The skeletal muscle is the largest organ in animals. In pigs, skeletal muscles have important economic significance for production, and understanding the development of skeletal muscles is important for improving productivity and meat quality. Previous studies have detected thousands of lincRNAs in skeletal muscle, but only a few number of lincRNAs have been characterized. In this study, 361 potential lincRNAs were identified on the basis of the designed pipeline, of which 53 lincRNAs were novel lincRNAs. Thereafter, 138 DELs were statistically obtained using the DeSeq2 software, of which 66 DELs were upregulated in the LW group and 72 were downregulated. The transcription length, exon length, number and expression characteristics of potential lincRNA and coding genes were compared and found to be consistent with the characteristics of other mammalian lincRNAs. The accuracy of our pipeline identification lincRNA was verified from the other hand.
The PTGs of DELs were predicted, and gene ontology analysis was used to understand the biological processes that PTGs may participate in to predict the possible role of corresponding DELs. Multiple biological processes are associated with skeletal muscle development. Such processes include muscle structure development, muscle organ development, muscle cell differentiation, and skeletal muscle tissue development. DELs may indirectly affect the development of skeletal muscle by regulating its target genes. In general, multiple genes need to cooperate with one another to play the ultimate biological function; thus, Kyoto Encyclopedia of Genes and Genomes pathway analysis was performed on PTGs. Path-based analysis is used to understand the possible role of DELs. Multiple genes were speculated to participate in the cAMP signaling pathway, the mTOR signaling pathway, and the Rap1 signaling pathway. QTL mapping analysis was performed to improve the credibility of DEL potential functional prediction. The results show that 1137 of the predicted 3018 QTLs are associated with skeletal muscle development. The proportion of loin muscle area QTL and body weight QTL is the highest. The muscle development-related PTGs were studied by generating the MDRPTG–DEL co-expression network. We then focused on muscle development related PTGs. Generating the MDRPTG–DEL co-expression network.
Based on the above analysis results, we comprehensive analysis found that multiple DELs may participate in the IGF-1-Akt-mTOR signaling pathway by regulating their PTGs. The growth of skeletal muscle depends on muscle fiber hypertrophy, and the size of muscle fibers is increased when the rate of protein synthesis is higher than the rate of degradation. Under normal physiological conditions, the IGF-1-Akt-mTOR pathway plays a key regulatory role in skeletal muscle protein synthesis[19, 20]. Interestingly, multiple PTGs are related to this pathway.
Mitochondrial calcium unidirectional transporter (MCU) is a highly selective channel for Ca2+ transport into the mitochondria. Mammucari et al. reported that MCU participates in IGF-1-Akt-mTOR signaling by increasing Ca2+ level in the mitochondria, activating the PGC-1α4, which is a transcriptional coactivator; the IGF-1 gene is activated through the PGC-1α4, leading to muscle hypertrophy[21, 22]. In the present study, our analysis showed that compared with the TP group, the LW group had higher MCU expression level (Figure.S3), which may be associated with the growth characteristics of Yorkshire pig breeds. More importantly, we found that DEL-MSTRG.8035 is positively related to the expression of MCU and highly expressed in the LW group. Insulin-like growth factor 2 (IGF-2) is a maternal blotting growth factor that regulates prenatal skeletal muscle development. It can be involved in the IGF1-Akt-mTOR signaling pathway by activating the IGF1 receptor. A significant positive correlation exists between the DEL-MSTRG.12010 and IGF-2, which were upregulated in the LW group. Interestingly, MSTRG.12010 was significantly negatively correlated with the troponin T-3 (TNNT3) gene. Wang et al. It is predicted that TNNT3 can regulate muscle growth and muscle fibers. TNNT3 is an important part of pig skeletal muscle filaments, that can affect the taste and tenderness of pork[26, 27]. Its expression level was low in the LW group. This may, on the other hand, find the reason for the decrease in meat quality as the growth rate increases. In addition, a potential target gene, serum response factor (SRF), plays an important role in controlling muscle fiber hypertrophy[28, 29]. SRF can control the transcription of miR-486, which as a potential regulator of PI3K/Akt signal transduction in muscle cells, can phosphorylate Akt and activate the IGF-1-Akt-mTOR signaling pathway, leading to muscle fiber hypertrophy. The co-expression network, suggests that DEL-MSTRG.21771 is significantly positively correlated with SRF expression and highly expressed in the LW group. It is speculated that MSTRG.21771 regulates the high expression of its potential target gene SRF, in the LW group, which possibly useful in maintaining the fast skeletal muscle rate in Yorkshire pig. PLD1 is an isoform of phospholipase D (PLD), which can stimulate phosphatidylcholine (PC) to produce phosphatidic acid (PA), which can bind to mTOR and participate in the IGF-1-Akt-mTOR signaling pathway. The substrate S6K1 of mTORC1 is phosphorylated to enhance protein translation, resulting in muscle fiber hypertrophy. Furthermore, DEL-MSTRG.6293 was positively correlated with its expression, and highly expressed in the LW group. QTL results indicated that MSTRG.8035, MSTRG12010, MSTRG21771, and MSTRG.6293 were all mapped to the QTL loci, such as Loin weight QTL, Loin muscle area QTL, and backfat above muscle dorsi QTL, which are related to muscle development. It is speculated that these DELs may be related to skeletal muscle development, and participate in the IGF-1-Akt-mTOR signaling pathway by regulating the expression of its potential target genes, thereby affecting the muscle fibrous hypertrophy process. However, the specific molecular regulation of this phenomenon remains unclear,and further studies are needed. Figure 8 shows that DELs may affect muscle protein synthesis by regulating their PTGs to participate in the IGF-1-Akt-mTOR pathway.