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CRISPR Screen Identifies the RNA-Binding Protein Eef1a1 as a Key Regulator of Myogenesis

文献类型： 外文期刊

作者： Weiwei Liu;Wei Wang;Zishuai Wang;Xinhao Fan;Wangchang Li;Yuxin Huang;Xiaogan Yang;Zhonglin Tang

作者机构：

关键词： （2-1-4）CRISPR screen;Eef1a1;myogenesis;RNA-binding protein

期刊名称： International Journal of Molecular Sciences

ISSN： 1661-6596

年卷期： 2024 年 25 卷 9 期

页码：

收录情况： SCIE（2024版）

摘要： Skeletal muscle myogenesis hinges on gene regulation, meticulously orchestrated by molecular mechanisms. While the roles of transcription factors and non-coding RNAs in myogenesis are widely known, the contribution of RNA-binding proteins (RBPs) has remained unclear until now. Therefore, to investigate the functions of post-transcriptional regulators in myogenesis and uncover new functional RBPs regulating myogenesis, we employed CRISPR high-throughput RBP-KO (RBP-wide knockout) library screening. Through this approach, we successfully identified Eef1a1 as a novel regulatory factor in myogenesis. Using CRISPR knockout (CRISPRko) and CRISPR interference (CRISPRi) technologies, we successfully established cellular models for both CRISPRko and CRISPRi. Our findings demonstrated that Eef1a1 plays a crucial role in promoting proliferation in C2C12 myoblasts. Through siRNA inhibition and overexpression methods, we further elucidated the involvement of Eef1a1 in promoting proliferation and suppressing differentiation processes. RIP (RNA immunoprecipitation), miRNA pull-down, and Dual-luciferase reporter assays confirmed that miR-133a-3p targets Eef1a1. Co-transfection experiments indicated that miR-133a-3p can rescue the effect of Eef1a1 on C2C12 myoblasts. In summary, our study utilized CRISPR library high-throughput screening to unveil a novel RBP, Eef1a1, involved in regulating myogenesis. Eef1a1 promotes the proliferation of myoblasts while inhibiting the differentiation process. Additionally, it acts as an antagonist to miR-133a-3p, thus modulating the process of myogenesis.

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