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1.
Neuroimage ; 203: 116155, 2019 Sep 05.
Artigo em Inglês | MEDLINE | ID: mdl-31494251

RESUMO

Most of human genome is present in two copies (maternal and paternal). However, segments of the genome can be deleted or duplicated, and many of these genomic variations (known as Copy Number Variants) are associated with psychiatric disorders. 16p11.2 copy number variants (breakpoint 4-5) confer high risk for neurodevelopmental disorders and are associated with structural brain alterations of large effect-size. Methods used in previous studies were unable to investigate the onset of these alterations and whether they evolve with age. In this study, we aim at characterizing age-related effects of 16p11.2 copy number variants by analyzing a group with a broad age range including younger individuals. A large normative developmental dataset was used to accurately adjust for effects of age. We normalized volumes of segmented brain regions as well as volumes of each voxel defined by tensor-based morphometry. Results show that the total intracranial volumes, the global gray and white matter volumes are respectively higher and lower in deletion and duplication carriers compared to control subjects at 4.5 years of age. These differences remain stable through childhood, adolescence and adulthood until 23 years of age (range: 0.5 to 1.0 Z-score). Voxel-based results are consistent with previous findings in 16p11.2 copy number variant carriers, including increased volume in the calcarine cortex and insula in deletions, compared to controls, with an inverse effect in duplication carriers (1.0 Z-score). All large effect-size voxel-based differences are present at 4.5 years and seem to remain stable until the age of 23. Our results highlight the stability of a neuroimaging endophenotype over 2 decades during which neurodevelopmental symptoms evolve at a rapid pace.

2.
Nat Commun ; 10(1): 3465, 2019 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-31371714

RESUMO

Brain morphogenesis is an important process contributing to higher-order cognition, however our knowledge about its biological basis is largely incomplete. Here we analyze 118 neuroanatomical parameters in 1,566 mutant mouse lines and identify 198 genes whose disruptions yield NeuroAnatomical Phenotypes (NAPs), mostly affecting structures implicated in brain connectivity. Groups of functionally similar NAP genes participate in pathways involving the cytoskeleton, the cell cycle and the synapse, display distinct fetal and postnatal brain expression dynamics and importantly, their disruption can yield convergent phenotypic patterns. 17% of human unique orthologues of mouse NAP genes are known loci for cognitive dysfunction. The remaining 83% constitute a vast pool of genes newly implicated in brain architecture, providing the largest study of mouse NAP genes and pathways. This offers a complementary resource to human genetic studies and predict that many more genes could be involved in mammalian brain morphogenesis.

3.
Nat Commun ; 10(1): 3300, 2019 Jul 24.
Artigo em Inglês | MEDLINE | ID: mdl-31341166

RESUMO

Genome-wide association studies (GWAS) have identified thousands of variants associated with complex traits, but their biological interpretation often remains unclear. Most of these variants overlap with expression QTLs, indicating their potential involvement in regulation of gene expression. Here, we propose a transcriptome-wide summary statistics-based Mendelian Randomization approach (TWMR) that uses multiple SNPs as instruments and multiple gene expression traits as exposures, simultaneously. Applied to 43 human phenotypes, it uncovers 3,913 putatively causal gene-trait associations, 36% of which have no genome-wide significant SNP nearby in previous GWAS. Using independent association summary statistics, we find that the majority of these loci were missed by GWAS due to power issues. Noteworthy among these links is educational attainment-associated BSCL2, known to carry mutations leading to a Mendelian form of encephalopathy. We also find pleiotropic causal effects suggestive of mechanistic connections. TWMR better accounts for pleiotropy and has the potential to identify biological mechanisms underlying complex traits.

4.
Eur J Hum Genet ; 27(5): 760-771, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30700833

RESUMO

Precise breakpoint mapping of balanced chromosomal rearrangements is crucial to identify disease etiology. Ten female patients with X-autosome balanced translocations associated with phenotypic alterations were evaluated, by mapping and sequencing their breakpoints. The rearrangements' impact on the expression of disrupted genes, and inferred mechanisms of formation in each case were assessed. For four patients that presented one of the chromosomal breaks in heterochromatic and highly repetitive segments, we combined cytogenomic methods and short-read sequencing to characterize, at nucleotide resolution, breakpoints that occurred in reference genome gaps. Most of rearrangements were possibly formed by non-homologous end joining and have breakpoints at repeat elements. Seven genes were found to be disrupted in six patients. Six of the affected genes showed altered expression, and the functional impairment of three of them were considered pathogenic. One gene disruption was considered potentially pathogenic, and three had uncertain clinical significance. Four patients presented no gene disruptions, suggesting other pathogenic mechanisms. Four genes were considered potentially affected by position effect and the expression abrogation of one of them was confirmed. This study emphasizes the importance of breakpoint-junction characterization at nucleotide resolution in balanced rearrangements to reveal genetic mechanisms associated with the patients' phenotypes, mechanisms of formation that originated the rearrangements, and genomic nature of disrupted DNA sequences.

5.
Neuron ; 2018 Nov 14.
Artigo em Inglês | MEDLINE | ID: mdl-30449657

RESUMO

Corpus callosum malformations are associated with a broad range of neurodevelopmental diseases. We report that de novo mutations in MAST1 cause mega-corpus-callosum syndrome with cerebellar hypoplasia and cortical malformations (MCC-CH-CM) in the absence of megalencephaly. We show that MAST1 is a microtubule-associated protein that is predominantly expressed in post-mitotic neurons and is present in both dendritic and axonal compartments. We further show that Mast1 null animals are phenotypically normal, whereas the deletion of a single amino acid (L278del) recapitulates the distinct neurological phenotype observed in patients. In animals harboring Mast1 microdeletions, we find that the PI3K/AKT3/mTOR pathway is unperturbed, whereas Mast2 and Mast3 levels are diminished, indicative of a dominant-negative mode of action. Finally, we report that de novo MAST1 substitutions are present in patients with autism and microcephaly, raising the prospect that mutations in this gene give rise to a spectrum of neurodevelopmental diseases.

6.
Genet Med ; 2018 Sep 07.
Artigo em Inglês | MEDLINE | ID: mdl-30190612

RESUMO

PURPOSE: To assess the contribution of rare variants in the genetic background toward variability of neurodevelopmental phenotypes in individuals with rare copy-number variants (CNVs) and gene-disruptive variants. METHODS: We analyzed quantitative clinical information, exome sequencing, and microarray data from 757 probands and 233 parents and siblings who carry disease-associated variants. RESULTS: The number of rare likely deleterious variants in functionally intolerant genes ("other hits") correlated with expression of neurodevelopmental phenotypes in probands with 16p12.1 deletion (n=23, p=0.004) and in autism probands carrying gene-disruptive variants (n=184, p=0.03) compared with their carrier family members. Probands with 16p12.1 deletion and a strong family history presented more severe clinical features (p=0.04) and higher burden of other hits compared with those with mild/no family history (p=0.001). The number of other hits also correlated with severity of cognitive impairment in probands carrying pathogenic CNVs (n=53) or de novo pathogenic variants in disease genes (n=290), and negatively correlated with head size among 80 probands with 16p11.2 deletion. These co-occurring hits involved known disease-associated genes such as SETD5, AUTS2, and NRXN1, and were enriched for cellular and developmental processes. CONCLUSION: Accurate genetic diagnosis of complex disorders will require complete evaluation of the genetic background even after a candidate disease-associated variant is identified.

7.
Biol Psychiatry ; 84(4): 253-264, 2018 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-29778275

RESUMO

BACKGROUND: 16p11.2 breakpoint 4 to 5 copy number variants (CNVs) increase the risk for developing autism spectrum disorder, schizophrenia, and language and cognitive impairment. In this multisite study, we aimed to quantify the effect of 16p11.2 CNVs on brain structure. METHODS: Using voxel- and surface-based brain morphometric methods, we analyzed structural magnetic resonance imaging collected at seven sites from 78 individuals with a deletion, 71 individuals with a duplication, and 212 individuals without a CNV. RESULTS: Beyond the 16p11.2-related mirror effect on global brain morphometry, we observe regional mirror differences in the insula (deletion > control > duplication). Other regions are preferentially affected by either the deletion or the duplication: the calcarine cortex and transverse temporal gyrus (deletion > control; Cohen's d > 1), the superior and middle temporal gyri (deletion < control; Cohen's d < -1), and the caudate and hippocampus (control > duplication; -0.5 > Cohen's d > -1). Measures of cognition, language, and social responsiveness and the presence of psychiatric diagnoses do not influence these results. CONCLUSIONS: The global and regional effects on brain morphometry due to 16p11.2 CNVs generalize across site, computational method, age, and sex. Effect sizes on neuroimaging and cognitive traits are comparable. Findings partially overlap with results of meta-analyses performed across psychiatric disorders. However, the lack of correlation between morphometric and clinical measures suggests that CNV-associated brain changes contribute to clinical manifestations but require additional factors for the development of the disorder. These findings highlight the power of genetic risk factors as a complement to studying groups defined by behavioral criteria.

8.
Am J Hum Genet ; 102(1): 116-132, 2018 01 04.
Artigo em Inglês | MEDLINE | ID: mdl-29290337

RESUMO

Whole-exome and targeted sequencing of 13 individuals from 10 unrelated families with overlapping clinical manifestations identified loss-of-function and missense variants in KIAA1109 allowing delineation of an autosomal-recessive multi-system syndrome, which we suggest to name Alkuraya-Kucinskas syndrome (MIM 617822). Shared phenotypic features representing the cardinal characteristics of this syndrome combine brain atrophy with clubfoot and arthrogryposis. Affected individuals present with cerebral parenchymal underdevelopment, ranging from major cerebral parenchymal thinning with lissencephalic aspect to moderate parenchymal rarefaction, severe to mild ventriculomegaly, cerebellar hypoplasia with brainstem dysgenesis, and cardiac and ophthalmologic anomalies, such as microphthalmia and cataract. Severe loss-of-function cases were incompatible with life, whereas those individuals with milder missense variants presented with severe global developmental delay, syndactyly of 2nd and 3rd toes, and severe muscle hypotonia resulting in incapacity to stand without support. Consistent with a causative role for KIAA1109 loss-of-function/hypomorphic variants in this syndrome, knockdowns of the zebrafish orthologous gene resulted in embryos with hydrocephaly and abnormally curved notochords and overall body shape, whereas published knockouts of the fruit fly and mouse orthologous genes resulted in lethality or severe neurological defects reminiscent of the probands' features.

9.
Hum Mutat ; 39(2): 281-291, 2018 02.
Artigo em Inglês | MEDLINE | ID: mdl-29193635

RESUMO

We report five individuals with loss-of-function of the X-linked AMMECR1: a girl with a balanced X-autosome translocation and inactivation of the normal X-chromosome; two boys with maternally inherited and de novo nonsense variants; and two half-brothers with maternally inherited microdeletion variants. They present with short stature, cardiac and skeletal abnormalities, and hearing loss. Variants of unknown significance in AMMECR1 in four male patients from two families with partially overlapping phenotypes were previously reported. AMMECR1 is coexpressed with genes implicated in cell cycle regulation, five of which were previously associated with growth and bone alterations. Our knockdown of the zebrafish orthologous gene resulted in phenotypes reminiscent of patients' features. The increased transcript and encoded protein levels of AMMECR1L, an AMMECR1 paralog, in the t(X;9) patient's cells indicate a possible partial compensatory mechanism. AMMECR1 and AMMECR1L proteins dimerize and localize to the nucleus as suggested by their nucleic acid-binding RAGNYA folds. Our results suggest that AMMECR1 is potentially involved in cell cycle control and linked to a new syndrome with growth, bone, heart, and kidney alterations with or without elliptocytosis.

10.
Genome Res ; 28(1): 1-10, 2018 01.
Artigo em Inglês | MEDLINE | ID: mdl-29237728

RESUMO

The majority of aneuploid fetuses are spontaneously miscarried. Nevertheless, some aneuploid individuals survive despite the strong genetic insult. Here, we investigate if the survival probability of aneuploid fetuses is affected by the genome-wide burden of slightly deleterious variants. We analyzed two cohorts of live-born Down syndrome individuals (388 genotyped samples and 16 fibroblast transcriptomes) and observed a deficit of slightly deleterious variants on Chromosome 21 and decreased transcriptome-wide variation in the expression level of highly constrained genes. We interpret these results as signatures of embryonic selection, and propose a genetic handicap model whereby an individual bearing an extremely severe deleterious variant (such as aneuploidy) could escape embryonic lethality if the genome-wide burden of slightly deleterious variants is sufficiently low. This approach can be used to study the composition and effect of the numerous slightly deleterious variants in humans and model organisms.


Assuntos
Aneuploidia , Cromossomos Humanos Par 21/genética , Síndrome de Down , Genótipo , Transcriptoma , Aborto Espontâneo , Síndrome de Down/embriologia , Síndrome de Down/genética , Feminino , Humanos , Gravidez
11.
Nat Plants ; 3(12): 926-929, 2017 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-29209081

RESUMO

Because plants do not possess a defined germline, deleterious somatic mutations can be passed to gametes, and a large number of cell divisions separating zygote from gamete formation may lead to many mutations in long-lived plants. We sequenced the genome of two terminal branches of a 234-year-old oak tree and found several fixed somatic single-nucleotide variants whose sequential appearance in the tree could be traced along nested sectors of younger branches. Our data suggest that stem cells of shoot meristems in trees are robustly protected from the accumulation of mutations.


Assuntos
Genes de Plantas , Mutação , Quercus/genética , Árvores/genética , Longevidade/genética , Meristema/citologia , Meristema/genética , Taxa de Mutação , Brotos de Planta/citologia , Brotos de Planta/genética , Polimorfismo de Nucleotídeo Único , Quercus/citologia , Árvores/citologia
12.
Am J Hum Genet ; 101(4): 564-577, 2017 Oct 05.
Artigo em Inglês | MEDLINE | ID: mdl-28965845

RESUMO

Copy-number changes in 16p11.2 contribute significantly to neuropsychiatric traits. Besides the 600 kb BP4-BP5 CNV found in 0.5%-1% of individuals with autism spectrum disorders and schizophrenia and whose rearrangement causes reciprocal defects in head size and body weight, a second distal 220 kb BP2-BP3 CNV is likewise a potent driver of neuropsychiatric, anatomical, and metabolic pathologies. These two CNVs are engaged in complex reciprocal chromatin looping, intimating a functional relationship between genes in these regions that might be relevant to pathomechanism. We assessed the drivers of the distal 16p11.2 duplication by overexpressing each of the nine encompassed genes in zebrafish. Only overexpression of LAT induced a reduction of brain proliferating cells and concomitant microcephaly. Consistently, suppression of the zebrafish ortholog induced an increase of proliferation and macrocephaly. These phenotypes were not unique to zebrafish; Lat knockout mice show brain volumetric changes. Consistent with the hypothesis that LAT dosage is relevant to the CNV pathology, we observed similar effects upon overexpression of CD247 and ZAP70, encoding members of the LAT signalosome. We also evaluated whether LAT was interacting with KCTD13, MVP, and MAPK3, major driver and modifiers of the proximal 16p11.2 600 kb BP4-BP5 syndromes, respectively. Co-injected embryos exhibited an increased microcephaly, suggesting the presence of genetic interaction. Correspondingly, carriers of 1.7 Mb BP1-BP5 rearrangements that encompass both the BP2-BP3 and BP4-BP5 loci showed more severe phenotypes. Taken together, our results suggest that LAT, besides its well-recognized function in T cell development, is a major contributor of the 16p11.2 220 kb BP2-BP3 CNV-associated neurodevelopmental phenotypes.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Transtorno Autístico/genética , Encéfalo/patologia , Transtornos Cromossômicos/genética , Cromossomos Humanos Par 16 , Variações do Número de Cópias de DNA , Deficiência Intelectual/genética , Proteínas de Membrana/genética , Microcefalia/genética , Microcefalia/patologia , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/fisiologia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Transtorno Autístico/imunologia , Transtorno Autístico/patologia , Encéfalo/metabolismo , Criança , Pré-Escolar , Deleção Cromossômica , Transtornos Cromossômicos/imunologia , Transtornos Cromossômicos/patologia , Cromossomos Humanos Par 16/genética , Cromossomos Humanos Par 16/imunologia , Estudos de Coortes , Embrião não Mamífero/metabolismo , Embrião não Mamífero/patologia , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Humanos , Lactente , Deficiência Intelectual/imunologia , Deficiência Intelectual/patologia , Masculino , Proteínas de Membrana/metabolismo , Proteínas de Membrana/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Pessoa de Meia-Idade , Fenótipo , Fosfoproteínas/fisiologia , Transdução de Sinais , Adulto Jovem , Peixe-Zebra/embriologia , Peixe-Zebra/genética , Proteínas de Peixe-Zebra/genética , Proteínas de Peixe-Zebra/metabolismo
13.
Nat Commun ; 8: 15842, 2017 07 27.
Artigo em Inglês | MEDLINE | ID: mdl-28748955

RESUMO

The enormous variation in human lifespan is in part due to a myriad of sequence variants, only a few of which have been revealed to date. Since many life-shortening events are related to diseases, we developed a Mendelian randomization-based method combining 58 disease-related GWA studies to derive longevity priors for all HapMap SNPs. A Bayesian association scan, informed by these priors, for parental age of death in the UK Biobank study (n=116,279) revealed 16 independent SNPs with significant Bayes factor at a 5% false discovery rate (FDR). Eleven of them replicate (5% FDR) in five independent longevity studies combined; all but three are depleted of the life-shortening alleles in older Biobank participants. Further analysis revealed that brain expression levels of nearby genes (RBM6, SULT1A1 and CHRNA5) might be causally implicated in longevity. Gene expression and caloric restriction experiments in model organisms confirm the conserved role for RBM6 and SULT1A1 in modulating lifespan.

14.
J Pediatr Genet ; 6(2): 98-102, 2017 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-28496997

RESUMO

Split hand/foot malformation with long bone deficiency (SHFLD) is a congenital limb anomaly where hands and/or feet cleft and syndactyly are associated with long bone defects, usually involving the tibia. Previously published data reported that 17p13.3 chromosomal duplication, including the BHLHA9 gene, has been associated with the distinct entity, termed SHFLD3 (OMIM 612576), inherited as an autosomal dominant trait. Here, we present a family with three members affected by SHFLD harboring BHLHA9 duplication. We exploited in vitro differentiation system to promote proband's skin fibroblasts toward osteoblastic lineage, and we observed a slight but consistent delay in the mineralization pattern. This result possibly suggests an impairment of the osteogenic process in the affected members.

15.
BMC Genomics ; 18(1): 144, 2017 02 10.
Artigo em Inglês | MEDLINE | ID: mdl-28183275

RESUMO

BACKGROUND: Copy number variations (CNVs) are a significant source of genetic diversity and commonly found in mammalian genomes. We have generated a genome-wide CNV map for Cynomolgus monkeys (Macaca fascicularis). This crab-eating macaque is the closest animal model to humans that is used in biomedical research. RESULTS: We show that Cynomolgus monkey CNVs are in general much smaller in size than gene loci and are specific to the population of origin. Genome-wide expression data from five vitally important organs demonstrates that CNVs in close proximity to transcription start sites associate strongly with expression changes. Among these eQTL genes we find an overrepresentation of genes involved in metabolism, receptor activity, and transcription. CONCLUSION: These results provide evidence that CNVs shape tissue transcriptomes in monkey populations, potentially offering an adaptive advantage. We suggest that this genetic diversity should be taken into account when using Cynomolgus macaques as models.


Assuntos
Variações do Número de Cópias de DNA , Macaca fascicularis/genética , Animais , Perfilação da Expressão Gênica , Genótipo , Locos de Características Quantitativas/genética
16.
Eur J Med Genet ; 60(3): 154-158, 2017 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-28089741

RESUMO

Aicardi-Goutières syndrome (AGS) is an inflammatory disorder belonging to the recently characterized group of type I interferonopathies. The most consistently affected tissues in AGS are the central nervous system and skin, but various organ systems and tissues have been reported to be affected, pointing to the systemic nature of the disease. Here we describe a patient with AGS due to a homozygous p.Arg114His mutation in the TREX1 gene. The histologically proven inflammatory myopathy in our patient expands the range of clinical features of AGS. Histological signs of muscle biopsies in the proband, and in two other AGS patients described earlier, are similar to those seen in various autoimmune myositises and could be ascribed to inapproapriate IFN I activation. In view of signs of possible mitochondrial damage in AGS, we propose that mitochondrial DNA could be a trigger of autoimmune responses in AGS.


Assuntos
Doenças Autoimunes do Sistema Nervoso/genética , Doenças Autoimunes do Sistema Nervoso/patologia , Exodesoxirribonucleases/genética , Interferon Tipo I/imunologia , Mitocôndrias/patologia , Miosite/patologia , Malformações do Sistema Nervoso/genética , Malformações do Sistema Nervoso/patologia , Fosfoproteínas/genética , Sequência de Bases , Criança , DNA Mitocondrial/genética , Feminino , Predisposição Genética para Doença , Humanos , Mitocôndrias/genética , Análise de Sequência de DNA
17.
Genome Med ; 8(1): 105, 2016 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-27799067

RESUMO

BACKGROUND: Smith-Magenis syndrome (SMS) is a developmental disability/multiple congenital anomaly disorder resulting from haploinsufficiency of RAI1. It is characterized by distinctive facial features, brachydactyly, sleep disturbances, and stereotypic behaviors. METHODS: We investigated a cohort of 15 individuals with a clinical suspicion of SMS who showed neither deletion in the SMS critical region nor damaging variants in RAI1 using whole exome sequencing. A combination of network analysis (co-expression and biomedical text mining), transcriptomics, and circularized chromatin conformation capture (4C-seq) was applied to verify whether modified genes are part of the same disease network as known SMS-causing genes. RESULTS: Potentially deleterious variants were identified in nine of these individuals using whole-exome sequencing. Eight of these changes affect KMT2D, ZEB2, MAP2K2, GLDC, CASK, MECP2, KDM5C, and POGZ, known to be associated with Kabuki syndrome 1, Mowat-Wilson syndrome, cardiofaciocutaneous syndrome, glycine encephalopathy, mental retardation and microcephaly with pontine and cerebellar hypoplasia, X-linked mental retardation 13, X-linked mental retardation Claes-Jensen type, and White-Sutton syndrome, respectively. The ninth individual carries a de novo variant in JAKMIP1, a regulator of neuronal translation that was recently found deleted in a patient with autism spectrum disorder. Analyses of co-expression and biomedical text mining suggest that these pathologies and SMS are part of the same disease network. Further support for this hypothesis was obtained from transcriptome profiling that showed that the expression levels of both Zeb2 and Map2k2 are perturbed in Rai1 -/- mice. As an orthogonal approach to potentially contributory disease gene variants, we used chromatin conformation capture to reveal chromatin contacts between RAI1 and the loci flanking ZEB2 and GLDC, as well as between RAI1 and human orthologs of the genes that show perturbed expression in our Rai1 -/- mouse model. CONCLUSIONS: These holistic studies of RAI1 and its interactions allow insights into SMS and other disorders associated with intellectual disability and behavioral abnormalities. Our findings support a pan-genomic approach to the molecular diagnosis of a distinctive disorder.


Assuntos
Exoma/genética , Redes Reguladoras de Genes , Genômica/métodos , Mutação/genética , Síndrome de Smith-Magenis/genética , Fatores de Transcrição/fisiologia , Transcriptoma , Animais , Embrião de Mamíferos/citologia , Embrião de Mamíferos/metabolismo , Feminino , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fenótipo , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa
19.
Nat Commun ; 7: 12339, 2016 08 17.
Artigo em Inglês | MEDLINE | ID: mdl-27531712

RESUMO

Long non-coding RNAs (lncRNAs) constitute a large, yet mostly uncharacterized fraction of the mammalian transcriptome. Such characterization requires a comprehensive, high-quality annotation of their gene structure and boundaries, which is currently lacking. Here we describe RACE-Seq, an experimental workflow designed to address this based on RACE (rapid amplification of cDNA ends) and long-read RNA sequencing. We apply RACE-Seq to 398 human lncRNA genes in seven tissues, leading to the discovery of 2,556 on-target, novel transcripts. About 60% of the targeted loci are extended in either 5' or 3', often reaching genomic hallmarks of gene boundaries. Analysis of the novel transcripts suggests that lncRNAs are as long, have as many exons and undergo as much alternative splicing as protein-coding genes, contrary to current assumptions. Overall, we show that RACE-Seq is an effective tool to annotate an organism's deep transcriptome, and compares favourably to other targeted sequencing techniques.


Assuntos
Sequenciamento de Nucleotídeos em Larga Escala/métodos , Reação em Cadeia da Polimerase/métodos , RNA Longo não Codificante/genética , Análise de Sequência de RNA/métodos , Éxons/genética , Loci Gênicos , Humanos , Anotação de Sequência Molecular , Especificidade de Órgãos/genética , Estudo de Prova de Conceito , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Sítios de Splice de RNA/genética , RNA Longo não Codificante/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Transcriptoma/genética
20.
Nature ; 536(7615): 205-9, 2016 08 11.
Artigo em Inglês | MEDLINE | ID: mdl-27487209

RESUMO

Genetic differences that specify unique aspects of human evolution have typically been identified by comparative analyses between the genomes of humans and closely related primates, including more recently the genomes of archaic hominins. Not all regions of the genome, however, are equally amenable to such study. Recurrent copy number variation (CNV) at chromosome 16p11.2 accounts for approximately 1% of cases of autism and is mediated by a complex set of segmental duplications, many of which arose recently during human evolution. Here we reconstruct the evolutionary history of the locus and identify bolA family member 2 (BOLA2) as a gene duplicated exclusively in Homo sapiens. We estimate that a 95-kilobase-pair segment containing BOLA2 duplicated across the critical region approximately 282 thousand years ago (ka), one of the latest among a series of genomic changes that dramatically restructured the locus during hominid evolution. All humans examined carried one or more copies of the duplication, which nearly fixed early in the human lineage--a pattern unlikely to have arisen so rapidly in the absence of selection (P < 0.0097). We show that the duplication of BOLA2 led to a novel, human-specific in-frame fusion transcript and that BOLA2 copy number correlates with both RNA expression (r = 0.36) and protein level (r = 0.65), with the greatest expression difference between human and chimpanzee in experimentally derived stem cells. Analyses of 152 patients carrying a chromosome 16p11. rearrangement show that more than 96% of breakpoints occur within the H. sapiens-specific duplication. In summary, the duplicative transposition of BOLA2 at the root of the H. sapiens lineage about 282 ka simultaneously increased copy number of a gene associated with iron homeostasis and predisposed our species to recurrent rearrangements associated with disease.


Assuntos
Cromossomos Humanos Par 16/genética , Variações do Número de Cópias de DNA/genética , Evolução Molecular , Predisposição Genética para Doença , Proteínas/genética , Animais , Transtorno Autístico/genética , Quebra Cromossômica , Duplicação Gênica , Homeostase/genética , Humanos , Ferro/metabolismo , Pan troglodytes/genética , Pongo/genética , Proteínas/análise , Recombinação Genética , Especificidade da Espécie , Fatores de Tempo
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