Gene expression data of Lin28a-positive MuSCs using RNA-seq and RIP-seq

During the early stages of embryonic development, Lin28a is expressed at high levels and gradually decreases as the embryo develops. As an RNA-binding protein, Lin28a maintains a subset of adult muscle stem cells (MuSCs) in an embryonic-like state. However, the specific mechanism for regulating RNA metabolism is not yet clear. Through the analysis of Lin28a-associated genes, we have revealed that Lin28a promotes the expression of Igf2bp3 and interacts with the Igf2bp3 protein, controlling the proliferation of MuSCs. In response to stress stimuli, Lin28a rapidly upregulates the expression of Igf2bp3, recruits mRNAs by interacting with the N6-methyladenosine (m6A) reader Igf2bp3, and forms protein complexes with G3bp1 in stress granules. Sequencing of the transcriptome and RNAs immunoprecipitated by Lin28a, Igf2bp3, and m6A antibodies in Lin28a+ MuSCs further revealed that Lin28a and Igf2bp3 collaboratively regulate the expression of DNA repair-related genes such as Fancm and Usp1 to promote DNA repair after oxidative stress. Therefore, Lin28a regulates the expression of DNA damage repair-related genes and upregulates the DNA stress response of MuSCs through stress granule regulation of m6A-modified mRNAs. This positive regulation contributes to the self-renewal of MuSCs. To investigate the crucial role of lin28a in MuSCs, we assessed the gene expression data of Lin28a-positive MuSCs using RNA-seq and RIP-seq methods.在胚胎发育早期，Lin28a呈高表达状态，且随胚胎发育进程逐渐下调。作为一种RNA结合蛋白，Lin28a可维持成体肌肉干细胞（Muscle Stem Cells, MuSCs）的亚群处于类胚胎状态。然而，其调控RNA代谢的具体分子机制目前仍不明确。通过对Lin28a关联基因的分析，本研究发现Lin28a可促进Igf2bp3的表达，并与Igf2bp3蛋白相互作用，进而调控MuSCs的增殖。在应激刺激下，Lin28a会快速上调Igf2bp3的表达，通过与N6-甲基腺嘌呤（N6-methyladenosine, m6A）识别蛋白Igf2bp3结合招募mRNA，并在应激颗粒中与G3bp1形成蛋白复合物。对Lin28a阳性MuSCs中Lin28a、Igf2bp3及m6A抗体免疫沉淀的RNA与转录组进行测序后，本研究进一步揭示，Lin28a与Igf2bp3可协同调控Fancm、Usp1等DNA修复相关基因的表达，从而促进氧化应激后的DNA修复。因此，Lin28a通过应激颗粒调控m6A修饰的mRNA，进而调控DNA损伤修复相关基因的表达，并增强MuSCs的DNA应激响应能力。这一正向调控机制可促进MuSCs的自我更新。为探究Lin28a在MuSCs中的关键作用，本研究采用RNA测序（RNA-seq）与RNA免疫沉淀测序（RIP-seq）技术，对Lin28a阳性MuSCs的基因表达数据进行了分析。