| Meat production performance is an important economic index in meat purpose goat breeding,which directly affects the economic benefit of meat purpose goat breeding.Meat yield and meat quality are main genetic traits reflecting meat production performance.To investigate the molecular mechanism regulating caprine meat production traits,Longissimus dorsi muscle from Ziwuling black goats and Liaoning cashmere goats with significant differences in meat yield and meat quality,were selected to detect the difference of muscle fibers and collagen fibers.Subsequently,non-coding RNAs related to muscle growth and development,fat deposition and meat tenderness were screened by RNA sequencing.Furthermore,the molecular mechanism of non-coding RNAs regulating muscle growth and development of goats was investigated by cell experiments in vitro.The main results of this study are as follows:1.There are significant differences in the histological characteristics of muscle between Liaoning cashmere goats and Ziwuling black goats.The muscle fiber diameter,crosssectional area and collagen fiber content in Longissimus dorsi muscle of Liaoning cashmere goats were higher than those of Ziwuling black goats,but the density of muscle fibers was lower than those of Ziwuling black goats(P < 0.05).2.Small RNA sequencing results showed that a total of 410 known goat miRNAs and159 differentially expressed miRNAs were identified in Longissimus dorsi muscle tissues of Liaoning cashmere goats and Ziwuling black goats.Of these differentially expressed miRNAs,24 miRNAs were up-regulated in Longissimus dorsi muscle of Liaoning cashmere goats compared to Ziwuling black goats,while 135 miRNAs were down-regulated.Functional annotation results showed that the target genes of up-regulated miRNAs were mainly enriched in Ras and Rap1 signaling pathway related to muscle development and fat deposition(P <0.05).For down-regulated miRNAs,their target genes were mainly enriched in Hippo signaling pathway related to muscle fiber hypertrophy(P < 0.05).miRNAs-m RNAs regulatory network showed that differentially expressed miRNAs target genes SMPX,NOTCH1,PTEN and KLF10 related to muscle fiber hypertrophy and fat deposition.3.RNA-seq analysis results showed that a total of 2302 lncRNAs and 136 differentially expressed lncRNAs were identified in Longissimus dorsi muscle of Liaoning cashmere goats and Ziwuling black goats.Of these differentially expressed lncRNAs,62 lncRNAs were upregulated in Longissimus dorsi muscle of Liaoning cashmere goats when comparing to Ziwuling black goats,while 74 lncRNAs were down-regulated.GO and KEGG analysis showed that the target genes of differentially expressed lncRNAs were significantly enriched in muscle contraction,muscle cell differentiation,connective tissue development and p53 signaling pathway(P < 0.05).The lncRNAs-m RNAs regulatory network showed that differentially expressed lncRNAs trans-regulate the expression of genes such as SMPX,MYL6 B,RYR3 and LOXL2 to further regulate muscle fiber hypertrophy,adipose deposition and meat tenderness.Ce RNA analysis results showed that lncRNA(XR＿001917125.1)functions as the molecular sponges of miR-34-3p and miR-460-5p to regulate the expression of NCAM2 and further regulate muscle growth and development.While XR＿001918832.1can functions as the molecular sponges of miR-200 b and miR-200 c to regulate the development of muscle cells and adipocytes.4.RNA-seq analysis results showed that a total of 10,875 circRNAs and 214 differentially expressed circRNAs were identified in Longissimus dorsi muscle of Liaoning cashmere goats and Ziwuling black goats.Of these differentially expressed circRNAs,85 circRNAs were up-regulated in Longissimus dorsi muscle of Liaoning cashmere goats when comparing to Ziwuling black goats,while 129 circRNAs were down-regulated.GO and KEGG enrichment analysis results showed that the parent genes of differentially expressed circRNAs were significantly enriched in connective development process related to meat tenderness,as well as Rap1,c GMP-PKG,c AMP and Ras signaling pathway related to the development of muscle cells and adipocytes(P < 0.05).Ce RNA regulatory network analysis showed that circ＿008879,circ＿004971 and circ＿006117 can function as the molecular sponges of miRNAs to regulate skeletal muscle development and fat deposition.5.miR-381 and miR-200 b affect the proliferation and differentiation of caprine skeletal muscle satellite cells(SMSCs)by targeting the genes related to Notch signaling pathway.Over-expression of miR-381 and miR-200 b inhibited the viability and proliferation of caprine SMSCs,but promoted the formation of myotube and myogenic differentiation.On the contrary,silence of miR-381 and miR-200 b promoted the viability and proliferation of SMSCs,and inhibited the formation of myotube and myogenic differentiation.Dual luciferase reporter assay confirmed that miR-381 directly target JAG2 and PTEN related to Notch signaling pathway and negatively regulated their expression in SMSCs.Similarly,miR-200 b directly target both NOTCH1,a crucial molecular of Notch signaling pathway,and CDK2,a cell proliferation regulator,and also negatively regulated their expression.6.circ CTAGE5 has crucial regulatory effect on the proliferation and differentiation of caprine SMSCs.Silence of circ CTAGE5 inhibited viability and proliferation of SMSCs,increased the expression of myogenic differentiation marker genes My HC,Myo G and MEF2 C,decreased the expression of SMAD4 and TGFB2,and promoted the formation of myotubes.In this study,we obtained the expression profiles of miRNAs,lncRNAs and circRNAs in Longissimus dorsi muscle tissues of Liaoning cashmere goats and Ziwuling black goats,and initially explored the functions of non-coding RNAs in muscle development,fat deposition and meat tenderness.Furthermore,it was found that miR-381,miR-200 b and circ CTAGE5 regulated the proliferation and differentiation of SMSCs.The results provide the basic data for revealing molecular mechanism of caprine muscle development and formulating molecular markers of caprine meat production traits. |
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