Hara Satoshi, Takano Takashi, Fujikawa Tsugunari, Yamada Munehiro, Wakai Takuya, Kono Tomohiro, Obata Yayoi
Department of BioScience, Tokyo University of Agriculture, 1-1-1 Sakuragaoka, Setagaya-ku, Tokyo 156-8502, Japan.
Department of BioScience, Tokyo University of Agriculture, 1-1-1 Sakuragaoka, Setagaya-ku, Tokyo 156-8502, Japan
Hum Mol Genet. 2014 Jul 15;23(14):3853-64. doi: 10.1093/hmg/ddu100. Epub 2014 Mar 5.
In mammals, genomic imprinting governed by DNA methyltransferase DNMT3A and its cofactor DNMT3L is essential for functional gametes. Oocyte-specific methylation imprints are established during oocyte growth concomitant with DNMT3A/DNMT3L expression, although the mechanisms of oocyte-specific imprinting are not fully understood. To determine whether the presence of DNMT3A/DNMT3L in oocytes is sufficient for acquisition of methylation imprints, we produced transgenic mice to induce DNMT3A/DNMT3L expression prematurely in oogenesis and analyzed DNA methylation imprints. The results showed that 2- to 4-fold greater expression of DNMT3A/DNMT3L was achieved in non-growing (ng) oocytes versus fully grown oocytes derived from wild-type mice, but the analyzed imprint domains were not methylated. Thus, the presence of DNMT3A/DNMT3L in ng oocytes is insufficient for methylation imprints, and imprinted regions are resistant to DNMT3A/DNMT3L in ng oocytes. In contrast, excess DNMT3A/DNMT3L accelerated imprint acquisition at Igf2r, Lit1, Zac1 and Impact but not Snrpn and Mest in growing oocytes. Therefore, DNMT3A/DNMT3L quantity is an important factor for imprint acquisition. Transcription at imprinted domains is proposed to be involved in de novo methylation; however, transcription at Lit1, Snrpn and Impact was observed in ng oocytes. Thus, transcription cannot induce DNMT3A catalysis at imprinted regions even if DNMT3A/DNMT3L is present. However, the accelerated methylation imprints in oocytes, with the exception of Igf2r, were erased during embryogenesis. In conclusion, a sufficient amount of DNMT3A/DNMT3L and a shift from the resistant to permissive state are essential to establish oocyte-specific methylation imprints and that maintenance of the acquired DNA methylation imprints is essential for functional imprinting.
在哺乳动物中，由DNA甲基转移酶DNMT3A及其辅助因子DNMT3L调控的基因组印记对于功能性配子至关重要。卵母细胞特异性甲基化印记在卵母细胞生长过程中伴随着DNMT3A/DNMT3L的表达而建立，尽管卵母细胞特异性印记的机制尚未完全了解。为了确定卵母细胞中DNMT3A/DNMT3L的存在是否足以获得甲基化印记，我们制备了转基因小鼠，以在卵子发生过程中过早诱导DNMT3A/DNMT3L表达，并分析DNA甲基化印记。结果表明，与源自野生型小鼠的完全成熟卵母细胞相比，未成熟（ng）卵母细胞中DNMT3A/DNMT3L的表达量高2至4倍，但分析的印记区域未发生甲基化。因此，ng卵母细胞中DNMT3A/DNMT3L的存在不足以产生甲基化印记，并且印记区域对ng卵母细胞中的DNMT3A/DNMT3L具有抗性。相反，过量的DNMT3A/DNMT3L加速了生长中的卵母细胞中Igf2r、Lit1、Zac1和Impact的印记获得，但对Snrpn和Mest没有影响。因此，DNMT3A/DNMT3L的量是印记获得的重要因素。印记区域的转录被认为与从头甲基化有关；然而，在ng卵母细胞中观察到Lit1、Snrpn和Impact的转录。因此，即使存在DNMT3A/DNMT3L，转录也不能在印记区域诱导DNMT3A催化。然而，除Igf2r外，卵母细胞中加速的甲基化印记在胚胎发生过程中被消除。总之，足够量的DNMT3A/DNMT3L以及从抗性状态到许可状态的转变对于建立卵母细胞特异性甲基化印记至关重要，并且维持获得的DNA甲基化印记对于功能性印记至关重要。
