The quality of broiler meat affects consumers’ purchasing decisions. Numerous studies have shown that phosphorylation of proteins in muscle can affect muscle quality. Here, metabolomics and transcriptomics were used to systematically identify the genetic regulation of differences in meat flavor among different broiler. By constructing the meat flavor-related metabolite-gene networks, we identified that protein phosphatase magnesium/manganese-dependent 1J (PPM1J), which is known to regulate a range of biological processes by modulating reversible protein phosphorylation, was a differentially expressed gene with the highest connectivity to meat flavor-related metabolites. Gain- and loss-of-function analysis revealed that PPM1J induced muscular atrophy, improved meat quality and regulated the composition of glycerophospholipids. More importantly, phosphoproteome and metabolome results found that PPM1J participates in the regulation of meat quality feature and glycerophospholipids composition by catalyzing protein dephosphorylation. Our study provides a basis for further understanding the molecular mechanism of meat quality feature and glycerophospholipids composition in broiler.

Single nucleotide polymorphisms (SNPs) are valuable genetic markers that can provide insights into the genetic diversity and variation within chicken populations. In poultry breeding, SNP analysis is widely utilized to accelerate the selection of desirable traits, improving the efficiency and effectiveness of chicken breeding programs. In our previous research, we identified an association between LncEDCH1 and muscle development. To further investigate its specific mechanism, we conducted SNP detection and performed genotyping, linkage disequilibrium, and haplotype analysis. Our research findings indicate that 16 SNPs in the LncEDCH1. Among these SNPs, g.1703497 C>T and g.1704262 C>T were significantly associated with breast muscle weight percentage, g.1703497 C>T and g.1703613 T>C were significantly associated with leg weight percentage, and g.1703497 C>T, g.1703589 T>C, g.1703613 T>C, g.1703636 C>A, g.1703768 T>C, g.1704079 C>T, g.1704250 T>C, g.1704253 G>A were significantly associated with skin yellowness. Two haplotype blocks composed of 6 SNPs that were significantly associated with wing skin yellowness, breast skin yellowness, full-bore weight, and carcass weight percentage. Furthermore, through dual-luciferase reporter assays, biotin-coupled miRNA pull-down assays, 5-ethynyl-2'-deoxyuridine (EDU) assays, immunofluorescence, and quantitative real-time polymerase chain reaction (qPCR), it has been confirmed that miR-196-2-3p inhibits the expression of LncEDCH1 directly by binding to LncEDCH1 g.1703613T>C, thereby achieving indirect regulation of muscle development. These findings provide valuable molecular markers for chicken molecular breeding and broaden our understanding of the regulatory mechanisms.

Background Skeletal muscle development is pivotal for animal growth and health. Recently, long noncoding RNAs (lncRNAs) were found to interact with chromatin through diverse roles. However, little is known about how lncRNAs act as chromatin-associated RNAs to regulate skeletal muscle development. Here, we aim to investigate the regulation of chromatin-associated RNA (MYH1G-AS) during skeletal muscle development. Methods We provided comprehensive insight into the RNA profile and chromatin accessibility of different myofibers, combining RNA sequencing (RNA-seq) with an assay for transposase-accessible chromatin with high-throughput sequencing (ATAC-seq). The dual-luciferase reporter assay and chromatin immunoprecipitation (ChIP) assay were used to analyze the transcriptional regulation mechanism of MYH1G-AS. ALKBH5 -mediated MYH1G-AS N 6 -methyladenosine (m 6 A) demethylation was assessed by a single-base elongation and ligation-based qPCR amplification method (SELECT) assay. Functions of MYH1G-AS were investigated through a primary myoblast and lentivirus/cholesterol-modified antisense oligonucleotide (ASO)-mediated animal model. To validate the interaction of MYH1G-AS with fibroblast growth factor 18 (FGF18) protein, RNA pull down and an RNA immunoprecipitation (RIP) assay were performed. Specifically, the interaction between FGF18 and SWI/SNF-related matrix-associated actin-dependent regulator of chromatin subfamily A member 5 (SMARCA5) protein was analyzed by coimmunoprecipitation (Co-IP) and a yeast two-hybrid assay. Results A total of 45 differentially expressed (DE) lncRNAs, with DE ATAC-seq peaks in their promoter region, were classified as open chromatin-associated lncRNAs. A skeletal muscle-specific lncRNA (MSTRG.15576.9; MYH1G-AS), which is one of the open chromatin-associated lncRNA, was identified. MYH1G-AS transcription is coordinately regulated by transcription factors (TF) SMAD3 and SP2. Moreover, SP2 represses ALKBH5 transcription to weaken ALKBH5 -mediated m 6 A demethylation of MYH1G-AS, thus destroying MYH1G-AS RNA stability. MYH1G-AS accelerates myoblast proliferation but restrains myoblast differentiation. Moreover, MYH1G-AS drives a switch from slow-twitch to fast-twitch fibers and causes muscle atrophy. Mechanistically, MYH1G-AS inhibits FGF18 protein stabilization to reduce the interaction of FGF18 to SMARCA5, thus repressing chromatin accessibility of the SMAD4 promoter to activate the SMAD4 -dependent pathway. Conclusions Our results reveal a new pattern of the regulation of lncRNA expression at diverse levels and help expound the regulation of m 6 A methylation on chromatin status. Graphical Abstract

ABSTRACTRecent studies have found that the amount and proportion of meat flavor–related metabolites differ among different parts of muscle, which is partly attributed to their diverse myofiber type composition. However, the genetic regulatory factors behind these meat flavor–related metabolites remain unclear. Here, to systematically identify the differences in meat flavor–related metabolite of different parts of muscle and explore their genetic causes, metabolome and transcriptome were performed by using breast and drumstick muscle in Guangming‐2 chicken. Meat flavor–related metabolite‐gene networks were constructed, and phosphoglycerate kinase 2 (PGK2), which was found to be negatively correlated with flavor–related metabolites such as hypoxanthine, lysine, and glycerophospholipids, was selected for further identification. AlkB homolog 5 (ALKBH5)–mediated m6A demethylation enhances RNA stability of PGK2. Lentivirus‐mediated PGK2 overexpression and knockdown chicken models were constructed to study the function of PGK2. Gain‐ and loss‐of‐function analysis revealed that PGK2 promoted a switch from slow‐twitch to fast‐twitch fibers and suppressed the accumulation of hypoxanthine in vivo. Moreover, phosphoproteomic results demonstrated that PGK2 was involved in myofiber type transformation and meat flavor–related metabolite accumulation by modulating protein phosphorylation. Our findings deepen the understanding of the regulatory role of protein phosphorylation in meat flavor–related metabolite accumulation in broiler.

The quality of broiler meat affects consumers’ purchasing decisions. Numerous studies have shown that phosphorylation of proteins in muscle can affect muscle quality. Here, metabolomics and transcriptomics were used to systematically identify the genetic regulation of differences in meat flavor among different broiler. By constructing the meat flavor-related metabolite-gene networks, we identified that protein phosphatase magnesium/manganese-dependent 1J ( PPM1J ), which is known to regulate a range of biological processes by modulating reversible protein phosphorylation, was a differentially expressed gene with the highest connectivity to meat flavor-related metabolites. PPM1J facilitated the proportion of small myofiber (≤ 250 µm 2 ) and regulated the composition of glycerophospholipids. More importantly, phosphoproteomic and metabolome results found that PPM1J participates in the regulation of skeletal muscle development and glycerophospholipids composition by catalyzing protein dephosphorylation. Our study provides a basis for further understanding the molecular mechanism of skeletal muscle development and glycerophospholipids composition in broiler.

Consumers are increasingly concerned about the quality of meat products, and the smell and flavor of meat can affect the purchase of meat products by consumers. Here, to systematically identify the genetic regulation of meat flavor differences in different broiler breeds, a combination of metabolomics and transcriptomics. Protein phosphatase magnesium/manganese-dependent 1J (PPM1J) was one of the genes with the highest differential expression and connectivity among meat taste related metabolites. It is known to regulate a range of biological processes by modulating reversible protein phosphorylation. we found that facilitates proportion of small myofiber (≤ 250 μm2) and significantly up-regulate the content 1-stearoyl-2-docosahexaenoyl-sn-glycero-3-phosphocholine, glycine, and hypoxanthine was significantly up-regulated with PPM1J knockdown. Mechanistically, we found that most of the downregulated phosphopeptides were related to actin binding and muscle fibre development-related proteins in PPM1J knockdown muscle. Our study provides a basis for further understanding the molecular mechanism of meat flavor formation in broiler.

Carcass traits play important roles in the broiler industry and single nucleotide polymor-phism (SNP) can be efficient molecular markers for marker-assisted breeding of chicken carcass traits. Based on our previous RNA-seq data (accession num-ber GSE58755), cysteine rich with epidermal growth factor like domains 1 (CRELD1) and DnaJ heat shock protein family member C30 (DNAJC30) are differentially expressed in breast muscle between white recessive rock chicken (WRR) and Xinghua chicken (XH). In this study, we further characterize the potential function and SNP mutation of CRELD1 and DNAJC30 in chicken for the first time. Accord-ing to protein interaction network and enrichment analysis, CRELD1 and DNAJC30 may play some roles in chicken muscle development and fat deposition. In WRR and XH, the results of the rela-tive tissue expression pattern demonstrated that CRELD1 and DNAJC30 are not only differentially expressed in breast muscle but also leg muscle and abdominal fat. Therefore, we identified 5 SNP sites of CRELD1 and 7 SNP sites of DNAJC30 and geno-typed them in an F2 chicken population. There are 4 sites of CRELD1 and 3 sites of DNAJC30 are associ-ated with chicken carcass traits like breast muscle weight, body weight, dressed weight, leg weight per-centage, eviscerated weight with giblet percentage, intermuscular adipose width, shank length, and girth. These results suggest that the SNP sites of CRELD1 and DNAJC30 can be potential molecular markers to improve the chicken carcass traits and lay the foun-dation for marker-assisted selection.

Background Skeletal muscle is comprised of heterogeneous myofibers that differ in their physiological and metabolic parameters. Of these, slow-twitch (type I; oxidative) myofibers have more myoglobin, more mitochondria, and higher activity of oxidative metabolic enzymes compared to fast-twitch (type II; glycolytic) myofibers. Methods In our previous study, we found a novel LncRNA-TBP (for “LncRNA directly binds TBP transcription factor”) is specifically enriched in the soleus (which has a higher proportion of slow myofibers). The primary myoblast cells and animal model were used to assess the biological function of the LncRNA-TBP in vitro or in vivo . Meanwhile, we performed a RNA immunoprecipitation (RIP) and pull-down analysis to validate this interaction between LncRNA-TBP and TBP. Results Functional studies demonstrated that LncRNA-TBP inhibits myoblast proliferation but promotes myogenic differentiation in vitro . In vivo , LncRNA-TBP reduces fat deposition, activating slow-twitch muscle phenotype and inducing muscle hypertrophy. Mechanistically, LncRNA-TBP acts as a regulatory RNA that directly interacts with TBP protein to regulate the transcriptional activity of TBP-target genes (such as KLF4 , GPI , TNNI2 , and CDKN1A ). Conclusion Our findings present a novel model about the regulation of LncRNA-TBP , which can regulate the transcriptional activity of TBP-target genes by recruiting TBP protein, thus modulating myogenesis progression and inducing slow-twitch fibers.

Skeletal muscle is a regulator of the body's energy expenditure and metabolism. Abnormal regulation of skeletal muscle-specific genes leads to various muscle diseases. Long non-coding RNAs (lncRNAs) have been demonstrated to play important roles in muscle growth and muscle atrophy. To explore the potential function of muscle-associated lncRNA, we analyzed our previous RNA-sequencing data and selected the lncRNA (LncEDCH1) as the research object. In this study, we report that LncEDCH1 is specifically enriched in skeletal muscle, and its transcriptional activity is positively regulated by transcription factor SP1. LncEDCH1 regulates myoblast proliferation and differentiation in vitro. In vivo, LncEDCH1 reduces intramuscular fat deposition, activates slow-twitch muscle phenotype, and inhibits muscle atrophy. Mechanistically, LncEDCH1 binds to sarcoplasmic/ER calcium ATPase 2 (SERCA2) protein to enhance SERCA2 protein stability and increase SERCA2 activity. Meanwhile, LncEDCH1 improves mitochondrial efficiency possibly through a SERCA2-mediated activation of the AMPK pathway. Our findings provide a strategy for using LncEDCH1 as an effective regulator for the treatment of muscle atrophy and energy metabolism.

Long noncoding RNA (lncRNA) plays a crucial part in all kinds of life activities, especially in myogenesis. SMARCD3 (SWI/SNF-related, matrix-associated, actin-dependent regulator of chromatin, subfamily d, member 3) is a member of the SWI/SNF protein complex and was reported to be required for cell proliferation and myoblast differentiation. In this study, we identified a new lncRNA named SMARCD3-OT1 (SMARCD3overlappinglncRNA), which strongly regulated the development of myogenesis by improving the expression of SMARCD3X4 (SMARCD3transcripts4). We overexpressed and knockdown the expression of SMARCD3-OT1 and SMARCD3X4 to investigate their function on myoblast proliferation and differentiation. Cell experiments proved that SMARCD3-OT1 and SMARCD3X4 promoted myoblast proliferation through the CDKN1A pathway and improved differentiation of differentiated myoblasts through the MYOD pathway. Moreover, they upregulated the fast-twitch fiber-related genes and downregulated the slow-twitch fiber-related genes, which indicated that they facilitated the slow-twitch fiber to transform into the fast-twitch fiber. The animals' experiments supported the results above, demonstrating that SMARCD3-OT1 could induce muscle hypertrophy and fast-twitch fiber transformation. In conclusion, SMARCD3-OT1 can improve the expression of SMARCD3X4, thus inducing muscle hypertrophy. In addition, SMARCD3-OT1 can facilitate slow-twitch fibers to transform into fast-twitch fibers.

Poultry plays an important role in the meat consumer market and is significant to further understanding the potential mechanism of muscle development in the broiler. Bone marrow-derived mesenchymal stem cells (BM-MSCs) can provide critical insight into muscle development due to their multi-lineage differentiation potential. To our knowledge, chicken BM-MSCs demonstrate limited myogenic differentiation potential under the treatment with dexamethasone (DXMS) and hydrocortisone (HC). 5-azacytidine (5-Aza), a DNA demethylating agent, which has been widely used in the myogenic differentiation of BM-MSCs in other species. There is no previous report that applies 5-Aza to myogenic-induced differentiation of chicken BM-MSCs. In this study, we evaluated the myogenic determination and differentiation effect of BM-MSCs under different inductive agents. BM-MSCs showed better differentiation potential under the 5-Aza-treatment. Transcriptome sequence analysis identified 2402 differentially expressed DEGs including 28 muscle-related genes after 5-Aza-treatment. The DEGs were significantly enriched in Gene Ontology database terms, including in the cell plasma membrane, molecular binding, and cell cycle and differentiation. KEGG pathway analysis revealed that DEGs were enriched in myogenic differentiation-associated pathways containing the PI3K-Akt signaling pathway, the TGF-β signaling pathway, Arrhythmogenic right ventricular cardiomyopathy, dilated cardiomyopathy, and hypertrophic cardiomyopathy, which suggested that BM-MSCs differentiated into a muscle-like phenotype under 5-Aza-treatment. Although BM-MSCs have not formed myotubes in our study, it is worthy of further study. In summary, our study lays the foundation for constructing a myogenic determination and differentiation model in chicken BM-MSCs.

Meat production performance is one of the most important factors in determining the economic value of poultry. Myofiber is the basic unit of skeletal muscle, and its physical and chemical properties determine the meat quality of livestock and poultry to a certain extent. Peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PPARGC1A) as a transcriptional coactivator has been found to be widely involved in a series of biological processes. However, PPARGC1A is still poorly understood in chickens. In this manuscript, we reported that PPARGC1A was highly expressed in slow-twitch myofibers. PPARGC1A facilitated mitochondrial biogenesis and regulated skeletal muscle metabolism by mediating the flux of glycolysis and the TCA cycle. Gain- and loss-of-function analyses revealed that PPARGC1A promoted intramuscular fatty acid oxidation, drove the transformation of fast-twitch to slow-twitch myofibers, and increased chicken skeletal muscle mass. Mechanistically, the expression level of PPARGC1A is regulated by miR-193b-3p. Our findings help to understand the genetic regulation of skeletal muscle development and provide a molecular basis for further research on the antagonism of skeletal muscle development and fat deposition in chickens.

Skeletal muscle is the largest metabolic organ in the body, and its metabolic flexibility is essential for maintaining systemic energy homeostasis. Metabolic inflexibility in muscles is a dominant cause of various metabolic disorders, impeding muscle development. In our previous study, we found lncRNA ZFP36L2-AS (for “ ZFP36L2 -antisense transcript”) is specifically enriched in skeletal muscle. Here, we report that ZFP36L2-AS is upregulated during myogenic differentiation, and highly expressed in breast and leg muscle. In vitro, ZFP36L2-AS inhibits myoblast proliferation but promotes myoblast differentiation. In vivo, ZFP36L2-AS facilitates intramuscular fat deposition, as well as activates fast-twitch muscle phenotype and induces muscle atrophy. Mechanistically, ZFP36L2-AS interacts with acetyl-CoA carboxylase alpha (ACACA) and pyruvate carboxylase (PC) to induce ACACA dephosphorylation and damaged PC protein stability, thus modulating muscle metabolism. Meanwhile, ZFP36L2-AS can activate ACACA to reduce acetyl-CoA content, which enhances the inhibition of PC activity. Our findings present a novel model about the regulation of lncRNA on muscle metabolism.

As it is well known, muscle atrophy is a process in which protein degradation increases and protein synthesis decreases. This process is regulated by a variety of links. Among them, microRNAs play an essential role in this process, which has attracted widespread attention. In this paper, we find that miR-27b-3p and Cbl-b genes are significantly differentially expressed in the induced atrophy model. The dual-luciferase experiment and Western blot analysis confirmed that miR-27b-3p could regulate the expression of Cbl-b. In C2C12-differentiated myotubes, the overexpression of the Cbl-b gene showed that Cbl-b could upregulate the expression of MuRF-1 and Atrogin-1, which are related marker genes of muscle atrophy, at both the mRNA and protein levels, indicating that the Cbl-b gene can specifically affect muscle atrophy. The knockdown of the Cbl-b gene after C2C12-differentiated myotubes induced atrophy treatment can downregulate the expression of muscle-atrophy-related genes, indicating that manual intervention to downregulate the expression of Cbl-b has a certain alleviating effect on muscle atrophy. These data suggest that miR-27b-3p can regulate the expression of the Cbl-b gene and then exert a particular influence on muscle atrophy through the Cbl-b gene.

Background: Mesenchymal stem cells (MSCs) have the potential to multilineage differentiation, which can be used for a good model to provide critical insight of chicken muscle development. Differential adhesion method is one of the commonest methods to isolate MSCs based on the ability of plastic adhesion. 5-azacytidine (5-Aza), dexamethasone (DXMS), hydrocortisone (HC) and horse serum had been proved the potential to induce the myogenic differentiation of MSCs. However, the myogenic differentiation of MSCs is still poorly understood in chicken. In present study, we isolated chicken mesenchymal stem cells (cMSCs) from bone using 4-hour differential adhesion method and analyzed the myogenic effect of cMSCs treated with different method based on 5-Aza, DXMS, HC and horse serum. Results: cMSCs isolated by 4-hour differential adhesion method expressed MSCs special surface markers and presented normal growth characteristic. cMSCs showed great potential of myogenic differentiation by the treatment of 5-Aza and horse serum. RNA-sequence, GO and KGEE enrichment analysis revealed that this effect might be based on demethylation of 5-Aza and ECM-receptor interaction, focal adhesion, PI3K-Akt, p53, TGF-beta signaling pathways. Moreover, DXMS, HC and horse serum also presented potential of myogenic differentiation, but the effect was not as good as 5-Aza and horse serum method. Conclusions: cMSCs showed potential of myogenic differentiation by the treatment of 5-Aza and horse serum or DXMS, HC and horse serum.

Long non-coding RNAs (lncRNAs) are well-known to participate in a variety of important regulatory processes in myogenesis. In our previous RNA-seq study (accession number GSE58755), we found that lncRNA-FKBP1C was differentially expressed between White Recessive Rock (WRR) and Xinghua (XH) chicken. Here, we have further demonstrated that lncRNA-FKBP1C interacted directly with MYH1B by biotinylated RNA pull-down assay and RNA immunoprecipitation (RIP). Protein stability and degradation experiments identified that lncRNA-FKBP1C enhanced the protein stability of MYH1B. Overexpression of lncRNA-FKBP1C inhibited myoblasts proliferation, promoted myoblasts differentiation, and participated in the formation of skeletal muscle fibers. LncRNA-FKBP1C could downregulate the fast muscle genes and upregulate slow muscle genes. Conversely, its interference promoted cell proliferation, repressed cell differentiation, and drove the transformation of slow-twitch muscle fibers to fast-twitch muscle fibers. Similar results were observed after knockdown of the MYH1B gene, but the difference was that the MYH1B gene had no effects on fast muscle fibers. In short, these data demonstrate that lncRNA-FKBP1C could bound with MYH1B and enhance its protein stability, thus affecting proliferation, differentiation of myoblasts and conversion of skeletal muscle fiber types.

ALDH1A1 (aldehyde dehydrogenase 1A1) is a crucial protein in retinoids’ metabolism, and the lack of ALDH1A1 inhibits the fat deposition in mice. However, whether ALDH1A1 has a similar effect on chickens’ fat-depot is still unknown. In this study, we investigate the role of ALDH1A1 in chickens’ adipogenesis. The immortalized chicken preadipocyte 1 (ICP1) cell line and chicken primary preadipocytes isolated from abdominal fat were used to perform a series of experiments in vitro to elucidate the effects of ALDH1A1. In addition, lentivirus was used to verify the results of cell experiments in vivo. The data showed that overexpression of ALDH1A1 significantly weakened the proliferation of preadipocytes and suppressed the differentiation of preadipocytes through the PPARγ pathway, and the knockdown experiments had the opposite results. Moreover, chickens injected with overexpression lentivirus had higher abdominal fat percentage, a bigger size of lipid droplets, and higher triglyceride content in abdominal fat, and chickens injected with interfering lentivirus had the opposite situation. We proved that ALDH1A1 not only inhibited the proliferation and differentiation of chickens’ preadipocytes in vitro, but also inhibited the fat-depot of chickens in vivo, which was completely opposite the function of ALDH1A1 in mice, indicating that ALDH1A1 may have a different mechanism that is still unknown.

As the world population grows, muscle atrophy leading to muscle wasting could become a bigger risk. Long noncoding RNAs (lncRNAs) are known to play important roles in muscle growth and muscle atrophy. Meanwhile, it has recently come to light that many putative small open reading frames (sORFs) are hidden in lncRNAs; however, their translational capabilities and functions remain unclear. In this study, we uncovered 104 myogenic-associated lncRNAs translated, in at least a small peptide, by integrated transcriptome and proteomic analyses. Furthermore, an upstream ORF (uORF) regulatory network was constructed, and a novel muscle atrophy-associated lncRNA named SMUL (Smad ubiquitin regulatory factor 2 [SMURF2] upstream lncRNA) was identified. SMUL was highly expressed in skeletal muscle, and its expression level was down regulated during myoblast differentiation. SMUL promoted myoblast proliferation and suppressed differentiation in vitro. In vivo, SMUL induced skeletal muscle atrophy and promoted a switch from slow-twitch to fast-twitch fibers. In the meantime, translation of the SMUL sORF disrupted the stability of SMURF2 mRNA. Mechanistically, SMUL restrained SMURF2 production via nonsense-mediated mRNA decay (NMD), participating in the regulation of the transforming growth factor beta (TGF-beta)/SMAD pathway and further regulating myogenesis and muscle atrophy. Taken together, these results suggest that SMUL could be a novel therapeutic target for muscle atrophy.

FOXO3 belongs to the Forkhead O transcription factor family and it is an important gene in multiple biological processes, such as cell cycle control, cell proliferation, cell apoptosis, human longevity, and oxidative stress. Previous studies have shown that FOXO3 is associated with skeletal muscle growth and adipose development in mammals. However, the sequence of chicken FOXO3 is still incomplete and the cellular functions of FOXO3 in chickens are poorly understood. Thus, we obtained the full-length sequence of chicken FOXO3 by 5' rapid amplification of cDNA ends (5' RACE) and the phylogenetic tree showed that the chicken FOXO3 sequence was homologous with those in other species. Flow cytometry analysis and 5-ethynyl-2'-deoxyuridine assays showed that FOXO3 repressed cellular proliferation and induced apoptosis in a chicken hepatocellular carcinoma cell line (LMH). Mutations were screened in the second exon of FOXO3 and 13 synonymous single nucleotide polymorphisms were found in the test population. Further analysis showed that rs317670452 and rs15379317 were associated with many growth and carcass traits, such as the body weight at different ages and breast muscle weight. Our results indicate that chicken FOXO3 has similar cellular functions to those found in mammals and it is significantly associated with chicken growth.

SummaryThe proliferation, apoptosis, and differentiation of myoblasts are essential processes in skeletal muscle development. During this developmental process, microRNAs (miRNAs) play crucial roles. In our previous RNA-seq study (accession number GSE62971), we found that miR-16-5p was differentially expressed between fast and slow growth in chicken. In this study, we report that miR-16-5p could inhibit myoblast proliferation, promote myoblast apoptosis, and repress myoblast differentiation by directly binding to the 3′ UTR of SESN1, which is also differentially expressed. Overexpression of SESN1 significantly promoted the proliferation, inhibited apoptosis, and induced differentiation of myoblasts. Conversely, its loss of function hampered myoblast proliferation, facilitated myoblast apoptosis, and inhibited myoblast differentiation. Interestingly, we found SESN1 could regulate p53 by a feedback mechanism, thereby participating in the regulation of p53 signaling pathway, which suggests that this feedback is indispensable for myoblast proliferation and apoptosis. Altogether, these data demonstrated that miR-16-5p directly targets SESN1 to regulate the p53 signaling pathway, and therefore affecting myoblast proliferation and apoptosis. Additionally, SESN1 targets myogenic genes to control myoblast differentiation.