RESUMO
BACKGROUND: Dystrophin is a large essential protein of skeletal and heart muscle. It is a filamentous scaffolding protein with numerous binding domains. Mutations in the DMD gene, which encodes dystrophin, mostly result in the deletion of one or several exons and cause Duchenne (DMD) and Becker (BMD) muscular dystrophies. The most common DMD mutations are frameshift mutations resulting in an absence of dystrophin from tissues. In-frame DMD mutations are less frequent and result in a protein with partial wild-type dystrophin function. The aim of this study was to highlight structural and functional modifications of dystrophin caused by in-frame mutations. METHODS AND RESULTS: We developed a dedicated database for dystrophin, the eDystrophin database. It contains 209 different non frame-shifting mutations found in 945 patients from a French cohort and previous studies. Bioinformatics tools provide models of the three-dimensional structure of the protein at deletion sites, making it possible to determine whether the mutated protein retains the typical filamentous structure of dystrophin. An analysis of the structure of mutated dystrophin molecules showed that hybrid repeats were reconstituted at the deletion site in some cases. These hybrid repeats harbored the typical triple coiled-coil structure of native repeats, which may be correlated with better function in muscle cells. CONCLUSION: This new database focuses on the dystrophin protein and its modification due to in-frame deletions in BMD patients. The observation of hybrid repeat reconstitution in some cases provides insight into phenotype-genotype correlations in dystrophin diseases and possible strategies for gene therapy. The eDystrophin database is freely available: http://edystrophin.genouest.org/.
Assuntos
Distrofina/genética , Estudos de Associação Genética , Distrofia Muscular de Duchenne/genética , Mutação , Fases de Leitura/genética , Adolescente , Biologia Computacional , Bases de Dados Genéticas , Éxons/genética , Feminino , Genótipo , Humanos , Internet , Masculino , Distrofia Muscular de Duchenne/patologiaRESUMO
BACKGROUND: Out-of-frame stop codons (OSCs) occur naturally in coding sequences of all organisms, providing a mechanism of early termination of translation in incorrect reading frame so that the metabolic cost associated with frameshift events can be reduced. Given such a functional significance, we expect statistically overrepresented OSCs in coding sequences as a result of a widespread selection. Accordingly, we examined available prokaryotic genomes to look for evidence of this selection. RESULTS: The complete genome sequences of 990 prokaryotes were obtained from NCBI GenBank. We found that low G+C content coding sequences contain significantly more OSCs and G+C content at specific codon positions were the principal determinants of OSC usage bias in the different reading frames. To investigate if there is overrepresentation of OSCs, we modeled the trinucleotide and hexanucleotide biases of the coding sequences using Markov models, and calculated the expected OSC frequencies for each organism using a Monte Carlo approach. More than 93% of 342 phylogenetically representative prokaryotic genomes contain excess OSCs. Interestingly the degree of OSC overrepresentation correlates positively with G+C content, which may represent a compensatory mechanism for the negative correlation of OSC frequency with G+C content. We extended the analysis using additional compositional bias models and showed that lower-order bias like codon usage and dipeptide bias could not explain the OSC overrepresentation. The degree of OSC overrepresentation was found to correlate negatively with the optimal growth temperature of the organism after correcting for the G+C% and AT skew of the coding sequence. CONCLUSIONS: The present study uses approaches with statistical rigor to show that OSC overrepresentation is a widespread phenomenon among prokaryotes. Our results support the hypothesis that OSCs carry functional significance and have been selected in the course of genome evolution to act against unintended frameshift occurrences. Some results also hint that OSC overrepresentation being a compensatory mechanism to make up for the decrease in OSCs in high G+C organisms, thus revealing the interplay between two different determinants of OSC frequency.
Assuntos
Códon de Terminação/genética , Mutação da Fase de Leitura/genética , Peptídeos/genética , Células Procarióticas/metabolismo , Fases de Leitura/genética , Seleção Genética/genética , Sequência de Aminoácidos , Bactérias/genética , Bactérias/crescimento & desenvolvimento , Composição de Bases/genética , Sequência de Bases , Viés , Simulação por Computador , Genoma Bacteriano/genética , Cadeias de Markov , Dados de Sequência Molecular , Método de Monte Carlo , Fases de Leitura Aberta/genética , Peptídeos/química , Análise de Componente Principal , TemperaturaRESUMO
BACKGROUND: Many current gene prediction methods use only one model to represent protein-coding regions in a genome, and so are less likely to predict the location of genes that have an atypical sequence composition. It is likely that future improvements in gene finding will involve the development of methods that can adequately deal with intra-genomic compositional variation. RESULTS: This work explores a new approach to gene-prediction, based on the Self-Organizing Map, which has the ability to automatically identify multiple gene models within a genome. The current implementation, named RescueNet, uses relative synonymous codon usage as the indicator of protein-coding potential. CONCLUSIONS: While its raw accuracy rate can be less than other methods, RescueNet consistently identifies some genes that other methods do not, and should therefore be of interest to gene-prediction software developers and genome annotation teams alike. RescueNet is recommended for use in conjunction with, or as a complement to, other gene prediction methods.
Assuntos
Mapeamento Cromossômico/métodos , Genes Arqueais/genética , Genes Bacterianos/genética , Modelos Genéticos , Família Multigênica/genética , Mapeamento Cromossômico/estatística & dados numéricos , Códon/genética , Biologia Computacional/métodos , Biologia Computacional/estatística & dados numéricos , Deinococcus/genética , Sequência Rica em GC/genética , Genoma Arqueal , Genoma Bacteriano , Bactérias Gram-Negativas/genética , Bactérias Gram-Positivas Formadoras de Endosporo/genética , Cadeias de Markov , Mathanococcus/genética , Valor Preditivo dos Testes , Fases de Leitura/genética , SoftwareRESUMO
Increased productivity in DNA sequencing would not be valid without a straightforward detection and estimation of errors in finished sequences. The sequence of the surfactin operon from Bacillus subtilis was obtained by two different groups and by chance we were also working on the same chromosome region. Taking advantage of this situation we report in this paper, the number and nature of errors found in the overlapping part of the DNA sequences obtained by the three laboratories. The coincidence of some of the errors with compression in sequence ladders and with secondary DNA structures as well as the detection of frameshift errors using computer programs, are demonstrated. Finally we discuss the definition of a new sequencing strategy that might minimize both the error rate and the cost of sequencing.