Essential roles of HDAC1 and 2 in lineage development and genome-wide DNA methylation during mouse preimplantation development

Epigenetic modifications, including DNA methylation and histone modifications, are reprogrammed considerably following fertilization during mammalian early embryonic development. Incomplete epigenetic reprogramming is a major factor leading to poor developmental outcome in embryos generated by assisted reproductive technologies, such as somatic cell nuclear transfer. However, the role of histone modifications in preimplantation development is poorly understood. Here, we show that co-knockdown (cKD) of Hdac1 and 2 (but not individually) resulted in developmental failure during the morula to blastocyst transition. This outcome was also confirmed with the use of small-molecule HDAC1/2-specific inhibitor FK228. We observed reduced cell proliferation and increased incidence of apoptosis in cKD embryos, which were likely caused by increased acetylation of TRP53. Importantly, both RNA-seq and immunostaining analysis revealed a failure of lineage specification to generate trophectoderm and pluripotent cells. Among many gene expression changes, a substantial decrease of Cdx2 may be partly accounted for by the aberrant Hippo pathway occurring in cKD embryos. In addition, we observed an increase in global DNA methylation, consistent with increased DNA methyltransferases and UHRF1. Interestingly, deficiency of RBBP4 and 7 (both are core components of several HDAC1/2-containing epigenetic complexes) results in similar phenotypes as those of cKD embryos. Overall, HDAC1 and 2 play redundant functions required for lineage specification, cell viability and accurate global DNA methylation, each contributing to critical developmental programmes safeguarding a successful preimplantation development.

在哺乳动物早期胚胎发育过程中，受精后的表观遗传修饰（Epigenetic modifications），包括DNA甲基化（DNA methylation）与组蛋白修饰（histone modifications），会发生显著重编程。表观遗传重编程不完全是辅助生殖技术（Assisted Reproductive Technologies）所制备胚胎发育结局不佳的主要诱因，其中体细胞核移植（Somatic Cell Nuclear Transfer）来源的胚胎便是典型代表。然而，学界对组蛋白修饰在胚胎着床前发育中的作用仍知之甚少。本研究发现，同时敲低（co-knockdown，简称cKD）Hdac1与Hdac2（而非单独敲低任一亚型）会导致胚胎在桑椹胚向囊胚的转变过程中出现发育停滞。该实验结果亦可通过使用小分子HDAC1/2特异性抑制剂FK228得到验证。研究人员在共敲低胚胎中观察到细胞增殖能力下降与细胞凋亡发生率升高，该现象大概率由TRP53乙酰化水平上调所介导。重要的是，RNA测序（RNA-seq）与免疫荧光染色分析均显示，共敲低胚胎的细胞谱系特化过程出现异常，无法生成滋养外胚层与多能细胞。在诸多基因表达变化中，Cdx2的显著下调可能部分源于共敲低胚胎中异常激活的Hippo信号通路（Hippo pathway）。此外，研究团队还观察到全基因组DNA甲基化水平升高，这与DNA甲基转移酶（DNA methyltransferases）与UHRF1的表达上调相一致。有趣的是，RBBP4与RBBP7（二者均为多种含HDAC1/2的表观遗传复合物的核心组分）的敲除会引发与共敲低胚胎高度相似的表型。综上，HDAC1与HDAC2发挥冗余功能，共同保障细胞谱系特化、细胞存活以及精确的全基因组DNA甲基化，二者均参与调控关键发育程序，以确保胚胎着床前发育顺利完成。