Table_11_Integration of RNA-seq and ATAC-seq identifies muscle-regulated hub genes in cattle.XLS

As the main product of livestock, muscle itself plays an irreplaceable role in maintaining animal body movement and regulating metabolism. Therefore, it is of great significance to explore its growth, development and regeneration to improve the meat yield and quality of livestock. In this study, we attempted to use RNA-seq and ATAC-seq techniques to identify differentially expressed genes (DEGs) specifically expressed in bovine skeletal muscle as potential candidates for studying the regulatory mechanisms of muscle development. Microarray data from 8 tissue samples were selected from the GEO database for analysis. First, we obtained gene modules related to each tissue through WGCNA analysis. Through Gene Ontology (GO) functional annotation, the module of lightyellow (MElightyellow) was closely related to muscle development, and 213 hub genes were screened as follow-up research targets. Further, the difference analysis showed that, except for PREB, all other candidate hub genes were up-regulated (muscle group vs. other-group). ATAC-seq analysis showed that muscle-specific accessible chromatin regions were mainly located in promoter of genes related to muscle structure development (GO:0061061), muscle cell development (GO:0055001) and muscle system process (GO:0003012), which were involved in cAMP, CGMP-PKG, MAPK, and other signaling pathways. Next, we integrated the results of RNA-seq and ATAC-seq analysis, and 54 of the 212 candidate hub genes were identified as key regulatory genes in skeletal muscle development. Finally, through motif analysis, 22 of the 54 key genes were found to be potential target genes of transcription factor MEF2C. Including CAPN3, ACTN2, MB, MYOM3, SRL, CKM, ALPK3, MAP3K20, UBE2G1, NEURL2, CAND2, DOT1L, HRC, MAMSTR, FSD2, LRRC2, LSMEM1, SLC29A2, FHL3, KLHL41, ATXN7L2, and PDRG1. This provides a potential reference for studying the molecular mechanism of skeletal muscle development in mammals.

作为畜禽的主要产物，肌肉本身在维持动物机体运动与调节代谢过程中发挥着不可替代的作用。因此，探究其生长、发育与再生机制，对于提升畜禽肉产量与品质具有重要意义。本研究尝试借助RNA测序（RNA-seq）与转座酶可及性测序（ATAC-seq）技术，筛选牛骨骼肌特异性表达的差异表达基因（DEGs），作为探究肌肉发育调控机制的潜在候选靶点。我们从基因表达综合数据库（GEO）中选取8份组织样本的微阵列数据开展分析：首先通过加权基因共表达网络分析（WGCNA）获得与各组织相关的基因模块；经基因本体（GO）功能注释后，发现浅黄色模块（MElightyellow）与肌肉发育密切相关，并筛选出213个核心基因作为后续研究靶点。进一步差异分析结果显示，除PREB基因外，其余候选核心基因在肌肉组与其他组的对比中均呈上调表达。转座酶可及性测序分析表明，肌肉特异性可及染色质区域主要位于肌肉结构发育（GO:0061061）、肌细胞发育（GO:0055001）及肌肉系统过程（GO:0003012）相关基因的启动子区域，这些区域涉及cAMP、CGMP-PKG、MAPK等信号通路。随后，我们整合RNA测序与转座酶可及性测序的分析结果，在212个候选核心基因中鉴定出54个骨骼肌发育关键调控基因。最后通过基序分析，发现54个关键基因中的22个为转录因子MEF2C的潜在靶基因，包括CAPN3、ACTN2、MB、MYOM3、SRL、CKM、ALPK3、MAP3K20、UBE2G1、NEURL2、CAND2、DOT1L、HRC、MAMSTR、FSD2、LRRC2、LSMEM1、SLC29A2、FHL3、KLHL41、ATXN7L2及PDRG1。本研究为解析哺乳动物骨骼肌发育的分子机制提供了潜在参考依据。