RESUMO
Mutations in the ATM gene are responsible for the multisystem disorder ataxia-telangiectasia, characterized by neurodegeneration, immune deficiency and cancer predisposition. While no alternative splicing was identified within the coding region, the first four exons of the ATM gene, which fall within the 5'untranslated region (UTR), undergo extensive alternative splicing. We identified 12 different 5'UTRs that show considerable diversity in length and sequence contents. These mRNA leaders, which range from 150 to 884 nucleotides (nt), are expected to form variable secondary structures and contain different numbers of AUG codons. The longest 5'UTR contains a total of 18 AUGs upstream of the translation start site. The 3'UTR of 3590 nt is contained within a single 3'exon. Alternative polyadenylation results in 3'UTRs of varying lengths. These structural features suggest that ATM expression might be subject to complex post-transcriptional regulation, enabling rapid modulation of ATM protein level in response to environmental stimuli or alterations in cellular physiological states.
Assuntos
Ataxia Telangiectasia/genética , Proteínas Serina-Treonina Quinases , Proteínas/genética , Processamento Pós-Transcricional do RNA , Proteínas Mutadas de Ataxia Telangiectasia , Sequência de Bases , Proteínas de Ciclo Celular , Linhagem Celular , DNA , Proteínas de Ligação a DNA , Humanos , Células Jurkat , Masculino , Dados de Sequência Molecular , Biossíntese de Proteínas , Proteínas/metabolismo , RNA Mensageiro/genética , Proteínas Supressoras de TumorRESUMO
A human gene encoding a putative RNA helicase, designated DDX10, was identified 400 kb telomeric to the ataxia-telangiectasia gene at chromosome 11q22-q23. The predicted amino acid sequence shows very high similarity to a subgroup of DEAD-box RNA helicases involved in ribosome biogenesis. This novel gene encodes a 3.2-kb transcript in a variety of human tissues. A processed pseudogene of DDX10 was detected at chromosome 9q21-q22. We observed a rare trinucleotide repeat length polymorphism within the coding sequence of DDX10.
Assuntos
Cromossomos Humanos Par 11 , RNA Nucleotidiltransferases/genética , Sequência de Aminoácidos , Animais , Ataxia Telangiectasia/genética , Sequência de Bases , Linhagem Celular , Cromossomos Humanos Par 9 , Cricetinae , DNA Complementar , Expressão Gênica , Humanos , Dados de Sequência Molecular , Pseudogenes , RNA Helicases , Homologia de Sequência de Aminoácidos , Repetições de TrinucleotídeosRESUMO
Ataxia-telangiectasia (A-T) is an autosomal recessive disorder involving cerebellar degeneration, immunodeficiency radiation sensitivity, and cancer predisposition. A-T heterozygotes are moderately cancer prone. The A-T gene, designated ATM, was recently identified in our laboratory by positional cloning, and a partial cDNA clone was found to encode a polypeptide with a PI-3 kinase domain. We report here the molecular cloning of a cDNA contig spanning the complete open reading frame of the ATM gene. The predicted protein of 3056 amino acids shows significant sequence similarities to several large proteins in yeast, Drosophila and mammals, all of which share the PI-3 kinase domain. Many of these proteins are involved in the detection of DNA damage and the control of cell cycle progression. Mutations in their genes confer a variety of phenotypes with features similar to those observed in human A-T cells. The complete sequence of the ATM gene product provides useful clues to the function of this protein, and furthers understanding of the pleiotropic nature of the A-T mutations.
Assuntos
Proteínas Serina-Treonina Quinases , Proteínas/genética , Sequência de Aminoácidos , Animais , Proteínas Mutadas de Ataxia Telangiectasia , Sequência de Bases , Ciclo Celular , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Clonagem Molecular , Reparo do DNA , Proteínas de Ligação a DNA , Humanos , Dados de Sequência Molecular , Fases de Leitura Aberta , Fosfatidilinositol 3-Quinases , Fosfotransferases (Aceptor do Grupo Álcool)/genética , Fosfotransferases (Aceptor do Grupo Álcool)/metabolismo , Proteínas/metabolismo , Homologia de Sequência de Aminoácidos , Proteínas Supressoras de TumorRESUMO
A gene, ATM, that is mutated in the autosomal recessive disorder ataxia telangiectasia (AT) was identified by positional cloning on chromosome 11q22-23. AT is characterized by cerebellar degeneration, immunodeficiency, chromosomal instability, cancer predisposition, radiation sensitivity, and cell cycle abnormalities. The disease is genetically heterogeneous, with four complementation groups that have been suspected to represent different genes. ATM, which has a transcript of 12 kilobases, was found to be mutated in AT patients from all complementation groups, indicating that it is probably the sole gene responsible for this disorder. A partial ATM complementary DNA clone of 5.9 kilobases encoded a putative protein that is similar to several yeast and mammalian phosphatidylinositol-3' kinases that are involved in mitogenic signal transduction, meiotic recombination, and cell cycle control. The discovery of ATM should enhance understanding of AT and related syndromes and may allow the identification of AT heterozygotes, who are at increased risk of cancer.
Assuntos
Ataxia Telangiectasia/genética , Cromossomos Humanos Par 11 , Fosfotransferases (Aceptor do Grupo Álcool)/genética , Proteínas Serina-Treonina Quinases , Proteínas/genética , Sequência de Aminoácidos , Proteínas Mutadas de Ataxia Telangiectasia , Ciclo Celular , Proteínas de Ciclo Celular , Mapeamento Cromossômico , Cromossomos Artificiais de Levedura , Clonagem Molecular , DNA Complementar/genética , Proteínas de Ligação a DNA , Feminino , Teste de Complementação Genética , Predisposição Genética para Doença , Heterozigoto , Humanos , Masculino , Meiose , Dados de Sequência Molecular , Neoplasias/genética , Hibridização de Ácido Nucleico , Fosfatidilinositol 3-Quinases , Fosfotransferases (Aceptor do Grupo Álcool)/química , Fosfotransferases (Aceptor do Grupo Álcool)/fisiologia , Proteínas/química , Proteínas/fisiologia , Tolerância a Radiação , Deleção de Sequência , Transdução de Sinais , Proteínas Supressoras de TumorRESUMO
The locus of the autosomal recessive disorder ataxia-telangiectasia (A-T) has been assigned by linkage analysis with biallelic markers to a 4-Mb interval on chromosome 11q22-23, between GRIA4 and D11S1897. We have undertaken to saturate the A-T region with highly polymorphic microsatellite markers. To this end, we have identified seven new polymorphic CA-repeats in this region, and have mapped to it five new markers generated by Genethon and the Cooperative Human Linkage Center. These markers are in addition to 12 others that we have previously mapped or generated at the A-T locus. All 24 markers have been integrated into a high-density microsatellite map spanning some 6 Mb DNA. This map, which contains the A-T locus and flanking sequences, allows the construction of extensive, highly informative haplotypes.
Assuntos
Ataxia Telangiectasia/genética , Mapeamento Cromossômico , DNA Satélite/genética , Sequência de Bases , Cromossomos Humanos Par 11/genética , Primers do DNA/química , Marcadores Genéticos , Humanos , Dados de Sequência Molecular , Reação em Cadeia da Polimerase , Polimorfismo GenéticoRESUMO
Genes responsible for genetic diseases with increased sensitivity to DNA-damaging agents can be identified using complementation cloning. This strategy is based on in vitro complementation of the cellular sensitivity by gene transfer. Ataxia-telangiectasia (A-T) is a multisystem autosomal recessive disorder involving cellular sensitivity to ionizing radiation and radiomimetic drugs. A-T is genetically heterogeneous, with four complementation groups. We attempted to identify cDNA clones that modify the radiomimetic sensitivity of A-T cells assigned to complementation group [A-T(A)]. The cells were transfected with human cDNA libraries cloned in episomal vectors, and various protocols of radiomimetic selection were applied. Thirteen cDNAs rescued from survivor cells were found to confer various degrees of radiomimetic resistance to A-T(A) cells upon repeated introduction, and one of them also partially influenced another feature of the A-T phenotype, radioresistant DNA synthesis. None of the clones mapped to the A-T locus on chromosome 11q22-23. Nine of the clones were derived from known genes, some of which are involved in cellular stress responses. We concluded that a number of different genes, not necessarily associated with A-T, can influence the response of A-T cells to radiomimetic drugs, and hence the complementation cloning approach may be less applicable to A-T than to other diseases involving abnormal processing of DNA damage.
