Involvement of DOT1L in the Remodeling of Heterochromatin Configuration During Early Preimplantation Development in Mice
Involvement of DOT1L in the Remodeling of Heterochromatin Configuration During Early Preimplantation Development in Mice
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DOI：
10.1095/biolreprod.113.113258
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发表时间：
2013-12-01
影响因子：
3.6
通讯作者：
Aoki, Fugaku
中科院分区：
文献类型：
--
作者：
Ooga, Masatoshi;Suzuki, Masataka G.;Aoki, Fugaku
在受精和早期植入前发育过程中，染色质结构发生了显著的改变。虽然在卵母细胞的核中观察到染色中心，即着丝粒周围的异染色质簇，但它们在受精后消失，然后在四细胞期重新出现。为了阐明异染色质重组的机制，我们研究了着床前发育过程中DOT 1 L的表达和核定位，DOT 1 L通过组蛋白H3赖氨酸79（H3 K79）甲基转移酶活性参与异染色质结构的调节。Dot 1 L mRNA水平在二细胞期较低。在免疫细胞化学分析中，在该阶段的细胞核中未观察到DOT 1 L蛋白。显微注射Flag标记的Dot 1 L cRNA显示，DOT 1 L蛋白定位于一细胞和四细胞期的胚胎的细胞核中，而不是在两细胞期。然而，C-末端截短的DOT 1 L定位于两细胞期胚胎的细胞核中。表达截短的DOT 1 L引起H3 K79的高甲基化，并在两细胞阶段形成类似于染色中心的结构。有趣的是，无催化活性的截短DOT 1 L的表达也导致了类似于染色中心的结构的形成，而没有增加H3 K79甲基化。大多数表达截短的DOT 1 L或其非活性形式的胚胎被阻止在两细胞阶段。这些结果表明，DOT 1 L的情况下，这是参与形成一个特定的配置异染色质在两个细胞阶段，是必不可少的早期植入前的发展。
The global chromatin configuration is dramatically remodeled during fertilization and early preimplantation development. Although the chromocenters, which are pericentromeric heterochromatin clusters, are observed in the nuclei of oocytes, they disappear after fertilization and then reappear at the fourcell stage. To elucidate the mechanism of this reorganization of heterochromatin, we investigated the expression and nuclear localization of DOT1L, which is involved in the regulation of heterochromatin structure through histone H3 lysine 79 (H3K79) methyltransferase activity, during preimplantation development. The Dot1L mRNA level was low at the two-cell stage. In the analysis by the immunocytochemistry, DOT1L protein was not observed in the nuclei at this stage. Microinjection of Flag-tagged Dot1L cRNA revealed that the DOT1L protein was localized in the nucleus of the embryos at the onecell and four-cell stages but not at the two-cell stage. However, C-terminus-truncated DOT1L was localized in the nucleus of two-cell-stage embryos. Expression of the truncated DOT1L caused hypermethylation on H3K79 and the formation of chromocenter-like structures at the two-cell stage. Intriguingly, the expression of catalytically inactive truncated DOT1L also caused the formation of chromocenter-like structures without an increase in H3K79 methylation. Most embryos expressing the truncated DOT1L or its inactive form were arrested at the twocell stage. These results suggest that the absence of DOT1L, which is involved in the formation of a specific configuration of heterochromatin at the two-cell stage, is essential for early preimplantation development.