METTL3 inhibits inflammation of retinal pigment epithelium cells by regulating NR2F1 in an m6A-dependent manner [meRIP-seq]

To investigate the fuction of METTL3 in autoimmune uveitis, we established ARPE-19 cell lines in which target gene has been knocked down by shRNA. In this study, we explored the effects and underlying mechanisms of methyltransferase-like 3 (METTL3) in retinal pigment epithelium (RPE) cells, which are crucial for the pathogenesis of autoimmune uveitis. Our results showed that the expression of METTL3 was increased in both human RPE (hRPE) cells and ARPE19 cells after lipopolysaccharide (LPS) stimulation. Moreover, these two types of RPE cells exhibited inhibited proliferation and increased barrier destruction and inflammatory factor secretion after METTL3 silencing. Mechanistically, we found that NR2F1, as a METTL3-methylated target gene, promotes the inflammation of RPE cells in an m6A-dependent manner. Interestingly, NR2F1 deficiency reversed the increased inflammation in the METTL3-defective RPE cells. In conclusion, our study revealed that METTL3 attenuates RPE cell inflammation by methylating NR2F1, suggesting the critical role of METTL3 in RPE cells.

为探究甲基转移酶样3（METTL3）在自身免疫性葡萄膜炎（autoimmune uveitis）中的功能，我们构建了通过短发夹RNA（short hairpin RNA，shRNA）敲低靶基因的ARPE-19细胞株。本研究聚焦于甲基转移酶样3（METTL3）在视网膜色素上皮（retinal pigment epithelium，RPE）细胞中的作用及潜在机制——RPE细胞是自身免疫性葡萄膜炎发病过程中的关键细胞群。研究结果显示，经脂多糖（lipopolysaccharide，LPS）刺激后，人视网膜色素上皮（hRPE）细胞与ARPE-19细胞中METTL3的表达水平均显著升高。此外，在METTL3沉默后，这两类RPE细胞均表现出增殖受抑、屏障功能破坏加剧以及炎症因子分泌增加的表型。机制研究表明，作为METTL3介导甲基化的靶基因，NR2F1可通过N6-甲基腺嘌呤（m6A）依赖的方式促进RPE细胞的炎症反应。值得注意的是，敲低NR2F1可逆转METTL3缺陷型RPE细胞中炎症反应增强的表型。综上，本研究证实METTL3可通过对NR2F1进行甲基化修饰以减轻RPE细胞的炎症反应，提示METTL3在RPE细胞中发挥关键调控作用。