Yu Zonggang, Xu Xueli, Ai Nini, Wang Kaiming, Zhang Peiwen, Li Xintong, LiuFu Sui, Liu Xiaolin, Jiang Jun, Gu Jingjing, Gao Ning, Ma Haiming
College of Animal Science and Technology, Hunan Agricultural University, Changsha, China.
Guangdong Laboratory for Lingnan Modern Agriculture, Guangzhou, China.
Front Cell Dev Biol. 2023 Jul 3;11:1185823. doi: 10.3389/fcell.2023.1185823. eCollection 2023.
The development of skeletal muscle is regulated by regulatory factors of genes and non-coding RNAs (ncRNAs). The objective of this study was to understand the transformation of muscle fiber type in the longissimus dorsi muscle of male Ningxiang pigs at four different growth stages (30, 90, 150, and 210 days after birth, n = 3) by histological analysis and whole transcriptome sequencing. Additionally, the study investigated the expression patterns of various RNAs involved in muscle fiber transformation and constructed a regulatory network for competing endogenous RNA (ceRNA) that includes circular RNA (circRNA)/long non-coding RNA (lncRNA)-microRNA (miRNA)-messenger RNA (mRNA). Histomorphology analysis showed that the diameter of muscle fiber reached its maximum at 150 days after birth. The slow muscle fiber transformation showed a pattern of initial decrease followed by an increase. 29,963 circRNAs, 2,683 lncRNAs, 986 miRNAs and 22,411 mRNAs with expression level ≥0 were identified by whole transcriptome sequencing. Furthermore, 642 differentially expressed circRNAs (DEc), 505 differentially expressed lncRNAs (DEl), 316 differentially expressed miRNAs (DEmi) and 6,090 differentially expressed mRNAs (DEm) were identified by differential expression analysis. Functions of differentially expressed mRNA were identified by gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG). GO enrichment analysis indicates that 40 known genes and 6 new genes are associated with skeletal muscle development. Additionally, KEGG analysis shows that these genes regulate skeletal muscle development via MAPK, FoxO, Hedgehog, PI3K-Akt, Notch, VEGF and other signaling pathways. Through protein-protein interaction (PPI) and transcription factor prediction (TFP), the action mode of skeletal muscle-related genes was explored. PPI analysis showed that there were stable interactions among 19 proteins, meanwhile, TFP analysis predicted 22 transcription factors such as HMG20B, MYF6, MYOD1 and MYOG, and 12 of the 19 interacting proteins were transcription factors. The regulatory network of ceRNA related to skeletal muscle development was constructed based on the correlation of various RNA expression levels and the targeted binding characteristics with miRNA. The regulatory network included 31 DEms, 59 miRNAs, 667 circRNAs and 224 lncRNAs. Overall, the study revealed the role of ceRNA regulatory network in the transformation of skeletal muscle fiber types in Ningxiang pigs, which contributes to the understanding of ceRNA regulatory network in Ningxiang pigs during the skeletal muscle development period.
骨骼肌的发育受基因调控因子和非编码RNA（ncRNAs）的调节。本研究的目的是通过组织学分析和全转录组测序，了解雄性宁乡猪在四个不同生长阶段（出生后30、90、150和210天，n = 3）背最长肌中肌纤维类型的转变。此外，该研究调查了参与肌纤维转变的各种RNA的表达模式，并构建了一个竞争性内源RNA（ceRNA）调控网络，该网络包括环状RNA（circRNA）/长链非编码RNA（lncRNA）-微小RNA（miRNA）-信使RNA（mRNA）。组织形态学分析表明，肌纤维直径在出生后150天达到最大值。慢肌纤维转变呈现出先下降后上升的模式。通过全转录组测序鉴定出表达水平≥0的29,963个circRNA、2,683个lncRNA、986个miRNA和22,411个mRNA。此外，通过差异表达分析鉴定出642个差异表达的circRNA（DEc）、505个差异表达的lncRNA（DEl）、316个差异表达的miRNA（DEmi）和6,090个差异表达的mRNA（DEm）。通过基因本体论（GO）和京都基因与基因组百科全书（KEGG）鉴定差异表达mRNA的功能。GO富集分析表明，40个已知基因和6个新基因与骨骼肌发育相关。此外，KEGG分析表明，这些基因通过丝裂原活化蛋白激酶（MAPK）、叉头框蛋白O（FoxO）、刺猬信号通路（Hedgehog）、磷脂酰肌醇-3激酶-蛋白激酶B（PI3K-Akt）、Notch、血管内皮生长因子（VEGF）等信号通路调节骨骼肌发育。通过蛋白质-蛋白质相互作用（PPI）和转录因子预测（TFP），探索了骨骼肌相关基因的作用模式。PPI分析表明，19种蛋白质之间存在稳定的相互作用，同时，TFP分析预测了22种转录因子，如高迁移率族蛋白20B（HMG20B）、肌细胞生成素（MYF6）、肌分化抗原（MYOD1）和肌生成调节因子（MYOG），19种相互作用蛋白中有12种是转录因子。基于各种RNA表达水平的相关性以及与miRNA的靶向结合特性，构建了与骨骼肌发育相关的ceRNA调控网络。该调控网络包括31个DEm、59个miRNA、667个circRNA和224个lncRNA。总体而言，该研究揭示了ceRNA调控网络在宁乡猪骨骼肌纤维类型转变中的作用，这有助于了解宁乡猪骨骼肌发育时期的ceRNA调控网络。
