Chen Tong, Wang Juan, Xu Yanyan, Zhu Yonghong, Jin Ying, Fan Qiuling
Department of Nephrology, Shenyang Seventh People's Hospital, Shenyang, Liaoning, China.
Department of Nephrology, Shanghai General Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China.
Ren Fail. 2025 Dec;47(1):2521455. doi: 10.1080/0886022X.2025.2521455. Epub 2025 Jul 20.
To explore the N6-methyladenosine (mA) modification mechanism of taurine upregulated gene 1 (TUG1) and whether methyltransferase 3 (METTL3) can promote peroxisome proliferators-activated receptor γ coactivator 1 alpha (PGC-1α) transcription and alleviate mitochondrial dysfunction.
high glucose (HG)-treated HK-2 cell models and db/db mice models injected with rAAV-METTL3 the tail vein were established. The expression levels were determined by RT-qPCR, western blot, and immunohistochemical staining. RNA mA modification was analyzed by the RNase Mazf. The biochemical indicators of mice were detected by enzyme-linked immunosorbent assay. Cell apoptosis was detected by flow cytometry. Histopathological staining was performed to evaluate kidney injury. mtDNA content, mitochondrial complex activity, and ATP were detected by RT-qPCR and detection kits, respectively, per the manufacturer's instructions. Mitochondrial reactive oxygen species production in HK-2 cells incubated with MitoSOX Red and mitochondrial morphology were observed under a fluorescence microscope and transmission electron microscope, respectively. Molecular interactions were verified through RNA immunoprecipitation, RNA pull-down, and dual-luciferase reporter gene assay.
METTL3 and TUG1 expression levels decreased in the kidneys of diabetic mice and HG-treated HK-2 cells. Mechanistically, METTL3-mediated mA modification increased the stability of TUG1 in an insulin-like growth factor 2 mRNA binding protein 2 (IGF2BP2)-dependent manner. METTL3-mediated mA modification of TUG1 promotes PGC-1α activation, thereby alleviating mitochondrial dysfunction in HG-treated HK-2 cells and db/db mice. Moreover, METTL3 overexpression alleviated kidney injury in db/db mice.
METTL3 targets TUG1/PGC-1α and ameliorates mitochondrial dysfunction in diabetic nephropathy in an IGF2BP2-dependent manner.
探讨牛磺酸上调基因1（TUG1）的N6-甲基腺苷（m⁶A）修饰机制，以及甲基转移酶3（METTL3）是否能促进过氧化物酶体增殖物激活受体γ共激活因子1α（PGC-1α）转录并减轻线粒体功能障碍。
建立高糖（HG）处理的HK-2细胞模型和经尾静脉注射rAAV-METTL3的db/db小鼠模型。通过RT-qPCR、蛋白质免疫印迹和免疫组织化学染色测定表达水平。用RNase Mazf分析RNA m⁶A修饰。通过酶联免疫吸附测定法检测小鼠的生化指标。通过流式细胞术检测细胞凋亡。进行组织病理学染色以评估肾损伤。分别根据制造商的说明，通过RT-qPCR和检测试剂盒检测线粒体DNA含量、线粒体复合物活性和ATP。分别在荧光显微镜和透射电子显微镜下观察用MitoSOX Red孵育的HK-2细胞中线粒体活性氧的产生和线粒体形态。通过RNA免疫沉淀、RNA下拉和双荧光素酶报告基因测定验证分子相互作用。
糖尿病小鼠肾脏和HG处理的HK-2细胞中METTL3和TUG1表达水平降低。机制上，METTL3介导的m⁶A修饰以胰岛素样生长因子-2 mRNA结合蛋白2（IGF2BP2）依赖的方式增加了TUG1的稳定性。METTL3介导的TUG1的m⁶A修饰促进PGC-1α激活，从而减轻HG处理的HK-2细胞和db/db小鼠中的线粒体功能障碍。此外，METTL3过表达减轻了db/db小鼠的肾损伤。
METTL3以IGF2BP2依赖的方式靶向TUG1/PGC-1α并改善糖尿病肾病中的线粒体功能障碍。
