Recent studies of the demethylation process in murine zygotes have shown that 5-methylcytosine (5mC) is first converted into 5-hydroxymethylcytosine (5hmC) or further-oxidized cytosines in the paternal genome by the maternal ten-eleven translocation 3 (TET3) enzyme. This process is crucial for normal embryogenesis, and our aim was to elucidate the effect of Tet3 on the maternal genome during female germ-line development. Immunofluorescence analysis showed that 5hmC was clearly present in fully grown oocytes but not in nongrowing and early growth-stage oocytes. The 5hmC in the maternal genome was clearly detectable in DNA methyltransferase 3-like enzyme (Dnmt3L)-null oocytes and their fertilized zygotes, although Dnmt3L is essential for DNA methylation in oocytes. An analysis using an enzyme digestion-based method showed that 5hmC was present in LTR retrotransposons from the late growth period of oocytes. Quantitative RT-PCR analysis showed that Tet3 expression was enhanced during oocyte growth and exhibited an approximately 40-fold increase between nongrowing and fully grown oocytes. Our results show that 5hmC is generated since the oocyte growth stage, accompanied by up-regulation of Tet3; 5hmC is located mainly in LTR retrotransposons, indicating that 5hmC generated in growth-stage oocytes is responsible for genomewide demethylation after fertilization.
ASJC Scopus subject areas
- Cell Biology