RESUMEN
Chronic infection with the hepatitis C virus (HCV) affects 170 million people worldwide and is an important cause of liver-related morbidity and mortality. The standard of care therapy combines pegylated interferon (pegIFN) alpha and ribavirin (RBV), and is associated with a range of treatment-limiting adverse effects. One of the most important of these is RBV-induced haemolytic anaemia, which affects most patients and is severe enough to require dose modification in up to 15% of patients. Here we show that genetic variants leading to inosine triphosphatase deficiency, a condition not thought to be clinically important, protect against haemolytic anaemia in hepatitis-C-infected patients receiving RBV.
Asunto(s)
Anemia Hemolítica/inducido químicamente , Anemia Hemolítica/genética , Variación Genética/genética , Hepatitis C Crónica/tratamiento farmacológico , Pirofosfatasas/genética , Alelos , Anemia Hemolítica/complicaciones , Antivirales , Cromosomas Humanos Par 20 , Europa (Continente)/etnología , Estudio de Asociación del Genoma Completo , Hemoglobinas/deficiencia , Hemoglobinas/metabolismo , Hepatitis C Crónica/complicaciones , Humanos , Polimorfismo de Nucleótido Simple/genética , Pirofosfatasas/deficiencia , Pirofosfatasas/metabolismo , Grupos Raciales/genética , Ribavirina/uso terapéutico , Estados Unidos , Inosina TrifosfatasaRESUMEN
Human genetic variation contributes to differences in susceptibility to HIV-1 infection. To search for novel host resistance factors, we performed a genome-wide association study (GWAS) in hemophilia patients highly exposed to potentially contaminated factor VIII infusions. Individuals with hemophilia A and a documented history of factor VIII infusions before the introduction of viral inactivation procedures (1979-1984) were recruited from 36 hemophilia treatment centers (HTCs), and their genome-wide genetic variants were compared with those from matched HIV-infected individuals. Homozygous carriers of known CCR5 resistance mutations were excluded. Single nucleotide polymorphisms (SNPs) and inferred copy number variants (CNVs) were tested using logistic regression. In addition, we performed a pathway enrichment analysis, a heritability analysis, and a search for epistatic interactions with CCR5 Δ32 heterozygosity. A total of 560 HIV-uninfected cases were recruited: 36 (6.4%) were homozygous for CCR5 Δ32 or m303. After quality control and SNP imputation, we tested 1 081 435 SNPs and 3686 CNVs for association with HIV-1 serostatus in 431 cases and 765 HIV-infected controls. No SNP or CNV reached genome-wide significance. The additional analyses did not reveal any strong genetic effect. Highly exposed, yet uninfected hemophiliacs form an ideal study group to investigate host resistance factors. Using a genome-wide approach, we did not detect any significant associations between SNPs and HIV-1 susceptibility, indicating that common genetic variants of major effect are unlikely to explain the observed resistance phenotype in this population.
Asunto(s)
Resistencia a la Enfermedad/genética , Estudio de Asociación del Genoma Completo , Infecciones por VIH/genética , Hemofilia A/genética , Adulto , Variaciones en el Número de Copia de ADN , Epistasis Genética , Factor VIII/uso terapéutico , Femenino , Eliminación de Gen , Predisposición Genética a la Enfermedad , Seropositividad para VIH/genética , Heterocigoto , Homocigoto , Humanos , Modelos Logísticos , Masculino , Metaanálisis como Asunto , Persona de Mediana Edad , Fenotipo , Polimorfismo de Nucleótido Simple , Estudios Prospectivos , Receptores CCR5/genética , Receptores CCR5/metabolismoRESUMEN
Since the discovery of HIV as the cause of AIDS, numerous insights have been gained from studies of its natural history and epidemiology. It has become clear that there are substantial interindividual differences in the risk of HIV acquisition and course of disease. Meanwhile, the field of human genetics has undergone a series of rapid transitions that have fundamentally altered the approach to studying HIV host genetics. We aim to describe the field as it has transitioned from the era of candidate-gene studies and the era of genome-wide association studies (GWAS) to its current state in the infancy of comprehensive sequencing. In some ways the field has come full circle, having evolved from being driven almost exclusively by our knowledge of immunology, to a bias-free GWAS approach, to a point where our ability to catalogue human variation far outstrips our ability to biologically interpret it.
Asunto(s)
Predisposición Genética a la Enfermedad , Estudio de Asociación del Genoma Completo/métodos , Infecciones por VIH/genética , VIH , HumanosRESUMEN
Although there are many methods available for inferring copy-number variants (CNVs) from next-generation sequence data, there remains a need for a system that is computationally efficient but that retains good sensitivity and specificity across all types of CNVs. Here, we introduce a new method, estimation by read depth with single-nucleotide variants (ERDS), and use various approaches to compare its performance to other methods. We found that for common CNVs and high-coverage genomes, ERDS performs as well as the best method currently available (Genome STRiP), whereas for rare CNVs and high-coverage genomes, ERDS performs better than any available method. Importantly, ERDS accommodates both unique and highly amplified regions of the genome and does so without requiring separate alignments for calling CNVs and other variants. These comparisons show that for genomes sequenced at high coverage, ERDS provides a computationally convenient method that calls CNVs as well as or better than any currently available method.
Asunto(s)
Variaciones en el Número de Copia de ADN , Genoma Humano , Análisis de Secuencia de ADN/métodos , Algoritmos , Eliminación de Gen , Técnicas de Genotipaje , Humanos , Estudios de Validación como AsuntoRESUMEN
To date, the widely used genome-wide association studies (GWASs) of the human genome have reported thousands of variants that are significantly associated with various human traits. However, in the vast majority of these cases, the causal variants responsible for the observed associations remain unknown. In order to facilitate the identification of causal variants, we designed a simple computational method called the "preferential linkage disequilibrium (LD)" approach, which follows the variants discovered by GWASs to pinpoint the causal variants, even if they are rare compared with the discovery variants. The approach is based on the hypothesis that the GWAS-discovered variant is better at tagging the causal variants than are most other variants evaluated in the original GWAS. Applying the preferential LD approach to the GWAS signals of five human traits for which the causal variants are already known, we successfully placed the known causal variants among the top ten candidates in the majority of these cases. Application of this method to additional GWASs, including those of hepatitis C virus treatment response, plasma levels of clotting factors, and late-onset Alzheimer disease, has led to the identification of a number of promising candidate causal variants. This method represents a useful tool for delineating causal variants by bringing together GWAS signals and the rapidly accumulating variant data from next-generation sequencing.
Asunto(s)
Estudio de Asociación del Genoma Completo , Desequilibrio de Ligamiento , Biología Computacional/métodos , Frecuencia de los Genes , Predisposición Genética a la Enfermedad , Genoma Humano , Humanos , Polimorfismo de Nucleótido SimpleRESUMEN
We studied five individuals from three Jewish Bukharian families affected by an apparently autosomal-recessive form of hereditary spastic paraparesis accompanied by severe intellectual disability, fluctuating central hypoventilation, gastresophageal reflux disease, wake apnea, areflexia, and unique dysmorphic features. Exome sequencing identified one homozygous variant shared among all affected individuals and absent in controls: a 1 bp frameshift TECPR2 deletion leading to a premature stop codon and predicting significant degradation of the protein. TECPR2 has been reported as a positive regulator of autophagy. We thus examined the autophagy-related fate of two key autophagic proteins, SQSTM1 (p62) and MAP1LC3B (LC3), in skin fibroblasts of an affected individual, as compared to a healthy control, and found that both protein levels were decreased and that there was a more pronounced decrease in the lipidated form of LC3 (LC3II). siRNA knockdown of TECPR2 showed similar changes, consistent with aberrant autophagy. Our results are strengthened by the fact that autophagy dysfunction has been implicated in a number of other neurodegenerative diseases. The discovered TECPR2 mutation implicates autophagy, a central intracellular mechanism, in spastic paraparesis.
Asunto(s)
Autofagia/genética , Proteínas Portadoras/genética , Mutación , Proteínas del Tejido Nervioso/genética , Paraparesia Espástica/genética , Encéfalo/patología , Exones , Femenino , Fibroblastos/metabolismo , Fibroblastos/ultraestructura , Genotipo , Células HeLa , Humanos , Judíos/genética , Imagen por Resonancia Magnética , Masculino , Neuroimagen , Paraparesia Espástica/diagnóstico , Paraparesia Espástica/metabolismo , Linaje , Fenotipo , Análisis de Secuencia de ADNRESUMEN
Schizophrenia is a severe psychiatric disorder with strong heritability and marked heterogeneity in symptoms, course, and treatment response. There is strong interest in identifying genetic risk factors that can help to elucidate the pathophysiology and that might result in the development of improved treatments. Linkage and genome-wide association studies (GWASs) suggest that the genetic basis of schizophrenia is heterogeneous. However, it remains unclear whether the underlying genetic variants are mostly moderately rare and can be identified by the genotyping of variants observed in sequenced cases in large follow-up cohorts or whether they will typically be much rarer and therefore more effectively identified by gene-based methods that seek to combine candidate variants. Here, we consider 166 persons who have schizophrenia or schizoaffective disorder and who have had either their genomes or their exomes sequenced to high coverage. From these data, we selected 5,155 variants that were further evaluated in an independent cohort of 2,617 cases and 1,800 controls. No single variant showed a study-wide significant association in the initial or follow-up cohorts. However, we identified a number of case-specific variants, some of which might be real risk factors for schizophrenia, and these can be readily interrogated in other data sets. Our results indicate that schizophrenia risk is unlikely to be predominantly influenced by variants just outside the range detectable by GWASs. Rather, multiple rarer genetic variants must contribute substantially to the predisposition to schizophrenia, suggesting that both very large sample sizes and gene-based association tests will be required for securely identifying genetic risk factors.
Asunto(s)
Exoma/genética , Predisposición Genética a la Enfermedad/genética , Esquizofrenia/genética , Secuencia de Bases , Finlandia , Estudio de Asociación del Genoma Completo , Genotipo , Humanos , Datos de Secuencia Molecular , Factores de Riesgo , Alineación de Secuencia , Análisis de Secuencia de ADN , Estados UnidosRESUMEN
Whole-exome sequencing (WES), which analyzes the coding sequence of most annotated genes in the human genome, is an ideal approach to studying fully penetrant autosomal-recessive diseases, and it has been very powerful in identifying disease-causing mutations even when enrollment of affected individuals is limited by reduced survival. In this study, we combined WES with homozygosity analysis of consanguineous pedigrees, which are informative even when a single affected individual is available, to identify genetic mutations responsible for Walker-Warburg syndrome (WWS), a genetically heterogeneous autosomal-recessive disorder that severely affects the development of the brain, eyes, and muscle. Mutations in seven genes are known to cause WWS and explain 50%-60% of cases, but multiple additional genes are expected to be mutated because unexplained cases show suggestive linkage to diverse loci. Using WES in consanguineous WWS-affected families, we found multiple deleterious mutations in GTDC2 (also known as AGO61). GTDC2's predicted role as an uncharacterized glycosyltransferase is consistent with the function of other genes that are known to be mutated in WWS and that are involved in the glycosylation of the transmembrane receptor dystroglycan. Therefore, to explore the role of GTDC2 loss of function during development, we used morpholino-mediated knockdown of its zebrafish ortholog, gtdc2. We found that gtdc2 knockdown in zebrafish replicates all WWS features (hydrocephalus, ocular defects, and muscular dystrophy), strongly suggesting that GTDC2 mutations cause WWS.
Asunto(s)
Glicosiltransferasas/genética , Síndrome de Walker-Warburg/genética , Exoma , Humanos , MutaciónRESUMEN
Idiopathic generalized epilepsy (IGE) is a complex disease with high heritability, but little is known about its genetic architecture. Rare copy-number variants have been found to explain nearly 3% of individuals with IGE; however, it remains unclear whether variants with moderate effect size and frequencies below what are reliably detected with genome-wide association studies contribute significantly to disease risk. In this study, we compare the exome sequences of 118 individuals with IGE and 242 controls of European ancestry by using next-generation sequencing. The exome-sequenced epilepsy cases include study subjects with two forms of IGE, including juvenile myoclonic epilepsy (n = 93) and absence epilepsy (n = 25). However, our discovery strategy did not assume common genetic control between the subtypes of IGE considered. In the sequence data, as expected, no variants were significantly associated with the IGE phenotype or more specific IGE diagnoses. We then selected 3,897 candidate epilepsy-susceptibility variants from the sequence data and genotyped them in a larger set of 878 individuals with IGE and 1,830 controls. Again, no variant achieved statistical significance. However, 1,935 variants were observed exclusively in cases either as heterozygous or homozygous genotypes. It is likely that this set of variants includes real risk factors. The lack of significant association evidence of single variants with disease in this two-stage approach emphasizes the high genetic heterogeneity of epilepsy disorders, suggests that the impact of any individual single-nucleotide variant in this disease is small, and indicates that gene-based approaches might be more successful for future sequencing studies of epilepsy predisposition.
Asunto(s)
Epilepsia Generalizada/genética , Exoma/genética , Predisposición Genética a la Enfermedad/genética , Secuencia de Bases , Estudio de Asociación del Genoma Completo , Genotipo , Humanos , Datos de Secuencia Molecular , Alineación de Secuencia , Análisis de Secuencia de ADN , Población Blanca/genéticaRESUMEN
BACKGROUND: Mendelian analysis of disorders of immune regulation can provide insight into molecular pathways associated with host defense and immune tolerance. METHODS: We identified three families with a dominantly inherited complex of cold-induced urticaria, antibody deficiency, and susceptibility to infection and autoimmunity. Immunophenotyping methods included flow cytometry, analysis of serum immunoglobulins and autoantibodies, lymphocyte stimulation, and enzymatic assays. Genetic studies included linkage analysis, targeted Sanger sequencing, and next-generation whole-genome sequencing. RESULTS: Cold urticaria occurred in all affected subjects. Other, variable manifestations included atopy, granulomatous rash, autoimmune thyroiditis, the presence of antinuclear antibodies, sinopulmonary infections, and common variable immunodeficiency. Levels of serum IgM and IgA and circulating natural killer cells and class-switched memory B cells were reduced. Linkage analysis showed a 7-Mb candidate interval on chromosome 16q in one family, overlapping by 3.5 Mb a disease-associated haplotype in a smaller family. This interval includes PLCG2, encoding phospholipase Cγ(2) (PLCγ(2)), a signaling molecule expressed in B cells, natural killer cells, and mast cells. Sequencing of complementary DNA revealed heterozygous transcripts lacking exon 19 in two families and lacking exons 20 through 22 in a third family. Genomic sequencing identified three distinct in-frame deletions that cosegregated with disease. These deletions, located within a region encoding an autoinhibitory domain, result in protein products with constitutive phospholipase activity. PLCG2-expressing cells had diminished cellular signaling at 37°C but enhanced signaling at subphysiologic temperatures. CONCLUSIONS: Genomic deletions in PLCG2 cause gain of PLCγ(2) function, leading to signaling abnormalities in multiple leukocyte subsets and a phenotype encompassing both excessive and deficient immune function. (Funded by the National Institutes of Health Intramural Research Programs and others.).
Asunto(s)
Enfermedades Autoinmunes/genética , Síndromes Periódicos Asociados a Criopirina/genética , Síndromes de Inmunodeficiencia/genética , Fosfolipasa C gamma/genética , Eliminación de Secuencia , Frío/efectos adversos , ADN Complementario/análisis , ADN Complementario/aislamiento & purificación , Femenino , Humanos , Masculino , Linaje , Fenotipo , Fosfolipasa C gamma/metabolismo , Polimorfismo de Nucleótido Simple , Análisis de Secuencia de ADNRESUMEN
Chronic infection with hepatitis C virus (HCV) affects 170 million people worldwide and is the leading cause of cirrhosis in North America. Although the recommended treatment for chronic infection involves a 48-week course of peginterferon-alpha-2b (PegIFN-alpha-2b) or -alpha-2a (PegIFN-alpha-2a) combined with ribavirin (RBV), it is well known that many patients will not be cured by treatment, and that patients of European ancestry have a significantly higher probability of being cured than patients of African ancestry. In addition to limited efficacy, treatment is often poorly tolerated because of side effects that prevent some patients from completing therapy. For these reasons, identification of the determinants of response to treatment is a high priority. Here we report that a genetic polymorphism near the IL28B gene, encoding interferon-lambda-3 (IFN-lambda-3), is associated with an approximately twofold change in response to treatment, both among patients of European ancestry (P = 1.06 x 10(-25)) and African-Americans (P = 2.06 x 10(-3)). Because the genotype leading to better response is in substantially greater frequency in European than African populations, this genetic polymorphism also explains approximately half of the difference in response rates between African-Americans and patients of European ancestry.
Asunto(s)
Variación Genética/genética , Hepacivirus/efectos de los fármacos , Hepatitis C Crónica/tratamiento farmacológico , Hepatitis C Crónica/genética , Interferón-alfa/farmacología , Interleucinas/genética , Polietilenglicoles/farmacología , Carga Viral , Negro o Afroamericano/genética , Cromosomas Humanos Par 19/genética , Ensayos Clínicos como Asunto , Europa (Continente)/etnología , Asia Oriental/etnología , Frecuencia de los Genes , Genoma Humano/genética , Estudio de Asociación del Genoma Completo , Genotipo , Hepatitis C Crónica/etnología , Hepatitis C Crónica/virología , Hispánicos o Latinos/genética , Humanos , Interferón alfa-2 , Interferón-alfa/efectos adversos , Interferón-alfa/uso terapéutico , Interferones , Farmacogenética , Polietilenglicoles/efectos adversos , Polietilenglicoles/uso terapéutico , Polimorfismo de Nucleótido Simple/genética , Proteínas RecombinantesRESUMEN
One of the longest running debates in evolutionary biology concerns the kind of genetic variation that is primarily responsible for phenotypic variation in species. Here, we address this question for humans specifically from the perspective of population allele frequency of variants across the complete genome, including both coding and noncoding regions. We establish simple criteria to assess the likelihood that variants are functional based on their genomic locations and then use whole-genome sequence data from 29 subjects of European origin to assess the relationship between the functional properties of variants and their population allele frequencies. We find that for all criteria used to assess the likelihood that a variant is functional, the rarer variants are significantly more likely to be functional than the more common variants. Strikingly, these patterns disappear when we focus on only those variants in which the major alleles are derived. These analyses indicate that the majority of the genetic variation in terms of phenotypic consequence may result from a mutation-selection balance, as opposed to balancing selection, and have direct relevance to the study of human disease.
Asunto(s)
Variación Genética , Alelos , Secuencia Conservada , Evolución Molecular , Frecuencia de los Genes , Genes Reguladores , Genoma Humano , Estudio de Asociación del Genoma Completo , Humanos , Modelos Genéticos , Mutación , Fenotipo , Polimorfismo de Nucleótido Simple , Selección Genética , Población Blanca/genéticaRESUMEN
BACKGROUND: Carbamazepine causes various forms of hypersensitivity reactions, ranging from maculopapular exanthema to severe blistering reactions. The HLA-B*1502 allele has been shown to be strongly correlated with carbamazepine-induced Stevens-Johnson syndrome and toxic epidermal necrolysis (SJS-TEN) in the Han Chinese and other Asian populations but not in European populations. METHODS: We performed a genomewide association study of samples obtained from 22 subjects with carbamazepine-induced hypersensitivity syndrome, 43 subjects with carbamazepine-induced maculopapular exanthema, and 3987 control subjects, all of European descent. We tested for an association between disease and HLA alleles through proxy single-nucleotide polymorphisms and imputation, confirming associations by high-resolution sequence-based HLA typing. We replicated the associations in samples from 145 subjects with carbamazepine-induced hypersensitivity reactions. RESULTS: The HLA-A*3101 allele, which has a prevalence of 2 to 5% in Northern European populations, was significantly associated with the hypersensitivity syndrome (P=3.5×10(-8)). An independent genomewide association study of samples from subjects with maculopapular exanthema also showed an association with the HLA-A*3101 allele (P=1.1×10(-6)). Follow-up genotyping confirmed the variant as a risk factor for the hypersensitivity syndrome (odds ratio, 12.41; 95% confidence interval [CI], 1.27 to 121.03), maculopapular exanthema (odds ratio, 8.33; 95% CI, 3.59 to 19.36), and SJS-TEN (odds ratio, 25.93; 95% CI, 4.93 to 116.18). CONCLUSIONS: The presence of the HLA-A*3101 allele was associated with carbamazepine-induced hypersensitivity reactions among subjects of Northern European ancestry. The presence of the allele increased the risk from 5.0% to 26.0%, whereas its absence reduced the risk from 5.0% to 3.8%. (Funded by the U.K. Department of Health and others.).
Asunto(s)
Anticonvulsivantes/efectos adversos , Carbamazepina/efectos adversos , Hipersensibilidad a las Drogas/genética , Antígenos HLA-A/genética , Población Blanca/genética , Anticonvulsivantes/uso terapéutico , Carbamazepina/uso terapéutico , Exantema/inducido químicamente , Exantema/genética , Estudio de Asociación del Genoma Completo , Genotipo , Prueba de Histocompatibilidad , Humanos , Polimorfismo de Nucleótido Simple , Síndrome de Stevens-Johnson/inducido químicamente , Síndrome de Stevens-Johnson/genéticaRESUMEN
A genome-wide screen for large structural variants showed that a copy number variant (CNV) in the region encoding killer cell immunoglobulin-like receptors (KIR) associates with HIV-1 control as measured by plasma viral load at set point in individuals of European ancestry. This CNV encompasses the KIR3DL1-KIR3DS1 locus, encoding receptors that interact with specific HLA-Bw4 molecules to regulate the activation of lymphocyte subsets including natural killer (NK) cells. We quantified the number of copies of KIR3DS1 and KIR3DL1 in a large HIV-1 positive cohort, and showed that an increase in KIR3DS1 count associates with a lower viral set point if its putative ligand is present (p = 0.00028), as does an increase in KIR3DL1 count in the presence of KIR3DS1 and appropriate ligands for both receptors (p = 0.0015). We further provide functional data that demonstrate that NK cells from individuals with multiple copies of KIR3DL1, in the presence of KIR3DS1 and the appropriate ligands, inhibit HIV-1 replication more robustly, and associated with a significant expansion in the frequency of KIR3DS1+, but not KIR3DL1+, NK cells in their peripheral blood. Our results suggest that the relative amounts of these activating and inhibitory KIR play a role in regulating the peripheral expansion of highly antiviral KIR3DS1+ NK cells, which may determine differences in HIV-1 control following infection.
Asunto(s)
Variaciones en el Número de Copia de ADN , VIH-1/fisiología , Receptores KIR/genética , Estudios de Cohortes , VIH-1/inmunología , Humanos , Células Asesinas Naturales/metabolismo , Células Asesinas Naturales/fisiología , Activación de Linfocitos , Modelos Inmunológicos , Receptores KIR/metabolismo , Carga Viral , Replicación ViralRESUMEN
Reduced fecundity, associated with severe mental disorders, places negative selection pressure on risk alleles and may explain, in part, why common variants have not been found that confer risk of disorders such as autism, schizophrenia and mental retardation. Thus, rare variants may account for a larger fraction of the overall genetic risk than previously assumed. In contrast to rare single nucleotide mutations, rare copy number variations (CNVs) can be detected using genome-wide single nucleotide polymorphism arrays. This has led to the identification of CNVs associated with mental retardation and autism. In a genome-wide search for CNVs associating with schizophrenia, we used a population-based sample to identify de novo CNVs by analysing 9,878 transmissions from parents to offspring. The 66 de novo CNVs identified were tested for association in a sample of 1,433 schizophrenia cases and 33,250 controls. Three deletions at 1q21.1, 15q11.2 and 15q13.3 showing nominal association with schizophrenia in the first sample (phase I) were followed up in a second sample of 3,285 cases and 7,951 controls (phase II). All three deletions significantly associate with schizophrenia and related psychoses in the combined sample. The identification of these rare, recurrent risk variants, having occurred independently in multiple founders and being subject to negative selection, is important in itself. CNV analysis may also point the way to the identification of additional and more prevalent risk variants in genes and pathways involved in schizophrenia.
Asunto(s)
Predisposición Genética a la Enfermedad/genética , Esquizofrenia/genética , Eliminación de Secuencia/genética , China , Cromosomas Humanos Par 1/genética , Cromosomas Humanos Par 15/genética , Europa (Continente) , Dosificación de Gen/genética , Genoma Humano/genética , Genotipo , Humanos , Pérdida de Heterocigocidad , Modelos Genéticos , Polimorfismo de Nucleótido Simple/genética , Trastornos Psicóticos/genéticaRESUMEN
Deletions at 16p13.11 are associated with schizophrenia, mental retardation, and most recently idiopathic generalized epilepsy. To evaluate the role of 16p13.11 deletions, as well as other structural variation, in epilepsy disorders, we used genome-wide screens to identify copy number variation in 3812 patients with a diverse spectrum of epilepsy syndromes and in 1299 neurologically-normal controls. Large deletions (> 100 kb) at 16p13.11 were observed in 23 patients, whereas no control had a deletion greater than 16 kb. Patients, even those with identically sized 16p13.11 deletions, presented with highly variable epilepsy phenotypes. For a subset of patients with a 16p13.11 deletion, we show a consistent reduction of expression for included genes, suggesting that haploinsufficiency might contribute to pathogenicity. We also investigated another possible mechanism of pathogenicity by using hybridization-based capture and next-generation sequencing of the homologous chromosome for ten 16p13.11-deletion patients to look for unmasked recessive mutations. Follow-up genotyping of suggestive polymorphisms failed to identify any convincing recessive-acting mutations in the homologous interval corresponding to the deletion. The observation that two of the 16p13.11 deletions were larger than 2 Mb in size led us to screen for other large deletions. We found 12 additional genomic regions harboring deletions > 2 Mb in epilepsy patients, and none in controls. Additional evaluation is needed to characterize the role of these exceedingly large, non-locus-specific deletions in epilepsy. Collectively, these data implicate 16p13.11 and possibly other large deletions as risk factors for a wide range of epilepsy disorders, and they appear to point toward haploinsufficiency as a contributor to the pathogenicity of deletions.
Asunto(s)
Cromosomas Humanos Par 16 , Susceptibilidad a Enfermedades , Epilepsia/genética , Mutación , Eliminación de Secuencia , Humanos , Hibridación de Ácido Nucleico/genética , SíndromeRESUMEN
BACKGROUND: There is considerable interest in the use of next-generation sequencing to help diagnose unidentified genetic conditions, but it is difficult to predict the success rate in a clinical setting that includes patients with a broad range of phenotypic presentations. METHODS: The authors present a pilot programme of whole-exome sequencing on 12 patients with unexplained and apparent genetic conditions, along with their unaffected parents. Unlike many previous studies, the authors did not seek patients with similar phenotypes, but rather enrolled any undiagnosed proband with an apparent genetic condition when predetermined criteria were met. RESULTS: This undertaking resulted in a likely genetic diagnosis in 6 of the 12 probands, including the identification of apparently causal mutations in four genes known to cause Mendelian disease (TCF4, EFTUD2, SCN2A and SMAD4) and one gene related to known Mendelian disease genes (NGLY1). Of particular interest is that at the time of this study, EFTUD2 was not yet known as a Mendelian disease gene but was nominated as a likely cause based on the observation of de novo mutations in two unrelated probands. In a seventh case with multiple disparate clinical features, the authors were able to identify homozygous mutations in EFEMP1 as a likely cause for macular degeneration (though likely not for other features). CONCLUSIONS: This study provides evidence that next-generation sequencing can have high success rates in a clinical setting, but also highlights key challenges. It further suggests that the presentation of known Mendelian conditions may be considerably broader than currently recognised.
Asunto(s)
Exoma , Enfermedades Genéticas Congénitas/diagnóstico , Técnicas de Diagnóstico Molecular/métodos , Análisis de Secuencia de ADN/métodos , Adolescente , Adulto , Factores de Transcripción Básicos con Cremalleras de Leucinas y Motivos Hélice-Asa-Hélice/genética , Niño , Preescolar , Femenino , Enfermedades Genéticas Congénitas/genética , Humanos , Lactante , Masculino , Modelos Genéticos , Mutación , Canal de Sodio Activado por Voltaje NAV1.2/genética , Péptido-N4-(N-acetil-beta-glucosaminil) Asparagina Amidasa/genética , Proteína Smad4/genética , Factor de Transcripción 4 , Factores de Transcripción/genéticaRESUMEN
Although more than 2,400 genes have been shown to contain variants that cause Mendelian disease, there are still several thousand such diseases yet to be molecularly defined. The ability of new whole-genome sequencing technologies to rapidly indentify most of the genetic variants in any given genome opens an exciting opportunity to identify these disease genes. Here we sequenced the whole genome of a single patient with the dominant Mendelian disease, metachondromatosis (OMIM 156250), and used partial linkage data from her small family to focus our search for the responsible variant. In the proband, we identified an 11 bp deletion in exon four of PTPN11, which alters frame, results in premature translation termination, and co-segregates with the phenotype. In a second metachondromatosis family, we confirmed our result by identifying a nonsense mutation in exon 4 of PTPN11 that also co-segregates with the phenotype. Sequencing PTPN11 exon 4 in 469 controls showed no such protein truncating variants, supporting the pathogenicity of these two mutations. This combination of a new technology and a classical genetic approach provides a powerful strategy to discover the genes responsible for unexplained Mendelian disorders.
Asunto(s)
Ligamiento Genético , Predisposición Genética a la Enfermedad , Genoma Humano , Proteína Tirosina Fosfatasa no Receptora Tipo 11/genética , Exones , Femenino , Estudio de Asociación del Genoma Completo , Humanos , Masculino , Mutación , Linaje , Análisis de Secuencia de ADNRESUMEN
We present the analysis of twenty human genomes to evaluate the prospects for identifying rare functional variants that contribute to a phenotype of interest. We sequenced at high coverage ten "case" genomes from individuals with severe hemophilia A and ten "control" genomes. We summarize the number of genetic variants emerging from a study of this magnitude, and provide a proof of concept for the identification of rare and highly-penetrant functional variants by confirming that the cause of hemophilia A is easily recognizable in this data set. We also show that the number of novel single nucleotide variants (SNVs) discovered per genome seems to stabilize at about 144,000 new variants per genome, after the first 15 individuals have been sequenced. Finally, we find that, on average, each genome carries 165 homozygous protein-truncating or stop loss variants in genes representing a diverse set of pathways.
Asunto(s)
Genoma Humano/genética , Análisis de Secuencia de ADN , Secuencia de Bases , Estudios de Casos y Controles , Variaciones en el Número de Copia de ADN/genética , Bases de Datos Genéticas , Exones/genética , Factor VIII/genética , Duplicación de Gen/genética , Técnicas de Inactivación de Genes , Genética de Población , Genotipo , Hemofilia A/genética , Humanos , Mutación INDEL/genética , Análisis de Secuencia por Matrices de Oligonucleótidos , Sistemas de Lectura Abierta/genética , Polimorfismo Genético , Polimorfismo de Nucleótido Simple/genéticaRESUMEN
By impairing both function and survival, the severe reduction in oxygen availability associated with high-altitude environments is likely to act as an agent of natural selection. We used genomic and candidate gene approaches to search for evidence of such genetic selection. First, a genome-wide allelic differentiation scan (GWADS) comparing indigenous highlanders of the Tibetan Plateau (3,200-3,500 m) with closely related lowland Han revealed a genome-wide significant divergence across eight SNPs located near EPAS1. This gene encodes the transcription factor HIF2alpha, which stimulates production of red blood cells and thus increases the concentration of hemoglobin in blood. Second, in a separate cohort of Tibetans residing at 4,200 m, we identified 31 EPAS1 SNPs in high linkage disequilibrium that correlated significantly with hemoglobin concentration. The sex-adjusted hemoglobin concentration was, on average, 0.8 g/dL lower in the major allele homozygotes compared with the heterozygotes. These findings were replicated in a third cohort of Tibetans residing at 4,300 m. The alleles associating with lower hemoglobin concentrations were correlated with the signal from the GWADS study and were observed at greatly elevated frequencies in the Tibetan cohorts compared with the Han. High hemoglobin concentrations are a cardinal feature of chronic mountain sickness offering one plausible mechanism for selection. Alternatively, as EPAS1 is pleiotropic in its effects, selection may have operated on some other aspect of the phenotype. Whichever of these explanations is correct, the evidence for genetic selection at the EPAS1 locus from the GWADS study is supported by the replicated studies associating function with the allelic variants.