Single-cell analysis reveals an indispensable role of SRSF2 in controlling fate of Myf5-expressing cells during skeletal muscle development

Skeletal muscle is an important tissue of animal body, which plays an important role in the process of motion, support and energy metabolism. Skeletal muscles are composed of striated myofibers bundles which formed during embryonic myogenesis and may undergo remodeling in the process of muscle regeneration after birth. SRSF2 is an important member of the splicing factor SR protein family. It plays an active role in transcription elongation, coupling transcription and splicing. A large number of studies have shown that it plays a key role in maintaining the normal development of thymus, heart and bone marrow. Here, we have used single cell RNA-seq and lineage analysis and gene knockout animal model to explore the regulation of splicing factor SRSF2 on myoblast cell fate and differentiation, as well as the mechanism in mouse skeletal muscle development. scRNA-seq: The SRSF2f/f mice were crossed with transgenic C57BL/6J mice expressing Cre recombinase driven by the Myf5 (myogenic factor 5) promoter to generate progeny (SRSF2f/w/Myf5-Cre). The SRSF2f/f mice were crossed with Rosa26-LSL-tdtomato (hereafter tdT) mice. After generated SRSF2f/f/tdT mice (both SRSF2 and tdT are homozygous). Next, mating SRSF2f/f/tdT with SRSF2f/w/Myf5-Cre mice to generate SRSF2f/f/Myf5-Cre/tdT mice. SRSF2f/w/Myf5-Cre/tdT littermates served as controls.骨骼肌是动物机体的重要组织，在运动、支撑及能量代谢过程中发挥关键作用。骨骼肌由横纹肌纤维束构成，此类纤维束于胚胎肌发生阶段形成，并可在出生后肌肉再生过程中发生重塑。 剪接因子SRSF2是SR蛋白家族的重要成员，在转录延伸、转录与剪接的偶联过程中发挥积极调控作用。大量研究表明，其在维持胸腺、心脏及骨髓的正常发育中具有关键作用。 本研究采用单细胞RNA测序（single cell RNA-seq）、谱系分析及基因敲除动物模型，探究了剪接因子SRSF2对小鼠骨骼肌发育过程中成肌细胞命运与分化的调控作用及其分子机制。 单细胞RNA测序相关实验流程如下：将SRSF2f/f小鼠与表达由Myf5（肌因子5，myogenic factor 5）启动子驱动的Cre重组酶的转基因C57BL/6J小鼠杂交，获得子代小鼠（SRSF2f/w/Myf5-Cre）。将SRSF2f/f小鼠与Rosa26-LSL-tdtomato（以下简称tdT）小鼠杂交，构建得到SRSF2f/f/tdT小鼠（SRSF2与tdT均为纯合子）。随后将SRSF2f/f/tdT小鼠与SRSF2f/w/Myf5-Cre小鼠交配，获得SRSF2f/f/Myf5-Cre/tdT小鼠。以同窝出生的SRSF2f/w/Myf5-Cre/tdT小鼠作为实验对照组。