Chen Si-Qi, Li Jia-Qi, Wang Xiao-Qiao, Lei Wen-Jing, Li Hao, Wan Jiao, Hu Zheng, Zou Yao-Wei, Wu Xiao-Yu, Niu Hong-Xin
Zhujiang Hospital, Southern Medical University, Guangzhou, 510282 China.
Division of Nephrology, Nanfang Hospital, Southern Medical University, North Guangzhou Ave 1838, Guangzhou, 510515 People's Republic of China.
Cell Div. 2020 May 25;15:8. doi: 10.1186/s13008-020-00064-3. eCollection 2020.
The enhancer of zeste homolog 2 (EZH2) is a histone methyltransferase and induces the trimethylation of histone H3 lysine 27 (H3K27me3) in the promoter of many key genes; EZH2 acts as a transcriptional repressor and is an epigenetic regulator for several cancers. However, the role of EZH2 in nonneoplastic diseases, such as kidney diseases, is unknown and has been investigated.
NRK-52E cells were treated with DZNep, a potent inhibitor of EZH2, with different concentrations and for different times to evaluate the apoptosis level of NRK-52E cells by Western blot and Flow cytometry analysis. The binding of EZH2 to the Deptor promoter was determined by ChIP assay.
The inhibition of EZH2 with 3-deazaneplanocin A (DZNep), a specific inhibitor of EZH2, led to the apoptosis of NRK-52E cells and the inhibition of mTORC1 and mTORC2 activity. A ChIP assay demonstrated that EZH2 bound the promoter region of Deptor, an endogenous inhibitor of mTORC1 and mTORC2, and regulated the transcription of Deptor by modulating H3K27me3 in its promoter region. Further experiments were performed to examine the effects of EZH2 inhibition on cisplatin-induced injured cells. Cisplatin induced the activation of mTORC1 and mTORC2 and apoptosis in NRK-52E cells, and DZNep inhibited mTORC1 and mTORC2 activity and aggravated cell apoptosis.
These data suggested that EZH2 inhibition increased the transcription of Deptor by modifying H3K27me3 in its promoter region, subsequently inhibited mTORC1 and mTORC2 activities, downregulated the expression of apoptosis suppressor genes, and finally led to apoptosis in renal tubular cells. The inhibition of EZH2 aggravated the cisplatin-induced injury in renal tubular cells by inactivating the mTOR complexes. The present study provides new insight into renal protection and suggests that EZH2 might be a target.
zeste 同源物 2（EZH2）增强子是一种组蛋白甲基转移酶，可诱导许多关键基因启动子中组蛋白 H3 赖氨酸 27（H3K27me3）的三甲基化；EZH2 作为转录抑制因子，是多种癌症的表观遗传调节因子。然而，EZH2 在非肿瘤性疾病（如肾脏疾病）中的作用尚不清楚，且已被研究。
用 EZH2 的强效抑制剂 DZNep 以不同浓度和不同时间处理 NRK-52E 细胞，通过蛋白质免疫印迹法和流式细胞术分析评估 NRK-52E 细胞的凋亡水平。通过染色质免疫沉淀（ChIP）试验确定 EZH2 与 Deptor 启动子的结合情况。
用 EZH2 的特异性抑制剂 3-去氮杂氮胞苷（DZNep）抑制 EZH2 导致 NRK-52E 细胞凋亡，并抑制 mTORC1 和 mTORC2 活性。ChIP 试验表明，EZH2 结合 mTORC1 和 mTORC2 的内源性抑制剂 Deptor 的启动子区域，并通过调节其启动子区域的 H3K27me3 来调控 Deptor 的转录。进行了进一步实验以研究 EZH2 抑制对顺铂诱导的损伤细胞的影响。顺铂诱导 NRK-52E 细胞中 mTORC1 和 mTORC2 的激活以及细胞凋亡，而 DZNep 抑制 mTORC1 和 mTORC2 活性并加重细胞凋亡。
这些数据表明，EZH2 抑制通过修饰 Deptor 启动子区域的 H3K27me3 增加 Deptor 的转录，随后抑制 mTORC1 和 mTORC2 活性，下调凋亡抑制基因的表达，最终导致肾小管细胞凋亡。EZH2 的抑制通过使 mTOR 复合物失活加重顺铂诱导的肾小管细胞损伤。本研究为肾脏保护提供了新的见解，并表明 EZH2 可能是一个靶点。
