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1.
PLoS Genet ; 9(10): e1003823, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-24098143

RESUMO

Agenesis of the corpus callosum (ACC), cerebellar hypoplasia (CBLH), and polymicrogyria (PMG) are severe congenital brain malformations with largely undiscovered causes. We conducted a large-scale chromosomal copy number variation (CNV) discovery effort in 255 ACC, 220 CBLH, and 147 PMG patients, and 2,349 controls. Compared to controls, significantly more ACC, but unexpectedly not CBLH or PMG patients, had rare genic CNVs over one megabase (p = 1.48×10⁻³; odds ratio [OR] = 3.19; 95% confidence interval [CI] = 1.89-5.39). Rare genic CNVs were those that impacted at least one gene in less than 1% of the combined population of patients and controls. Compared to controls, significantly more ACC but not CBLH or PMG patients had rare CNVs impacting over 20 genes (p = 0.01; OR = 2.95; 95% CI = 1.69-5.18). Independent qPCR confirmation showed that 9.4% of ACC patients had de novo CNVs. These, in comparison to inherited CNVs, preferentially overlapped de novo CNVs previously observed in patients with autism spectrum disorders (p = 3.06×10⁻4; OR = 7.55; 95% CI = 2.40-23.72). Interestingly, numerous reports have shown a reduced corpus callosum area in autistic patients, and diminished social and executive function in many ACC patients. We also confirmed and refined previously known CNVs, including significantly narrowing the 8p23.1-p11.1 duplication present in 2% of our current ACC cohort. We found six novel CNVs, each in a single patient, that are likely deleterious: deletions of 1p31.3-p31.1, 1q31.2-q31.3, 5q23.1, and 15q11.2-q13.1; and duplications of 2q11.2-q13 and 11p14.3-p14.2. One ACC patient with microcephaly had a paternally inherited deletion of 16p13.11 that included NDE1. Exome sequencing identified a recessive maternally inherited nonsense mutation in the non-deleted allele of NDE1, revealing the complexity of ACC genetics. This is the first systematic study of CNVs in congenital brain malformations, and shows a much higher prevalence of large gene-rich CNVs in ACC than in CBLH and PMG.


Assuntos
Agenesia do Corpo Caloso/genética , Cerebelo/anormalidades , Variações do Número de Cópias de DNA , Malformações do Desenvolvimento Cortical/genética , Malformações do Sistema Nervoso/genética , Adolescente , Adulto , Agenesia do Corpo Caloso/patologia , Cerebelo/patologia , Criança , Pré-Escolar , Deficiências do Desenvolvimento/genética , Deficiências do Desenvolvimento/patologia , Feminino , Genoma Humano , Estudo de Associação Genômica Ampla , Humanos , Lactente , Recém-Nascido , Masculino , Malformações do Desenvolvimento Cortical/patologia , Pessoa de Meia-Idade , Malformações do Sistema Nervoso/patologia , Polimorfismo de Nucleotídeo Único
2.
Nature ; 452(7190): 961-5, 2008 Apr 24.
Artigo em Inglês | MEDLINE | ID: mdl-18432238

RESUMO

Escherichia coli AlkB and its human homologues ABH2 and ABH3 repair DNA/RNA base lesions by using a direct oxidative dealkylation mechanism. ABH2 has the primary role of guarding mammalian genomes against 1-meA damage by repairing this lesion in double-stranded DNA (dsDNA), whereas AlkB and ABH3 preferentially repair single-stranded DNA (ssDNA) lesions and can repair damaged bases in RNA. Here we show the first crystal structures of AlkB-dsDNA and ABH2-dsDNA complexes, stabilized by a chemical cross-linking strategy. This study reveals that AlkB uses an unprecedented base-flipping mechanism to access the damaged base: it squeezes together the two bases flanking the flipped-out one to maintain the base stack, explaining the preference of AlkB for repairing ssDNA lesions over dsDNA ones. In addition, the first crystal structure of ABH2, presented here, provides a structural basis for designing inhibitors of this human DNA repair protein.


Assuntos
Enzimas Reparadoras do DNA/química , DNA/metabolismo , Dioxigenases/química , Dioxigenases/metabolismo , Proteínas de Escherichia coli/química , Proteínas de Escherichia coli/metabolismo , Oxigenases de Função Mista/química , Oxigenases de Função Mista/metabolismo , RNA/metabolismo , Adenina/análogos & derivados , Adenina/metabolismo , Homólogo AlkB 2 da Dioxigenase Dependente de alfa-Cetoglutarato , Sítios de Ligação , Reagentes de Ligações Cruzadas/química , Cristalografia por Raios X , DNA/química , Dano ao DNA , Reparo do DNA , Enzimas Reparadoras do DNA/metabolismo , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/metabolismo , Humanos , Modelos Moleculares , Ligação Proteica
3.
Nat Genet ; 44(4): 440-4, S1-2, 2012 Feb 26.
Artigo em Inglês | MEDLINE | ID: mdl-22366783

RESUMO

Brain malformations are individually rare but collectively common causes of developmental disabilities. Many forms of malformation occur sporadically and are associated with reduced reproductive fitness, pointing to a causative role for de novo mutations. Here, we report a study of Baraitser-Winter syndrome, a well-defined disorder characterized by distinct craniofacial features, ocular colobomata and neuronal migration defect. Using whole-exome sequencing of three proband-parent trios, we identified de novo missense changes in the cytoplasmic actin-encoding genes ACTB and ACTG1 in one and two probands, respectively. Sequencing of both genes in 15 additional affected individuals identified disease-causing mutations in all probands, including two recurrent de novo alterations (ACTB, encoding p.Arg196His, and ACTG1, encoding p.Ser155Phe). Our results confirm that trio-based exome sequencing is a powerful approach to discover genes causing sporadic developmental disorders, emphasize the overlapping roles of cytoplasmic actin proteins in development and suggest that Baraitser-Winter syndrome is the predominant phenotype associated with mutation of these two genes.


Assuntos
Anormalidades Múltiplas/genética , Actinas/genética , Encéfalo/anormalidades , Adolescente , Adulto , Sequência de Aminoácidos , Sequência de Bases , Criança , Coloboma/genética , Variações do Número de Cópias de DNA , Deficiências do Desenvolvimento/genética , Feminino , Humanos , Deficiência Intelectual/genética , Masculino , Dados de Sequência Molecular , Mutação de Sentido Incorreto , Malformações do Sistema Nervoso/genética , Fator de Transcrição PAX9/genética , Alinhamento de Sequência , Análise de Sequência de DNA , Deleção de Sequência , Síndrome
4.
Nat Genet ; 44(8): 934-40, 2012 Jun 24.
Artigo em Inglês | MEDLINE | ID: mdl-22729224

RESUMO

Megalencephaly-capillary malformation (MCAP) and megalencephaly-polymicrogyria-polydactyly-hydrocephalus (MPPH) syndromes are sporadic overgrowth disorders associated with markedly enlarged brain size and other recognizable features. We performed exome sequencing in 3 families with MCAP or MPPH, and our initial observations were confirmed in exomes from 7 individuals with MCAP and 174 control individuals, as well as in 40 additional subjects with megalencephaly, using a combination of Sanger sequencing, restriction enzyme assays and targeted deep sequencing. We identified de novo germline or postzygotic mutations in three core components of the phosphatidylinositol 3-kinase (PI3K)-AKT pathway. These include 2 mutations in AKT3, 1 recurrent mutation in PIK3R2 in 11 unrelated families with MPPH and 15 mostly postzygotic mutations in PIK3CA in 23 individuals with MCAP and 1 with MPPH. Our data highlight the central role of PI3K-AKT signaling in vascular, limb and brain development and emphasize the power of massively parallel sequencing in a challenging context of phenotypic and genetic heterogeneity combined with postzygotic mosaicism.


Assuntos
Malformações do Desenvolvimento Cortical/genética , Megalencefalia/genética , Mutação , Fosfatidilinositol 3-Quinases/genética , Proteínas Proto-Oncogênicas c-akt/genética , Classe I de Fosfatidilinositol 3-Quinases , Exoma , Mutação em Linhagem Germinativa , Humanos , Hidrocefalia/enzimologia , Hidrocefalia/genética , Hidrocefalia/patologia , Malformações do Desenvolvimento Cortical/enzimologia , Malformações do Desenvolvimento Cortical/patologia , Megalencefalia/enzimologia , Megalencefalia/patologia , Mutação de Sentido Incorreto , Síndrome
6.
Virology ; 302(2): 405-12, 2002 Oct 25.
Artigo em Inglês | MEDLINE | ID: mdl-12441084

RESUMO

Retroviruses require a balance of spliced and unspliced RNA for efficient replication. Here, we examined the effect of mutations in a splicing suppressor sequence called the negative regulator of splicing (NRS), located within the gag gene of Rous sarcoma virus. While the NRS mutant viruses showed only small changes in the levels of spliced env mRNAs, they had significant increases in src mRNA levels and transformed cells more efficiently than wild-type virus. None of these mutations prevented viral replication; however, some of the mutant viruses replicated more slowly than wild-type virus. In addition, increased transcriptional readthrough of the poly(A) site in the 3' LTR was observed with the NRS mutant viruses, suggesting that the wild-type NRS sequence promotes polyadenylation.


Assuntos
Vírus do Sarcoma Aviário/genética , Elementos Facilitadores Genéticos , Regulação Viral da Expressão Gênica , Genes src/genética , Poliadenilação , Splicing de RNA , Animais , Células Cultivadas , Embrião de Galinha , Fibroblastos , Genes gag , Mutação Puntual , RNA Mensageiro/metabolismo , RNA Viral/metabolismo
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