The de novo DNA methyltransferases Dnmt3a and Dnmt3b play crucial roles in developmental and cellular processes. Their enzymatic activities are stimulated by a regulatory protein Dnmt3L (Dnmt3-like) in vitro. However, genetic evidence indicates that Dnmt3L functions predominantly as a regulator of Dnmt3a in germ cells. How Dnmt3a and Dnmt3b activities are regulated during embryonic development and in somatic cells remains largely unknown. Here we show that Dnmt3b3, a catalytically inactive Dnmt3b isoform expressed in differentiated cells, positively regulates de novo methylation by Dnmt3a and Dnmt3b with a preference for Dnmt3b. Dnmt3b3 is equally potent as Dnmt3L in stimulating the activities of Dnmt3a2 and Dnmt3b2 in vitro. Like Dnmt3L, Dnmt3b3 forms a complex with Dnmt3a2 with a stoichiometry of 2:2. However, rescue experiments in Dnmt3a/3b/3l triple-knockout (TKO) mouse embryonic stem cells (mESCs) reveal that Dnmt3b3 prefers Dnmt3b2 over Dnmt3a2 in remethylating genomic sequences. Dnmt3a2, an active isoform that lacks the N-terminal uncharacterized region of Dnmt3a1 including a nuclear localization signal, has very low activity in TKO mESCs, indicating that an accessory protein is absolutely required for its function. Our results suggest that Dnmt3b3 and perhaps similar Dnmt3b isoforms facilitate de novo DNA methylation during embryonic development and in somatic cells.

中文翻译：

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无活性的 Dnmt3b3 亚型优先增强 Dnmt3b 介导的 DNA 甲基化


DNA 从头甲基转移酶 Dnmt3a 和 Dnmt3b 在发育和细胞过程中发挥着至关重要的作用。它们的酶活性在体外受到调节蛋白 Dnmt3L（Dnmt3 样）的刺激。然而，遗传证据表明 Dnmt3L 在生殖细胞中主要充当 Dnmt3a 的调节因子。 Dnmt3a 和 Dnmt3b 活性在胚胎发育和体细胞中如何受到调节仍然很大程度上未知。在这里，我们表明，Dnmt3b3（一种在分化细胞中表达的催化失活的 Dnmt3b 亚型）可通过 Dnmt3a 和 Dnmt3b 积极调节 Dnmt3a 和 Dnmt3b 的从头甲基化，并优先调节 Dnmt3b。 Dnmt3b3 在体外刺激 Dnmt3a2 和 Dnmt3b2 活性方面​​与 Dnmt3L 同等有效。与 Dnmt3L 一样，Dnmt3b3 与 Dnmt3a2 形成复合物，化学计量为 2:2。然而，在Dnmt3a/3b/3l三敲除 (TKO) 小鼠胚胎干细胞 (mESC) 中进行的拯救实验表明，在重新甲基化基因组序列方面，Dnmt3b3 比 Dnmt3a2 更喜欢 Dnmt3b2。 Dnmt3a2 是一种活性亚型，缺乏 Dnmt3a1 N 端未表征区域（包括核定位信号），在 TKO mESC 中活性非常低，表明辅助蛋白对其功能来说绝对是必需的。我们的结果表明，Dnmt3b3 和可能类似的 Dnmt3b 亚型在胚胎发育和体细胞中促进 DNA 从头甲基化。