Qian Benheng, Wang Ping, Zhang Donghong, Wu Lianpin
Department of Cardiology, The Second Affiliated Hospital of Wenzhou Medical University, 109 Xueyuan Road, Wenzhou, 325027, Zhejiang, PR China.
School of Medical Imaging, Tianjin Medical University, Tianjin, 300203, China.
Cell Death Discov. 2021 Jun 26;7(1):157. doi: 10.1038/s41420-021-00552-7.
Both N6-methyladenosine (m6A) RNA modification and microRNAs (miRNAs) are common regulatory mechanisms for gene post-transcription by modulating mRNA stability and translation. They also share the same 3'-untranslated regions (UTRs) regions for their target gene. However, little is known about their potential interaction in cell development and biology. Here, we aimed to investigate how m6A regulates the specific miRNA repression during cardiac development and hypertrophy. Our multiple lines of bioinformatic and molecular biological evidence have shown that m6A modification on cardiac miR-133a target sequence promotes miR-133a repressive effect via AGO2-IGF2BP2 (Argonaute 2-Insulin-like growth factor 2 mRNA binding protein 2) complex. Among 139 cardiac miRNAs, only the seed sequence of miR-133a was inversely complement to m6A consensus motif "GGACH" by sequence alignment analysis. Immunofluorescence staining, luciferase reporter, and m6A-RIP (RNA immunoprecipitation) assays revealed that m6A modification facilitated miR-133a binding to and repressing their targets. The inhibition of the miR-133a on cardiac proliferation and hypertrophy could be prevented by silencing of Fto (FTO alpha-ketoglutarate dependent dioxygenase) which induced m6A modification. IGF2BP2, an m6A binding protein, physically interacted with AGO2 and increased more miR-133a accumulation on its target site, which was modified by m6A. In conclusion, our study revealed a novel and precise regulatory mechanism that the m6A modification promoted the repression of specific miRNA during heart development and hypertrophy. Targeting m6A modification might provide a strategy to repair hypertrophic gene expression induced by miR-133a.
N6-甲基腺苷（m6A）RNA修饰和微小RNA（miRNA）都是通过调节mRNA稳定性和翻译来进行基因转录后调控的常见机制。它们的靶基因也共享相同的3'-非翻译区（UTR）区域。然而，关于它们在细胞发育和生物学中的潜在相互作用却知之甚少。在这里，我们旨在研究m6A如何在心脏发育和肥大过程中调节特定miRNA的抑制作用。我们多方面的生物信息学和分子生物学证据表明，心脏miR-133a靶序列上的m6A修饰通过AGO2-IGF2BP2（Argonaute 2-胰岛素样生长因子2 mRNA结合蛋白2）复合物促进miR-133a的抑制作用。通过序列比对分析，在139种心脏miRNA中，只有miR-133a的种子序列与m6A共有基序“GGACH”反向互补。免疫荧光染色、荧光素酶报告基因和m6A-RIP（RNA免疫沉淀）分析表明，m6A修饰促进了miR-133a与其靶标的结合并抑制其靶标。沉默诱导m6A修饰的Fto（FTOα-酮戊二酸依赖性双加氧酶）可阻止miR-133a对心脏增殖和肥大的抑制作用。IGF2BP2是一种m6A结合蛋白，与AGO2发生物理相互作用，并增加了更多在其靶位点上被m6A修饰的miR-133a积累。总之，我们的研究揭示了一种新的精确调控机制，即m6A修饰在心脏发育和肥大过程中促进特定miRNA的抑制作用。靶向m6A修饰可能为修复由miR-133a诱导的肥厚基因表达提供一种策略。
