1 State Key Laboratory of Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu, China.
2 West China School of Stomatology, Sichuan University, Chengdu, China.
J Dent Res. 2018 May;97(5):571-579. doi: 10.1177/0022034517746987. Epub 2018 Jan 2.
The enhancer of zeste homolog 2 (EZH2) is a catalytic subunit of PRC2 (polycomb repressor complex 2). It mediates gene silencing via methyltransferase activity and is involved in the determination of cell lineage. However, the function of EZH2 and the underlying mechanisms by which it affects the differentiation of human dental pulp cell (hDPC) have remained underexplored. In this research, we found that EZH2 expression decreased during the mineralization of hDPCs, with attenuated H3K27me3 (trimethylation on lysine 27 in histone H3). Overexpression of EZH2 impaired the odontogenic differentiation of hDPCs, while EZH2 without methyltransferase activity mutation (mutation of suppressed variegation of 3 to 9, enhancer of zeste and trithorax domain, EZH2ΔSET) did not display this phenotype. In addition, siRNA knockdown studies showed that EZH2 negatively modulated hDPC differentiation in vitro and inhibited mineralized nodule formation in transplanted β-tricalcium phosphate / hDPC composites. To further investigate the underlying mechanisms, we explored the Wnt/β-catenin signaling pathway in view of the fact that previous research had documented the essential role that it plays during hDPC mineralization, as well as its links to EZH2 in other cells. We demonstrated for the first time that EZH2 depletion activated the Wnt/β-catenin signaling pathway and enhanced the accumulation of β-catenin in hDPCs. Chromatin immunoprecipitation analysis suggested that these effects are attributable to the level of the EZH2-regulated H3K27me3 on the β-catenin promoter. We conclude that EZH2 plays a negative role during the odontogenic differentiation of hDPCs. Suppression of EZH2 could promote hDPC mineralization by epigenetically regulating the expression of β-catenin and activating the Wnt canonical signaling pathway.
增强子结合锌指蛋白 2（EZH2）是 PRC2（多梳抑制复合物 2）的催化亚基。它通过甲基转移酶活性介导基因沉默，并参与细胞谱系的确定。然而，EZH2 的功能以及它影响人牙髓细胞（hDPC）分化的潜在机制仍未得到充分探索。在这项研究中，我们发现 EZH2 的表达在 hDPC 矿化过程中降低，同时 H3K27me3（组蛋白 H3 赖氨酸 27 上的三甲基化）减弱。EZH2 的过表达会损害 hDPC 的成牙分化，而缺乏甲基转移酶活性突变的 EZH2（突变抑制 3 到 9 的变异性、增强子结合锌指蛋白和三结构域、EZH2ΔSET）则没有表现出这种表型。此外，siRNA 敲低研究表明，EZH2 在体外负调控 hDPC 分化，并抑制移植β-磷酸三钙/hDPC 复合材料中矿化结节的形成。为了进一步探讨潜在机制，我们研究了 Wnt/β-catenin 信号通路，因为之前的研究已经证明了它在 hDPC 矿化过程中的重要作用，以及它在其他细胞中与 EZH2 的联系。我们首次证明，EZH2 的耗竭激活了 Wnt/β-catenin 信号通路，并增强了 hDPC 中β-catenin 的积累。染色质免疫沉淀分析表明，这些效应归因于 EZH2 调节的 H3K27me3 水平对β-catenin 启动子的影响。我们的结论是，EZH2 在 hDPC 的成牙分化过程中发挥负性作用。抑制 EZH2 可以通过表观遗传调控β-catenin 的表达并激活 Wnt 经典信号通路来促进 hDPC 的矿化。
