Identification of target genes for DNA methylation in skeletal muscle and its medical application

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DNA methylation is essential for normal embryonic development, and altered DNA methylation patterns have been implicated in tumorigenesis. An epigenetic mechanism involving DNA methylation has also been suggested to be involved in the regulation of metabolic processes ; however, the molecular basis of this mechanism has not been clearly demonstrated. In this study, we attempted to get an insight into the role of DNA methylation in skeletal muscle, which plays important roles in exercise, energy expenditure and glucose metabolism. We made skeletal muscle-specific knockout(KO) mice with Dnmt3a(a de novo DNA methyltransferase highly expressed in skeletal muscle) by crossing Dnmt3a flox/flox mice and transgenic mice expressing Cre recombinase, driven by the skeletal muscle alpha-actin promoter. A quantitative real-time PCR analysis confirmed that Dnmt3a mRNA levels were markedly diminished in the skeletal muscle but not in other tissues of the KO mice. In this study, a genome-wide DNA methylation analysis called Microarray-based Integrated Analysis of Methylation by Isoschizomers(MIAMI) was preformed using the methylation-sensitive restriction enzyme HpaII and a genome microarray. MIAMI analysis revealed a marked decrease in DNA methylation in the KO mice, including that of several genes coding transcription factors. Despite this, the decreased DNA methylation did not correlate with the gene expression levels under the same condition. Because DNA methylation is considered a key epigenetic contributor in the maintenance of gene silencing, gene expression in the KO mice may be modified in the presence of additional metabolic stress. Therefore, analysis of metabolic phenotype of the KO mice will be essential to elucidate the epigenetic regulation of skeletal muscle gene expression and skeletal muscle-related metabolic diseases.