m6A modification impacts hepatic drug and lipid metabolism properties by regulating carboxylesterase 2
m6A modification impacts hepatic drug and lipid metabolism properties by regulating carboxylesterase 2
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DOI：
10.1016/j.bcp.2021.114766
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发表时间：
2021-09-22
影响因子：
5.8
通讯作者：
Nakajima, Miki
中科院分区：
文献类型：
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作者：
Takemoto, Seiya;Nakano, Masataka;Nakajima, Miki
N-6 位腺苷甲基化形成 N-6-甲基腺苷 (m(6)A) 是哺乳动物 mRNA 最常见的表观转录组修饰。这种修饰由类甲基转移酶 3 (METTL3)-METTL14 复合物催化，并被脂肪量和肥胖相关蛋白 (FTO) 或 AlkB 同源物 5 (ALKBH5) 等去甲基酶消除。 m(6)A 修饰通过读取蛋白（例如 YT521-B 同源 (YTH) 家族成员）的识别来调节 mRNA 稳定性、核输出、剪接和/或蛋白质翻译。羧酸酯酶 2 (CES2) 是一种丝氨酸酯酶，负责水解药物和内源性底物，例如甘油三酯和甘油二酯。在这里，我们研究了 m(6)A 修饰对人类 CES2 表达的潜在调节。在 HepaRG 和 HepG2 细胞中，双敲低 METTL3 和 METTL14 使 CES2 mRNA 水平显着增加，但通过敲低 FTO 或 ALKBH5 而降低，导致其蛋白质水平和 7-乙基-10-[4-(1-哌啶基)-1-哌啶基]羰氧基喜树碱 (CPT-11) 水解酶活性发生变化，表明 m(6)A 修饰对 CES2 负向调节表达。与 CES2 表达的变化一致，双敲低 METTL3 和 METTL14 减少了细胞中的脂质积累，但敲低 FTO 或 ALKBH5 则增加了细胞中的脂质积累。使用抗 m(6)A 抗体进行的 RNA 免疫沉淀测定表明，5'-非翻译区 (UTR) 和 CES2 最后一个外显子中的腺苷被甲基化。荧光素酶测定表明，YTHDC2 可降解含有 m(6)A 的 mRNA，通过识别 CES2 5'-UTR 中的 m(6)A 来下调 CES2 表达。总的来说，我们证明 m(6)A 修饰对 CES2 的调节有很大影响，影响药代动力学、药物反应和脂质代谢。
Methylation of adenosine at the N-6 position to form N-6-methyladenosine (m(6)A) is the most prevalent epitran-scriptomic modification of mammalian mRNA. This modification is catalyzed by a methyltransferase-like 3 (METTL3)-METTL14 complex and is erased by demethylases such as fat mass and obesity-associated protein (FTO) or AlkB homolog 5 (ALKBH5). m(6)A modification regulates mRNA stability, nuclear export, splicing, and/or protein translation via recognition by reader proteins such as members of YT521-B homology (YTH) family. Carboxylesterase 2 (CES2) is a serine esterase responsible for the hydrolysis of drugs and endogenous substrates, such as triglycerides and diacylglycerides. Here, we examined the potential regulation of human CES2 expression by m(6)A modification. CES2 mRNA level was significantly increased by double knockdown of METTL3 and METTL14 but was decreased by knockdown of FTO or ALKBH5 in HepaRG and HepG2 cells, leading to changes in its protein level and hydrolase activity for 7-ethyl-10-[4-(1-piperidino)-1-piperidino]carbonyloxycamptothecin (CPT-11), suggesting that m(6)A modification negatively regulates CES2 expression. Consistent with the changes in CES2 expression, lipid accumulation in the cells was decreased by double knockdown of METTL3 and METTL14 but was increased by knockdown of FTO or ALKBH5. RNA immunoprecipitation assays using an anti-m(6)A antibody showed that adenosines in the 5'-untranslated region (UTR) and the last exon of CES2 are methylated. Luciferase assays revealed that YTHDC2, which degrades m(6)A-containing mRNA, downregulates CES2 expression by recognition of m(6)A in the 5'-UTR of CES2. Collectively, we demonstrated that m(6)A modification has a great impact on the regulation of CES2, affecting pharmacokinetics, drug response and lipid metabolism.