| Budget Amount *help |
¥3,900,000 (Direct Cost: ¥3,900,000)
Fiscal Year 2005: ¥1,900,000 (Direct Cost: ¥1,900,000)
Fiscal Year 2004: ¥2,000,000 (Direct Cost: ¥2,000,000)
|
|---|
| Research Abstract |
The haploid spermatid specific gene, Tact1, is a single exon gene, which contains CpG-island within the gene. DNA methylation in the CpG-island represses the Tact1 gene expression in somatic cells. Demethylation is necessary for Tact1 expression in the round spermatids. In this study, we examined the timing, in which these CpGs are demethylated and remethylated, and involvement of the de novo methyltransferases, Dnmt3a and Dnmt3b with remethylation. We performed bisulfite-PCR sequencing for analyzing methylation status in the CpG sites within the Tact1 gene using genomic DNAs obtained from spermatogenic cells of various differentiation stages by laser micro dissection and elutriation. Demethylation occurs during proliferation stage of the spermatogonia, resulting from passive action due to inactivation of maintenance methyltransferase, Dnmt1. Hypomethylated status is maintained in pachytene spermatocytes, round spermatids, in which the Tact1 gene is expressed, and spermatozoa in epididymis. We found that remethylation occurs in mature spermatozoa in vas deferens. Analysis of germ line specific conditional knockout mice showed that genomic remethylation in spermatozoa may be redundantly regulated by two de novo methyltransferases, Dnmt3a and Dnmt3b. Alternatively, unidentified de novo methylation machinery may be necessary for newly methylation in sperm.
|
