Zhong Yulun, Zhang Rou, Lu Lingzi, Tan Huijian, You Yuyu, Mao Yang, Yuan Yanqiu
School of Pharmaceutical Sciences, Sun Yat-Sen University, Guangzhou, China.
Guangdong Provincial Key Laboratory of Drug Non-Clinical Evaluation and Research, Guangzhou, China.
Cell Commun Signal. 2025 Mar 8;23(1):126. doi: 10.1186/s12964-025-02130-1.
Protein arginine methylations are crucial post-translational modifications (PTMs) in eukaryotes, playing a significant regulatory role in diverse biological processes. Here, we present our investigation into the detailed arginine methylation pattern of the C-terminal RG-rich region of METTL14, a key component of the m6A RNA methylation machinery, and its functional implications in biology and disease.
Using ETD-based mass spectrometry and in vitro enzyme reactions, we uncover a specific arginine methylation pattern on METTL14. RNA methyltransferase activity assays were used to assess the impact of sDMA on METTL3:METTL14 complex activity. RNA immunoprecipitation was used to evaluate mRNA-m6A reader interactions. MeRIP-seq analysis was used to study the genome-wide effect of METTL14 sDMA on m6A modification in acute myeloid leukemia cells.
We demonstrate that PRMT5 catalyzes the site-specific symmetric dimethylation at R425 and R445 within the extensively methylated RGG/RG motifs of METTL14. We show a positive regulatory role of symmetric dimethylarginines (sDMA) in the catalytic efficiency of the METTL3:METTL14 complex and m6A-specific gene expression in HEK293T and acute myeloid leukemia cells, potentially through the action of m6A reader protein YTHDF1. In addition, the combined inhibition of METTL3 and PRMT5 further reduces the expression of several m6A substrate genes essential for AML proliferation, suggesting a potential therapeutic strategy for AML treatment.
The study confirms the coexistence of sDMA and aDMA modifications on METTL14's RGG/RG motifs, with sDMA at R425 and R445 enhancing METTL3:METTL14's catalytic efficacy and regulating gene expression through m6A deposition in cancer cells.
蛋白质精氨酸甲基化是真核生物中至关重要的翻译后修饰（PTM），在多种生物学过程中发挥着重要的调节作用。在此，我们展示了对METTL14 C末端富含RG区域的详细精氨酸甲基化模式的研究，METTL14是m6A RNA甲基化机制的关键组成部分，以及其在生物学和疾病中的功能意义。
使用基于电子转移解离（ETD）的质谱和体外酶反应，我们揭示了METTL14上特定的精氨酸甲基化模式。RNA甲基转移酶活性测定用于评估对称二甲基精氨酸（sDMA）对METTL3:METTL14复合物活性的影响。RNA免疫沉淀用于评估mRNA与m6A阅读蛋白的相互作用。甲基化RNA免疫沉淀测序（MeRIP-seq）分析用于研究METTL14的sDMA对急性髓系白血病细胞中m6A修饰的全基因组效应。
我们证明蛋白精氨酸甲基转移酶5（PRMT5）催化METTL14广泛甲基化的RGG/RG基序内R425和R445位点的位点特异性对称二甲基化。我们发现对称二甲基精氨酸（sDMA）在HEK293T细胞和急性髓系白血病细胞中对METTL3:METTL14复合物的催化效率和m6A特异性基因表达具有正向调节作用，可能是通过m6A阅读蛋白YTHDF1的作用。此外，METTL3和PRMT5的联合抑制进一步降低了几种对急性髓系白血病增殖至关重要的m6A底物基因的表达，提示了一种潜在的急性髓系白血病治疗策略。
该研究证实了METTL14的RGG/RG基序上存在sDMA和不对称二甲基精氨酸（aDMA）修饰，R425和R445位点的sDMA增强了METTL3:METTL14的催化效力，并通过癌细胞中的m6A沉积调节基因表达。
