Arai Daisuke, Hayakawa Koji, Ohgane Jun, Hirosawa Mitsuko, Nakao Yoichi, Tanaka Satoshi, Shiota Kunio
Laboratory of Cellular Biochemistry, Department of Animal Resource Sciences/Veterinary Medical Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan; Laboratory of Chemical Biology, Department of Chemistry and Biochemistry, School of Advanced Science and Engineering, Waseda University, 3-4-1 Ohkubo, Shinjuku-ku, Tokyo 169-8555, Japan.
Laboratory of Cellular Biochemistry, Department of Animal Resource Sciences/Veterinary Medical Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan.
Mech Dev. 2015 May;136:143-54. doi: 10.1016/j.mod.2014.12.003. Epub 2014 Dec 17.
Nodal signaling plays critical roles during embryonic development. The Nodal gene is not expressed in adult tissues but is frequently activated in cancer cells, contributing to progression toward malignancy. Although several regulatory elements of the Nodal gene have been identified, the epigenetic mechanisms by which Nodal expression is regulated over the long term remain unclear. We found a region exhibiting dynamic changes in DNA methylation at approximately -3.0 kb to -0.4 kb upstream from the transcriptional start site (TSS) that we termed the epigenetic regulatory element (ERE). The ERE was unmethylated in mouse embryonic stem cells (mESCs) but became increasingly methylated in differentiated cells and tissues, concomitant with the downregulation of Nodal mRNA expression. In vitro reporter assays identified an Oct3/4 binding motif within the ERE, indicating that the ERE is responsible for the activation of Nodal in mESCs. Furthermore, the ERE was a target of differentiation-associated Polycomb silencing, and the chromatin condensed when mESCs differentiated to embryoid bodies (EBs). Pharmacological inhibition of PRC2 led to the reactivation of Nodal expression in EBs and mouse embryonic fibroblasts (MEFs). The ERE was also targeted by PRC2 in normal human cells. In NODAL-expressing human cancer cells, accumulation of EZH2 and trimethylation of H3K27 at the ERE were diminished. In conclusion, Nodal is epigenetically controlled through the ERE in the mouse embryo and human cells.
Nodal信号在胚胎发育过程中发挥着关键作用。Nodal基因在成年组织中不表达，但在癌细胞中经常被激活，促进了肿瘤的进展。尽管已经确定了Nodal基因的几个调控元件，但Nodal表达长期受到调控的表观遗传机制仍不清楚。我们发现一个区域在转录起始位点（TSS）上游约-3.0 kb至-0.4 kb处的DNA甲基化呈现动态变化，我们将其称为表观遗传调控元件（ERE）。ERE在小鼠胚胎干细胞（mESCs）中未甲基化，但在分化细胞和组织中甲基化程度逐渐增加，同时伴随着Nodal mRNA表达的下调。体外报告基因检测在ERE内鉴定出一个Oct3/4结合基序，表明ERE负责在mESCs中激活Nodal。此外，ERE是分化相关的多梳抑制的靶点，当mESCs分化为胚状体（EBs）时染色质会浓缩。PRC2的药理学抑制导致EBs和小鼠胚胎成纤维细胞（MEFs）中Nodal表达的重新激活。ERE在正常人类细胞中也是PRC2的靶点。在表达NODAL的人类癌细胞中，ERE处EZH2的积累和H3K27的三甲基化减少。总之，在小鼠胚胎和人类细胞中，Nodal通过ERE受到表观遗传控制。
