cortical dementia with pyramidal signs, myoclonus and cerebellar abnormalities that closely resemble sporadic Creutzfeldt-Jakob disease. Autopsy shows perpendicular deposits of prion protein in the molecular layer of the cerebellum. Identity testing, PRNP microsatellite haplotyping and genealogical work confirm no cryptic close family relationships and suggests multiple progenitor disease haplotypes. All patients were homozygous for methionine at polymorphic codon 129. In addition, at a single nucleotide polymorphism upstream of PRNP thought to confer susceptibility to sporadic Creutzfeldt-Jakob disease (rs1029273), all patients were homozygous for the risk allele (combined P=5.9×10(-5)). The haplotype identified may also be a risk factor in other partially penetrant inherited prion diseases although it does not modify age of onset. Blood expression of PRNP in healthy individuals was modestly higher in carriers of the risk haplotype. These findings may provide a precedent for understanding apparently sporadic neurodegenerative diseases caused by rare high-risk mutations.
Assuntos
Saúde da Família , Predisposição Genética para Doença , Mutagênese Insercional , Oligopeptídeos/genética , Doenças Priônicas/genética , Príons/genética , Idoso , Idoso de 80 Anos ou mais , Córtex Cerebral/metabolismo , Córtex Cerebral/patologia , Distribuição de Qui-Quadrado , Transtornos Cognitivos/etiologia , Transtornos Cognitivos/genética , Eletroencefalografia , Feminino , Testes Genéticos , Haplótipos , Humanos , Masculino , Pessoa de Meia-Idade , Doenças Priônicas/complicações , Doenças Priônicas/diagnóstico por imagem , Príons/metabolismo , Tomografia Computadorizada por Raios X/métodosRESUMO
Mutation of the human prion protein gene (PRNP) open reading frame (ORF) accounts for almost all reported familial concurrence of prion disease. The more common mutations globally: octapeptide repeat insertions, P102L, D178N, E200K, and V210I have occurred in large multigenerational pedigrees and display autosomal dominant inheritance, however, many rare genetic changes have been reported that are of uncertain pathogenicity. Based on 19 years of PRNP sequencing at the MRC Prion Unit, London, and analysis of 3664 samples from patients referred with suspected prion disease and healthy populations, we present novel allele combinations, healthy control population data, results of screening the PRNP ORF in DNA from the entire referral series and the CEPH human genome diversity cell line panel. Of the 10 alleles detected in patients for which detailed cases histories are presented, 4 are unreported (G54S, D167N, V209M, Q212PP), two changes are thought to be pathogenic but have not been described in our regions (P105L from the UK, G114V from India and Turkey), and the remainder reported in healthy control populations or in trans to known pathogenic mutations suggesting non- or low pathogenicity (G54S, 1-OPRI, G142S, N171S, V209M, E219K). New genotype-phenotype correlations and population frequencies presented will help the diagnosis and genetic counselling of those with suspected inherited prion disease.
Assuntos
Alelos , Mutação de Sentido Incorreto , Doenças Priônicas/genética , Príons/genética , Análise Mutacional de DNA/métodos , Frequência do Gene , Estudos de Associação Genética , Genótipo , Humanos , Londres , Doenças Priônicas/diagnósticoRESUMO
BACKGROUND: Human and animal prion diseases are under genetic control, but apart from PRNP (the gene that encodes the prion protein), we understand little about human susceptibility to bovine spongiform encephalopathy (BSE) prions, the causal agent of variant Creutzfeldt-Jakob disease (vCJD). METHODS: We did a genome-wide association study of the risk of vCJD and tested for replication of our findings in samples from many categories of human prion disease (929 samples) and control samples from the UK and Papua New Guinea (4254 samples), including controls in the UK who were genotyped by the Wellcome Trust Case Control Consortium. We also did follow-up analyses of the genetic control of the clinical phenotype of prion disease and analysed candidate gene expression in a mouse cellular model of prion infection. FINDINGS: The PRNP locus was strongly associated with risk across several markers and all categories of prion disease (best single SNP [single nucleotide polymorphism] association in vCJD p=2.5 x 10(-17); best haplotypic association in vCJD p=1 x 10(-24)). Although the main contribution to disease risk was conferred by PRNP polymorphic codon 129, another nearby SNP conferred increased risk of vCJD. In addition to PRNP, one technically validated SNP association upstream of RARB (the gene that encodes retinoic acid receptor beta) had nominal genome-wide significance (p=1.9 x 10(-7)). A similar association was found in a small sample of patients with iatrogenic CJD (p=0.030) but not in patients with sporadic CJD (sCJD) or kuru. In cultured cells, retinoic acid regulates the expression of the prion protein. We found an association with acquired prion disease, including vCJD (p=5.6 x 10(-5)), kuru incubation time (p=0.017), and resistance to kuru (p=2.5 x 10(-4)), in a region upstream of STMN2 (the gene that encodes SCG10). The risk genotype was not associated with sCJD but conferred an earlier age of onset. Furthermore, expression of Stmn2 was reduced 30-fold post-infection in a mouse cellular model of prion disease. INTERPRETATION: The polymorphic codon 129 of PRNP was the main genetic risk factor for vCJD; however, additional candidate loci have been identified, which justifies functional analyses of these biological pathways in prion disease.
Assuntos
Síndrome de Creutzfeldt-Jakob/epidemiologia , Síndrome de Creutzfeldt-Jakob/genética , Adulto , Idade de Início , Idoso , Alelos , Cromossomos Humanos/genética , DNA/genética , Interpretação Estatística de Dados , Feminino , Estudo de Associação Genômica Ampla , Genótipo , Humanos , Kuru/epidemiologia , Desequilíbrio de Ligação/genética , Masculino , Proteínas de Membrana/genética , Pessoa de Meia-Idade , Papua Nova Guiné/epidemiologia , Polimorfismo de Nucleotídeo Único , Vigilância da População , Proteínas Priônicas , Príons/genética , Controle de Qualidade , Fatores de Risco , Estatmina , Reino Unido/epidemiologiaRESUMO
Highly accurate and high-throughput SNP genotyping platforms are increasingly popular but the performance of suboptimal DNA samples remains unclear. The aim of our study was to determine the best platform, amplification technique, and loading concentration to maximize genotype accuracy and call rate using degraded samples. We amplified high-molecular weight genomic DNA samples recently extracted from whole blood and degraded DNA samples extracted from 50-year-old patient sera. Two whole-genome amplification (WGA) methodologies were used: an isothermal multiple displacement amplification method (MDA) and a fragmentation-PCR-based method (GenomePlex [GPLEX]; Sigma-Aldrich, St. Louis, MO). Duplicate runs were performed on genome-wide dense SNP arrays (Nsp-Mendel; Affymetrix) and custom SNP platforms based on molecular inversion probes (Targeted Genotyping [TG]; Affymetrix) and BeadArray technology (Golden Gate [GG]; Illumina). Miscalls and no-calls on Mendel arrays were correlated with each other, with confidence scores from the Bayesian calling algorithm, and with average probe intensity. Degraded DNA amplified with MDA gave low call rates and concordance across all platforms at standard loading concentrations. The call rate with MDA on GG was improved when a 5 x concentration of amplified DNA was used. The GPLEX amplification gave high call rate and concordance for degraded DNA at standard and higher loading concentrations on both TG and GG platforms. Based on these analyses, after standard filtering for SNP and sample performance, we were able to achieve a mean call rate of 99.7% and concordance 99.7% using degraded samples amplified by GPLEX on GG technology at 2 x loading concentration. These findings may be useful for investigators planning case-control association studies with patient samples of suboptimal quality.
Assuntos
DNA/metabolismo , Técnicas Genéticas , Técnicas de Amplificação de Ácido Nucleico/métodos , Polimorfismo de Nucleotídeo Único , Genoma Humano , Humanos , Kuru/genética , Pessoa de Meia-IdadeRESUMO
Inherited prion diseases are neurodegenerative disorders caused by autosomal dominant mutations in the human prion protein gene (PRNP). Kindred with inherited prion disease can show remarkable phenotypic variability that has yet to be explained. Here we report analysis of protease resistant disease-related prion protein (PrP(Sc)) isoforms from a range of inherited prion disease cases (point mutations P102L, D178N, E200K and 2-, 4- and 6-octapeptide repeat insertions) and show that the glycoform ratios of PrP(Sc) associated with PRNP point mutations are distinct from those observed in sporadic, iatrogenic and variant Creutzfeldt-Jakob disease. Patients with the same PRNP mutation can also propagate PrP(Sc) with distinct conformations. These data extend the spectrum of recognized PrP(Sc) types seen in human prion diseases and provide further insight into the generation of diverse clinicopathological phenotypes associated with inherited prion disease.
Assuntos
Mutação Puntual , Doenças Priônicas/genética , Príons/genética , Síndrome de Creutzfeldt-Jakob/genética , Síndrome de Creutzfeldt-Jakob/metabolismo , Endopeptidase K , Glicosilação , Humanos , Immunoblotting/métodos , Oligopeptídeos/genética , Tonsila Palatina/química , Fenótipo , Proteínas PrPSc/análise , Proteínas PrPSc/metabolismo , Doenças Priônicas/metabolismo , Baço/químicaRESUMO
By far the largest known kindred with an inherited prion disease caused by a prion protein (PrP) octapeptide repeat insertion mutation originates from southeast England. This extended family shows very marked phenotypic heterogeneity and provides a unique opportunity to characterize this diversity and examine possible modifying factors amongst a large number of individuals in whom prion disease has been initiated by the same defined genetic mutation. As the inherited prion diseases comprise a significant proportion of familial early-onset dementia, an appreciation of their wide range of clinical presentation is important for differential diagnosis. Genealogical and clinical record review, together with the characterization of the mutation-linked single nucleotide polymorphism and microsatellite haplotype, suggested a single founder for both this large kindred and a smaller family in the mid-18th century. Here we report the phenotype of 86 affected individuals; at least another 84 individuals are known to be at risk of inheriting the disease. Clinical onset, typically with cognitive impairment, can be strikingly early in this kindred when compared with other inherited or sporadic prion diseases. We have investigated the effect of PrP genotype, candidate genes and prion strain type on clinical, neuroradiological and neuropathological phenotype. The transmission characteristics of prions from affected individuals resembled those of classical sporadic Creutzfeldt-Jakob disease. One surprising finding was a strong inverse correlation between age of onset and disease duration. The PrP gene polymorphic codon 129 was found to confer 41% of the variance in age of onset but interestingly this polymorphism had no effect on disease duration suggesting different molecular mechanisms are involved in determining disease onset and rate of clinical progression.
Assuntos
Fenótipo , Doenças Priônicas/genética , Príons/genética , Adulto , Idade de Início , Apolipoproteínas E/genética , Códon/genética , Elementos de DNA Transponíveis/genética , Saúde da Família , Feminino , Haplótipos/genética , Humanos , Masculino , Repetições de Microssatélites/genética , Pessoa de Meia-Idade , Mutagênese Insercional , Proteínas Mutantes , Mutação/genética , Linhagem , Peptídeos/genética , Polimorfismo de Nucleotídeo Único/genética , Proteínas PrPSc/genética , Doenças Priônicas/patologia , Doenças Priônicas/transmissão , Fatores Sexuais , Fatores de TempoRESUMO
Prion diseases are a diverse group of neurodegenerative conditions, caused by the templated misfolding of prion protein. Aside from the strong genetic risk conferred by multiple variants of the prion protein gene (PRNP), several other variants have been suggested to confer risk in the most common type, sporadic Creutzfeldt-Jakob disease (sCJD) or in the acquired prion diseases. Large and rare copy number variants (CNVs) are known to confer risk in several related disorders including Alzheimer's disease (at APP), schizophrenia, epilepsy, mental retardation, and autism. Here, we report the first genome-wide analysis for CNV-associated risk using data derived from a recent international collaborative association study in sCJD (n = 1147 after quality control) and publicly available controls (n = 5427). We also investigated UK patients with variant Creutzfeldt-Jakob disease (n = 114) and elderly women from the Eastern Highlands of Papua New Guinea who proved highly resistant to the epidemic prion disease kuru, who were compared with healthy young Fore population controls (n = 395). There were no statistically significant alterations in the burden of CNVs >100, >500, or >1000 kb, duplications, or deletions in any disease group or geographic region. After correction for multiple testing, no statistically significant associations were found. A UK blood service control sample showed a duplication CNV that overlapped PRNP, but these were not found in prion disease. Heterozygous deletions of a 3' region of the PARK2 gene were found in 3 sCJD patients and no controls (p = 0.001, uncorrected). A cell-based prion infection assay did not provide supportive evidence for a role for PARK2 in prion disease susceptibility. These data are consistent with a modest impact of CNVs on risk of late-onset neurologic conditions and suggest that, unlike APP, PRNP duplication is not a causal high-risk mutation.
Assuntos
Variações do Número de Cópias de DNA/genética , Doenças Priônicas/genética , Príons/genética , Regiões 3' não Traduzidas/genética , Idoso , Células Cultivadas , Síndrome de Creutzfeldt-Jakob/genética , Feminino , Predisposição Genética para Doença/genética , Estudo de Associação Genômica Ampla , Humanos , Kuru/genética , Perda de Heterozigosidade/genética , Masculino , Proteínas Priônicas , Risco , Ubiquitina-Proteína Ligases/genéticaRESUMO
An expanded hexanucleotide repeat in the C9orf72 gene is the most common genetic cause of amyotrophic lateral sclerosis and frontotemporal dementia (C9ALS/FTD). Although 0-30 hexanucleotide repeats are present in the general population, expansions >500 repeats are associated with C9ALS/FTD. Large C9ALS/FTD expansions share a common haplotype and whether these expansions derive from a single founder or occur more frequently on a predisposing haplotype is yet to be determined and is relevant to disease pathomechanisms. Furthermore, although cases carrying 50-200 repeats have been described, their role and the pathogenic threshold of the expansions remain to be identified and carry importance for diagnostics and genetic counseling. We present clinical and genetic data from a UK ALS cohort and report the detailed molecular study of an atypical somatically unstable expansion of 90 repeats. Our results across different tissues provide evidence for the pathogenicity of this repeat number by showing they can somatically expand in the central nervous system to the well characterized pathogenic range. Our results support the occurrence of multiple expansion events for C9ALS/FTD.
Assuntos
Esclerose Lateral Amiotrófica/genética , Estudos de Coortes , Expansão das Repetições de DNA/genética , Proteínas/genética , Proteína C9orf72 , Demência Frontotemporal/genética , Humanos , Reino UnidoRESUMO
OBJECTIVE: In many cases where Huntington disease (HD) is suspected, the genetic test for HD is negative: these are known as HD phenocopies. A repeat expansion in the C9orf72 gene has recently been identified as a major cause of familial and sporadic frontotemporal lobar degeneration and amyotrophic lateral sclerosis. Our objective was to determine whether this mutation causes HD phenocopies. METHODS: A cohort of 514 HD phenocopy patients were analyzed for the C9orf72 expansion using repeat primed PCR. In cases where the expansion was found, Southern hybridization was performed to determine expansion size. Clinical case notes were reviewed to determine the phenotype of expansion-positive cases. RESULTS: Ten subjects (1.95%) had the expansion, making it the most common identified genetic cause of HD phenocopy presentations. The size of expansion was not significantly different from that associated with other clinical presentations of C9orf72 expanded cases. The C9orf72 expansion-positive subjects were characterized by the presence of movement disorders, including dystonia, chorea, myoclonus, tremor, and rigidity. Furthermore, the age at onset in this cohort was lower than previously reported for subjects with the C9orf72 expansion and included one case with pediatric onset. DISCUSSION: This study extends the known phenotype of the C9orf72 expansion in both age at onset and movement disorder symptoms. We propose a revised clinico-genetic algorithm for the investigation of HD phenocopy patients based on these data.
Assuntos
Expansão das Repetições de DNA/genética , Doença de Huntington/genética , Proteínas/genética , Adolescente , Adulto , Idade de Início , Proteína C9orf72 , Criança , Transtornos Cognitivos/etiologia , Transtornos Cognitivos/genética , Estudos de Coortes , Análise Mutacional de DNA , Feminino , Humanos , Doença de Huntington/complicações , Imageamento por Ressonância Magnética , Masculino , Entrevista Psiquiátrica Padronizada , Pessoa de Meia-Idade , Fenótipo , Índice de Gravidade de Doença , Adulto JovemRESUMO
Identification of a specific genetic cause of early onset dementia (EOD) is important but can be difficult because of pleiotropy, locus heterogeneity and accessibility of gene tests. Here we assess the use of next generation sequencing (NGS) technologies as a quick, accurate and cost effective method to determine genetic diagnosis in EOD. We developed gene panel based technologies to assess 16 genes known to harbour mutations causal of dementia and combined these with PCR based assessments of the C9orf72 hexanucleotide repeat expansion and the octapeptide repeat region of PRNP. In a blinded study of 95 samples we show very high sensitivity and specificity are achievable using either Ion Torrent or MiSeq sequencing platforms. Modifications to the gene panel permit accurate detection of structural variation in APP. In 2/10 samples which had been selected because they possess a variant of uncertain significance the new technology discovered a causal mutation in genes not previously sequenced. A large proportion (23/85) of samples showed genetic variants of uncertain significance in addition to known mutations. The MRC Dementia Gene Panel and similar technologies are likely to be transformational in EOD diagnosis with a significant impact on the proportion of patients in whom a genetic cause is identified.
Assuntos
Demência/diagnóstico , Demência/genética , Técnicas de Diagnóstico Molecular/métodos , Análise de Sequência de DNA/métodos , Precursor de Proteína beta-Amiloide/genética , Proteína C9orf72 , Expansão das Repetições de DNA , Variação Genética , Humanos , Mutação , Reação em Cadeia da Polimerase/métodos , Proteínas Priônicas , Príons/genética , Proteínas/genética , Sensibilidade e EspecificidadeRESUMO
OBJECTIVES: To carry out a further survey of archived appendix samples to understand better the differences between existing estimates of the prevalence of subclinical infection with prions after the bovine spongiform encephalopathy epizootic and to see whether a broader birth cohort was affected, and to understand better the implications for the management of blood and blood products and for the handling of surgical instruments. DESIGN: Irreversibly unlinked and anonymised large scale survey of archived appendix samples. SETTING: Archived appendix samples from the pathology departments of 41 UK hospitals participating in the earlier survey, and additional hospitals in regions with lower levels of participation in that survey. SAMPLE: 32,441 archived appendix samples fixed in formalin and embedded in paraffin and tested for the presence of abnormal prion protein (PrP). RESULTS: Of the 32,441 appendix samples 16 were positive for abnormal PrP, indicating an overall prevalence of 493 per million population (95% confidence interval 282 to 801 per million). The prevalence in those born in 1941-60 (733 per million, 269 to 1596 per million) did not differ significantly from those born between 1961 and 1985 (412 per million, 198 to 758 per million) and was similar in both sexes and across the three broad geographical areas sampled. Genetic testing of the positive specimens for the genotype at PRNP codon 129 revealed a high proportion that were valine homozygous compared with the frequency in the normal population, and in stark contrast with confirmed clinical cases of vCJD, all of which were methionine homozygous at PRNP codon 129. CONCLUSIONS: This study corroborates previous studies and suggests a high prevalence of infection with abnormal PrP, indicating vCJD carrier status in the population compared with the 177 vCJD cases to date. These findings have important implications for the management of blood and blood products and for the handling of surgical instruments.
Assuntos
Apêndice/química , Portador Sadio/epidemiologia , Síndrome de Creutzfeldt-Jakob/epidemiologia , Encefalopatia Espongiforme Bovina/epidemiologia , Príons/análise , Animais , Portador Sadio/metabolismo , Bovinos , Códon/genética , Estudos de Coortes , Síndrome de Creutzfeldt-Jakob/genética , Síndrome de Creutzfeldt-Jakob/metabolismo , Encefalopatia Espongiforme Bovina/genética , Encefalopatia Espongiforme Bovina/transmissão , Feminino , Testes Genéticos , Homozigoto , Humanos , Masculino , Prevalência , Proteínas Priônicas , Príons/genética , Reino Unido/epidemiologiaRESUMO
Amyloid precursor protein gene (APP) duplications have been identified in screens of selected probands with early onset familial Alzheimer's disease (FAD). A causal role for copy number variation (CNV) in the prion protein gene (PRNP) in prion dementias is not known. We aimed to determine the prevalence of copy number variation in APP and PRNP in a large referral series, test a screening method for detection of the same, and expand knowledge of clinical phenotype. We used a 3-tiered screening assay for APP and PRNP duplication (exonic real-time quantitative polymerase chain reaction [exon-qPCR], fluorescent microsatellite quantitative PCR [fm-q-PCR], and Illumina array [Illumina Inc., San Diego, CA, USA]) for analysis of a heterogeneous referral series comprising 1531 probands. Five of 1531 probands screened showed APP duplication, a similar prevalence to APP missense mutation. Real-time quantitative PCR and fluorescent microsatellite quantitative PCR were similar individually but are theoretically complementary; we used Illumina arrays as our reference assay. Two of 5 probands were from an autosomal dominant early onset Alzheimer's disease (familial Alzheimer's disease) pedigree. One extensive, noncontiguous duplication on chromosome 21 was consistent with an unbalanced translocation not including the Down's syndrome critical region. Seizures were prominent in the other typical APP duplications. A range of imaging, neuropsychological, cerebrospinal fluid, and pathological findings are reported that extend the known phenotype. APP but not PRNP duplication is a significant cause of early onset dementia in the UK. The recognized phenotype may be expanded to include the possibility of early seizures and apparently sporadic disease which, in part, may be due to different mutational mechanisms. The pros and cons of our screening method are discussed.
Assuntos
Precursor de Proteína beta-Amiloide/genética , Duplicação Gênica/genética , Predisposição Genética para Doença/epidemiologia , Predisposição Genética para Doença/genética , Príons/genética , Idoso , Idoso de 80 Anos ou mais , Causalidade , Feminino , Humanos , Masculino , Prevalência , Medição de Risco , Fatores de Risco , Reino Unido/epidemiologiaRESUMO
The acquired prion disease kuru was restricted to the Fore and neighbouring linguistic groups of the Papua New Guinea highlands and largely affected children and adult women. Oral history documents the onset of the epidemic in the early twentieth century, followed by a peak in the mid-twentieth century and subsequently a well-documented decline in frequency. In the context of these strong associations (gender, region and time), we have considered the genetic factors associated with susceptibility and resistance to kuru. Heterozygosity at codon 129 of the human prion protein gene (PRNP) is known to confer relative resistance to both sporadic and acquired prion diseases. In kuru, heterozygosity is associated with older patients and longer incubation times. Elderly survivors of the kuru epidemic, who had multiple exposures at mortuary feasts, are predominantly PRNP codon 129 heterozygotes and this group show marked Hardy-Weinberg disequilibrium. The deviation from Hardy-Weinberg equilibrium is most marked in elderly women, but is also significant in a slightly younger cohort of men, consistent with their exposure to kuru as boys. Young Fore and the elderly from populations with no history of kuru show Hardy-Weinberg equilibrium. An increasing cline in 129V allele frequency centres on the kuru region, consistent with the effect of selection in elevating the frequency of resistant genotypes in the exposed population. The genetic data are thus strikingly correlated with exposure. Considering the strong coding sequence conservation of primate prion protein genes, the number of global coding polymorphisms in man is surprising. By intronic resequencing in a European population, we have shown that haplotype diversity at PRNP comprises two major and divergent clades associated with 129M and 129V. Kuru may have imposed the strongest episode of recent human balancing selection, which may not have been an isolated episode in human history.
Assuntos
Etnicidade/genética , Predisposição Genética para Doença/genética , Variação Genética , Kuru/epidemiologia , Kuru/genética , Príons/genética , Feminino , Frequência do Gene , Haplótipos/genética , Heterozigoto , Humanos , Masculino , Papua Nova Guiné/epidemiologia , Proteínas Priônicas , Seleção GenéticaRESUMO
Alzheimer's disease (AD) is the commonest neurodegenerative disease worldwide. Rare familial cases may be caused by mutations in one of three genes-amyloid precursor protein, presenilin-1 and presenilin-2; however, the molecular basis of >99% of AD cases is unknown. Somatic mutation has been considered to be a mechanism that may account for a proportion of sporadic cases of AD, but to date there has been no evidence for this. We now report a sporadic early-onset patient with AD, and show that this individual is a somatic mosaic for a mutation in the presenilin-1 gene, suggesting a novel molecular mechanism for AD. Quantification of the mosaicism demonstrated the degree of mosaicism at 8% in peripheral lymphocytes and 14% in cerebral cortex in the index patient; a clear gene dosage effect on age of presentation and clinical phenotypic presentation is demonstrated. This finding has important implications for the aetiology of sporadic AD, and for other apparently sporadic neurodegenerative diseases such as Parkinson's disease, motor neuron disease and Creutzfeldt-Jakob disease.