RESUMO
INTRODUCTION: Array comparative genomic hybridisation (array CGH) is a powerful method that detects alteration of gene copy number with greater resolution and efficiency than traditional methods. However, its ability to detect disease causing duplications in constitutional genomic DNA has not been shown. We developed an array CGH assay for X linked hypopituitarism, which is associated with duplication of Xq26-q27. METHODS: We generated custom BAC/PAC arrays that spanned the 7.3 Mb critical region at Xq26.1-q27.3, and used them to search for duplications in three previously uncharacterised families with X linked hypopituitarism. RESULTS: Validation experiments clearly identified Xq26-q27 duplications that we had previously mapped by fluorescence in situ hybridisation. Array CGH analysis of novel XH families identified three different Xq26-q27 duplications, which together refine the critical region to a 3.9 Mb interval at Xq27.2-q27.3. Expression analysis of six orthologous mouse genes from this region revealed that the transcription factor Sox3 is expressed at 11.5 and 12.5 days after conception in the infundibulum of the developing pituitary and the presumptive hypothalamus. DISCUSSION: Array CGH is a robust and sensitive method for identifying X chromosome duplications. The existence of different, overlapping Xq duplications in five kindreds indicates that X linked hypopituitarism is caused by increased gene dosage. Interestingly, all X linked hypopituitarism duplications contain SOX3. As mutation of this gene in human beings and mice results in hypopituitarism, we hypothesise that increased dosage of Sox3 causes perturbation of pituitary and hypothalamic development and may be the causative mechanism for X linked hypopituitarism.
Assuntos
Cromossomos Humanos X/genética , Proteínas de Ligação a DNA/genética , Duplicação Gênica , Genes Duplicados/genética , Doenças Genéticas Ligadas ao Cromossomo X/genética , Proteínas de Grupo de Alta Mobilidade/genética , Hipopituitarismo/genética , Fatores de Transcrição/genética , Adolescente , Adulto , Animais , Criança , Pré-Escolar , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Ligação Genética/genética , Genoma Humano , Humanos , Hipotálamo/embriologia , Hipotálamo/metabolismo , Hibridização in Situ Fluorescente , Lactente , Recém-Nascido , Masculino , Camundongos , Hibridização de Ácido Nucleico , Linhagem , Hipófise/embriologia , Hipófise/metabolismo , Reprodutibilidade dos Testes , Fatores de Transcrição SOXB1RESUMO
Mutations of the flavin-containing monooxygenase type 3 gene (FMO3) that encode the major functional form present in adult human liver, have been shown to cause trimethylaminuria. We now report a novel homozygous deletion of exons 1 and 2 in an Australian of Greek ancestry with TMAuria, the first report of a deletion causative of trimethylaminuria. The deletion occurs 328 bp upstream from exon 1. The 3'-end of the deletion occurs in intron 2, 10013 base pairs downstream from the end of exon 2. The deletion is 12226 bp long. For the proband homozygous for the human FMO3 gene deletion, it is predicted that in addition to loss of monooxygenase function for human FMO3 substrates, such as TMA and other amines, the proband will exhibit decreased tolerance of biogenic amines, both medicinal and those found in foods.
Assuntos
Deleção de Genes , Erros Inatos do Metabolismo/genética , Metilaminas/urina , Oxigenases/genética , Adolescente , Sequência de Bases , Southern Blotting , Primers do DNA , Éxons , Grécia , Homozigoto , Humanos , Masculino , Erros Inatos do Metabolismo/enzimologia , Reação em Cadeia da PolimeraseRESUMO
BACKGROUND: Spinocerebellar ataxia type 2 (SCA2) is a recently delineated cause of autosomal dominant cerebellar ataxia type I. The basic clinical neurologic features of SCA2 have been described in the literature, but neuropsychological features have not, despite statements that some patients became demented. OBJECTIVE: To describe the clinical and neuropsychological features of patients from a pedigree with SCA2. PATIENTS AND METHODS: We studied 8 affected members of an Australian pedigree of northern Italian origin with autosomal dominant cerebellar ataxia type I caused by SCA2. Patients underwent clinical neurologic examination and abbreviated neuropsychological testing, while some also underwent magnetic resonance imaging. The results were compared with pooled results from previously published studies of patients with SCA2. RESULTS: The pedigree displayed anticipation, with earlier onset in later generations, and there was an inverse correlation between repeat number and age at onset. The principal difference from other clinical reports of SCA2 was our finding of unequivocal frontal-executive dysfunction in 5 of 6 individuals who could be tested quantitatively, despite Mini-Mental State Examination scores in the nondemented range. This feature did not appear to correlate with either repeat size or duration of illness. CONCLUSIONS: In light of a recent report of frontal-executive dysfunction in spinocerebellar ataxia type III, we postulate that this pattern may be common to the autosomal dominant cerebellar ataxias and frequently may be overlooked because of the insensitivity of routine screening tests such as the Mini-Mental State Examination.
Assuntos
Lobo Frontal/fisiopatologia , Degenerações Espinocerebelares/fisiopatologia , Adulto , Idade de Início , Movimentos Oculares/fisiologia , Feminino , Humanos , Imageamento por Ressonância Magnética , Masculino , Pessoa de Meia-Idade , Testes Neuropsicológicos , Linhagem , Reflexo Vestíbulo-Ocular/fisiologia , Degenerações Espinocerebelares/genética , Degenerações Espinocerebelares/psicologia , Repetições de TrinucleotídeosRESUMO
A kindred is described with a dominantly inherited "pure" cerebellar ataxia in which the currently known spinocerebellar ataxias have been excluded. In the eight subjects studied, a notable clinical feature is slow progression, with the three least affected having only a mild degree of gait ataxia after three or more decades of disease duration. Pending an actual chromosomal locus discovery, the name spinocerebellar ataxia (SCA)15 is expectantly applied.
Assuntos
Aberrações Cromossômicas , Genes Dominantes , Ataxias Espinocerebelares/genética , Adulto , Idoso , Atrofia , Cerebelo/patologia , Mapeamento Cromossômico , Feminino , Ligação Genética , Genótipo , Humanos , Imageamento por Ressonância Magnética , Masculino , Pessoa de Meia-Idade , Exame Neurológico , Linhagem , Ataxias Espinocerebelares/diagnósticoRESUMO
The frequencies of various genetically defined spinocerebellar ataxias (SCAs) vary in different populations presumably due to founder effects. No data have been published on the Australian population. Although predominantly of Anglo-Celtic extraction, Australia has also received considerable influx from southeastern Europe and more recently eastern and southeastern Asia. We examined the frequency of mutations for SCA types 1, 2, 3, 6, and 7 in southeastern Australia. Of 88 pedigrees with multiple-affected members, SCA type 1 (SCA1) accounted for 16%, SCA2 for 6%, SCA3 for 12%, SCA6 for 17%, SCA7 for 2%, and 47% (41 pedigrees) were negative for each of SCA1, 2, 3, and 6. Twenty of the 41 negative pedigrees were also negative for dentatorubralpallidoluysian atrophy, and indeed dentatorubralpallidoluysian atrophy has not been reported in Australia. In addition, no pedigree information was available on a further four patients with SCA1, three patients with SCA2, three patients with SCA3, and three patients with SCA6. One SCA1 and two SCA2 patients had no other known affected family members. In total, of 63 pedigrees or individuals with positive tests, 30% were those with SCA1, 15% with SCA2, 22% with SCA3, 30% with SCA6, and 3% with SCA7. Judging by pedigree names, four of the nine SCA2 positive individuals/pedigrees were of Italian extraction, and four of the 14 SCA3 positive individuals/pedigrees were of Chinese descent, whereas only 1 of the 20 SCA1 positive individuals/pedigrees were non-Anglo-Celtic. These results are in accordance with the known ethnic composition of the Australian population and with gene frequencies in these constituent ethnic groups reported by others. The frequency of large-normal alleles for SCA1 and SCA3 in the population reflects the prevalence of these two diseases, supporting the hypothesis that disease alleles arise by expansion of large-normal alleles.
Assuntos
Frequência do Gene , Ataxias Espinocerebelares/epidemiologia , Ataxias Espinocerebelares/genética , Alelos , Efeito Fundador , Genótipo , Humanos , New South Wales/epidemiologia , Linhagem , Prevalência , Ataxias Espinocerebelares/classificação , Ataxias Espinocerebelares/diagnóstico , Tasmânia/epidemiologia , Repetições de Trinucleotídeos/genética , Vitória/epidemiologiaRESUMO
We have characterized deletions of the dystrophin gene in patients suffering from relatively mild muscular dystrophy. Our data show that most of the Becker muscular dystrophy (BMD) patients have intragenic deletions which leave the protein reading frame in phase. Remarkably, large deletions of the region corresponding to the central triple helical repeats in the protein can result in an exceptionally mild phenotype. Three brothers suffering from BMD, glycerol kinase deficiency, and adrenal hypoplasia possess a deletion at the 3' end of the gene. They also display developmental delay. Thus the 3' processing of the gene must be necessary for the correct function of the dystrophin molecule.
Assuntos
Deleção Cromossômica , Distrofina/genética , Distrofias Musculares/genética , Glândulas Suprarrenais/anormalidades , Mapeamento Cromossômico , Glicerol Quinase/deficiência , Humanos , Masculino , Distrofias Musculares/metabolismo , Distrofias Musculares/patologia , Linhagem , FenótipoRESUMO
Friedreich ataxia is an autosomal recessive disorder caused by mutations in the FRDA gene that encodes a 210-amino acid protein called frataxin. An expansion of a GAA trinucleotide repeat in intron 1 of the gene is present in more than 95% of mutant alleles. Of the 83 people we studied who have mutations in FRDA, 78 are homozygous for an expanded GAA repeat; the other five patients have an expansion in one allele and a point mutation in the other. Here we present a detailed clinical and genetic study of a subset of 51 patients homozygous for an expansion of the GAA repeat. We found a correlation between the size of the smaller of the two expanded alleles and age at onset, age into wheelchair, scoliosis, impaired vibration sense, and the presence of foot deformity. There was no significant correlation between the size of the smaller allele and cardiomyopathy, diabetes mellitus, loss of proprioception, or bladder symptoms. The larger allele size correlated with bladder symptoms and the presence of foot deformity. The duration of disease is correlated with wheelchair use and the presence of diabetes, scoliosis, bladder symptoms and impaired proprioception, and vibration sense but no other complications studied.
Assuntos
Ataxia de Friedreich/genética , Proteínas de Ligação ao Ferro , Fosfotransferases (Aceptor do Grupo Álcool)/genética , Expansão das Repetições de Trinucleotídeos/genética , Adolescente , Adulto , Idade de Início , Austrália , Cardiomiopatias/genética , Criança , Pré-Escolar , Consanguinidade , Diabetes Mellitus/genética , Europa (Continente)/etnologia , Feminino , Ataxia de Friedreich/patologia , Ataxia de Friedreich/fisiopatologia , Homozigoto , Humanos , Lactente , Íntrons/genética , Masculino , Pessoa de Meia-Idade , Doenças Musculoesqueléticas/genética , Propriocepção/genética , Expansão das Repetições de Trinucleotídeos/fisiologia , Doenças da Bexiga Urinária/genética , Cadeiras de Rodas , FrataxinaRESUMO
The attenuated form of familial adenomatous polyposis coli (AAPC) is associated with mutations in the adenomatous polyposis coli (APC) gene which cluster in the 5' region of the gene. It has been proposed that a 'genotype-phenotype boundary' exists at codons 159-163, and mutations that are 5' of this boundary will produce AAPC. Herein we document a three-generation family with an exon 3 mutation well to the 5' side of the proposed boundary, in which two affected individuals have had, in their 40s, a profuse form of familial adenomatous polyposis coli. We conclude that the codon 159-163 'boundary' is indicative rather than definitive. These two patients also had postoperative intra-abdominal adhesions, severely so in one.
Assuntos
Polipose Adenomatosa do Colo/genética , Códon/genética , Genes APC , Idoso , Éxons/genética , Genótipo , Humanos , Masculino , Linhagem , FenótipoRESUMO
Mutations in the presenilin 1 (PS1) gene are responsible for approximately 50% of early onset autosomal-dominant Alzheimer's disease cases. A PCR based mutation detection method, chemical cleavage of mismatch, was used to detect a novel PS1 mutation in the coding sequence of the PS1 gene. Sequencing confirmed a T to C transition altering a leucine to proline at codon 219 of the PS1 gene. This is a novel mutation in exon 7 of the PS1 gene occurring outside the transmembrane regions of IV and V.
Assuntos
Doença de Alzheimer/epidemiologia , Doença de Alzheimer/genética , Códon/genética , Proteínas de Membrana/genética , Mutação de Sentido Incorreto/genética , Idade de Início , Idoso , Substituição de Aminoácidos , DNA/genética , Análise Mutacional de DNA , Éxons/genética , Feminino , Genoma , Humanos , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase , Presenilina-1RESUMO
Pyrolysis of solid complexes of aromatic diazo compounds with cyclomaltoheptaose (beta-cyclodextrin) yields either derivatives via insertion of carbene into hydroxyl groups. The distribution of the 2-, 3-, and 6-O-isomers indicates that the regioselectivity is moderate. The guest geometry is not as important as its size in determining the ratios of regioisomers. The origins of the regioselectivity are discussed.
Assuntos
Ciclodextrinas/química , Compostos de Diazônio/química , beta-Ciclodextrinas , Configuração de Carboidratos , Indicadores e Reagentes , Espectroscopia de Ressonância Magnética , Estrutura Molecular , Relação Estrutura-AtividadeAssuntos
Inversão Cromossômica , Cromossomos Humanos Par 3/genética , Deficiências do Desenvolvimento/genética , Transtornos Mentais/genética , Adulto , Transtorno do Deficit de Atenção com Hiperatividade/genética , Transtorno do Deficit de Atenção com Hiperatividade/psicologia , Criança , Deficiências do Desenvolvimento/psicologia , Feminino , Humanos , Cariotipagem , Masculino , Transtornos Mentais/psicologiaRESUMO
Friedreich ataxia, an autosomal recessive neurodegenerative disease, is the most common of the inherited ataxias. The recent discovery of the gene that is mutated in this condition, FRDA, has led to rapid advances in the understanding of the pathogenesis of Friedreich ataxia. About 98% of mutant alleles have an expansion of a GAA trinucleotide repeat in intron 1 of the gene. This leads to reduced levels of the protein, frataxin. There is mounting evidence to suggest that Friedreich ataxia is the result of accumulation of iron in mitochondria leading to excess production of free radicals, which then results in cellular damage and death. Currently there is no known treatment that alters the natural course of the disease. The discovery of the FRDA gene and its possible function has raised hope that rational therapeutic strategies will be developed.
Assuntos
Cromossomos Humanos Par 9 , Ataxia de Friedreich/genética , Proteínas do Tecido Nervoso/genética , Mutação Puntual , Proteínas Adaptadoras de Transdução de Sinal , Mapeamento Cromossômico , Ataxia de Friedreich/metabolismo , Humanos , Ferro/metabolismo , Mitocôndrias/metabolismoRESUMO
Although several genes that determine left-right asymmetry for structural syndromes such as situs inversus have been characterised in recent years (Supp, Witte, Potter, & Brueckner, 1997), there has been little progress in determining which genes or loci predispose to left-right handedness in humans. Linkage analysis has been used widely for the localisation of genes followed by their positional cloning. The complex genetics of handedness is one of the greatest problems for standard linkage analysis. Several genetic models have been proposed for the inheritance of handedness in humans. On the basis of these models, left-handedness can be considered a common single gene trait with a high gene frequency and a non-mendelian inheritance pattern. We report here a possible strategy, using these genetic models, that can be applied for the identification for genes determining handedness in humans.
RESUMO
Illegitimately transcribed phenylalanine hydroxylase mRNA was amplified using the polymerase chain reaction from both fibroblasts and Epstein-Barr virus-transformed lymphocytes. This method was used to study mutations of this gene in patients with phenylketonuria and known point mutations were easily detected. Illegitimate transcription was successful for studying splicing defects and it was found that the previously described mutation which changes G to A at the 5' donor site of intron 7 causes exon 7 to be spliced out.
Assuntos
Mutação , Fenilalanina Hidroxilase/genética , Fenilcetonúrias/genética , Transcrição Gênica , Sequência de Aminoácidos , Sequência de Bases , DNA/genética , DNA/isolamento & purificação , Éxons , Triagem de Portadores Genéticos , Humanos , Íntrons , Dados de Sequência Molecular , Oligodesoxirribonucleotídeos , Fenilcetonúrias/diagnóstico , Fenilcetonúrias/enzimologia , Reação em Cadeia da Polimerase/métodos , Splicing de RNA , Valores de Referência , Mapeamento por RestriçãoRESUMO
The most common mutation in alpha-1-antitrypsin deficiency, conversion of a G to an A at base 9989 (PI-Z), was detected with the chemical cleavage of mismatch method, demonstrating the power of the method for prenatal diagnosis. Exon V of the gene was amplified using the polymerase chain reaction and heteroduplexes were formed to test for the presence of the mutation. The predicted C mismatch was readily detectable with hydroxylamine, and by making the probe from the chorionic villus sample it was possible to determine that the fetus was heterozygous, not homozygous, for the mutation.
Assuntos
Diagnóstico Pré-Natal/métodos , Deficiência de alfa 1-Antitripsina , Autorradiografia , Sequência de Bases , Amostra da Vilosidade Coriônica , Feminino , Humanos , Hidroxilamina , Hidroxilaminas , Dados de Sequência Molecular , Mutação , Reação em Cadeia da Polimerase , Gravidez , alfa 1-Antitripsina/genéticaRESUMO
Flavin-containing monooxygenase form 3 (FMO3) is one of the major enzyme systems that protect humans from the potentially toxic properties of drugs and chemicals. FMO3 converts nucleophilic heteroatom-containing chemicals and endogenous materials to polar metabolites, which facilitates their elimination. For example, the tertiary amine trimethylamine is N-oxygenated by human FMO3 to trimethylamine N-oxide, and trimethylamine N-oxide is excreted in a detoxication and deoderation process. In normal humans, virtually all trimethylamine is metabolized to trimethylamine N-oxide. In a few humans, trimethylamine is not efficiently metabolized to trimethylamine N-oxide, and those individuals suffer from trimethylaminuria, or fishlike odor syndrome. Previously, we identified mutations of the FMO3 gene that cause trimethylaminuria. We now report two prevalent polymorphisms of this gene (K158E and V257M) that modulate the activity of human FMO3. These polymorphisms are widely distributed in Canadian and Australian white populations. In vitro analysis of wild-type and variant human FMO3 proteins expressed from the cDNA for the two naturally occurring polymorphisms showed differences in substrate affinities for nitrogen-containing substrates. Thus, for polymorphic forms of human FMO3, lower k(cat)/K(m) values for N-oxygenation of 10-(N, N-dimethylaminopentyl)-2-(trifluoromethyl) phenothiazine, trimethylamine, and tyramine were observed. On the basis of in vitro kinetic parameters, human FMO1 does not significantly contribute to human metabolism of trimethylamine or tyramine. The results imply that prevalent polymorphisms of the human FMO3 gene may contribute to low penetrance predispositions to diseases associated with adverse environmental exposures to heteroatom-containing chemicals, drugs, and endogenous amines.
Assuntos
Inativação Metabólica/genética , Oxigenases/genética , Polimorfismo Genético/genética , Austrália , Canadá , Clonagem Molecular , DNA/biossíntese , DNA/genética , Eletroforese em Gel de Poliacrilamida , Flavina-Adenina Dinucleotídeo/metabolismo , Genética Populacional , Humanos , Immunoblotting , Cinética , Oxigenases/biossíntese , Quebeque , Especificidade por SubstratoRESUMO
The mutation in a patient with dihydropteridine reductase deficiency has been located and characterized. Polymerase chain reaction (PCR) was used to amplify the coding sequence of human dihydropteridine reductase from the messenger RNA of skin fibroblasts. Chemical cleavage of mismatches indicated a mismatched thymine and cytosine at approximately 117 and 147 bases, respectively, from the end of the probe. Cloning and sequencing of the mutant PCR products revealed the insertion of the triplet ACT (threonine), after alanine 122 (base 390). Amplification of a small region around this mutation by using genomic DNA as the PCR target indicates that the mutation is completely within an exon. Unequal crossing-over at the second base in the preceding alanine codon and duplication of the bases CTA may be the mechanism of mutagenesis. The cleavage site 147 bases from the end of the probe corresponded to the conversion of guanine to adenine at base 420 (CTG to CTA) and does not alter the code for leucine. This change, which was also seen in another dihydropteridine reductase-deficient child and in a control subject probably represents a common neutral polymorphism.
Assuntos
Códon , NADH NADPH Oxirredutases/deficiência , Fenilcetonúrias , RNA Mensageiro , Treonina/genética , Sequência de Aminoácidos , Sequência de Bases , DNA/genética , Sondas de DNA , Di-Hidropteridina Redutase/genética , Humanos , Dados de Sequência Molecular , Ácidos Nucleicos Heteroduplexes/genética , Reação em Cadeia da PolimeraseRESUMO
The tyrP gene which codes for a component of the tyrosine-specific transport system of Escherichia coli has been cloned on a 2.8-kilobase insert into plasmid pBR322. Transposon mutagenesis, using Tn1000, indicates that the tyrP+ gene is at least 1.1 kilobase in length. Labeling of the tyrP protein in maxicells with [35S]methionine indicates an apparent molecular weight of ca. 24,500. Sedimentation analysis reveals that the tyrP protein is associated with the cell membrane and is not free in the cytoplasm or periplasm. Strains with many copies of the tyrP+ gene show an enhanced uptake of tyrosine, but the expression of the system is still modulated by tyrosine and phenylalanine in the presence of the tyrR+ regulator protein. Accumulated radioactive tyrosine is rapidly effluxed by the addition either of energy uncouplers or of excess nonradioactive tyrosine, indicating that the transport system is energized by the proton motive force and that the internal pool is readily exchangeable. The effect of increasing expression of the tyrP gene on the steady-state level of tyrosine accumulated by cells indicates that although the transport system may be dependent on the proton motive force to drive uptake, the system never reaches thermodynamic equilibrium with it.