Identification Of Genes Related To Muscle Growth And Lipid Deposition From Transcriptomic And Proteomic Profiles Of Pig

Muscle growth and meat quality are complex quantitative traits and the main economic traits in pig production.Chinese indigenous pig breeds,the Diannan Small Ear pig(DSP)and the Tibetan pig(TP),have lower growth rate,more fat deposition,and better meat quality than the introduced pig breeds,such as the Landrace(LL)and Yorkshire(YY),which are lean-type pig breeds characterized by a fast growth rate and high lean meat content.RNA-seq,miRNA-seq,iTRAQ and PRM were applied to analyze the transcriptomic and proteomic profiles from the tissues of longissimus dorsi muscle of 4 breeds to identify the differentially expressed genes and miRNAs related to muscle growth and lipid deposition.We detected that 18,208 genes were positively expressed and 14,633 of them were co-expressed in the muscle tissues of the four samples.When the filter criteria of |log2FC|?1 and P<0.05 was applied,315 DEGs were acquired from the DSP-TP and the LL-YY combined.Of these DEGs,140 showed upregulated and 175 downregulated expression in the DSP-TP group.Based on DAVID functional annotation,we identified 85 DEGs related to growth traits that were mainly involved in muscle tissue development,muscle system process,regulation of cell development,and growth factor binding,and 27 DEGs related to lipid deposition that were mainly involved in lipid metabolic process and fatty acid biosynthetic process.With miRNA-seq,we obtained 320 positively expressed miRNAs,including 271 known pig miRNAs and 49 novel miRNAs.In those 271 known miRNAs,20 were higher and 10 lower expressed in DSP-TP than in LL-YY(p<0.01 and |log2FC|?1).The target genes of the 30 miRNAs were mainly participated in MAPK,GnRH,insulin and Calcium signaling pathway and others were involved in cell development,growth and proliferation,etc.Combining the DEGs and the differentially expressed(DE)miRNAs,we drafted a network of 46 genes and 18 miRNAs for regulating muscle growth and a network of 15 genes and 16 miRNAs for regulating lipid deposition.We identified that CAV2,MYOZ2,FRZB,miR-29b,miR-122,miR-145-5p and miR-let-7c,etc.were key genes or miRNAs regulating muscle growth,and FASN,SCD,ADORA1,miR-4332,miR-182,miR-92b-3p,miR-let-7a and miR-let-7e,etc.were key genes or miRNAs regulating lipid deposition.In total,4,815 peptides corresponding to 969 proteins were detected through iTRAQ analysis.Comparison of expression patterns between DSP-TP and LL-YY revealed 288 differentially expressed proteins(DEPs),of which 169 were up-regulated and 119 were down-regulated(FC>1.3or FC<0.77).Functional annotation suggested that 28 DEPs were related to muscle growth and 15 to lipid deposition.Combining the DEPs and the DEmiRNAs,we drafted a network of 17 miRNAs and their 54 target genes,of which 2 miRNAs and 2 proteins were related to muscle growth and 3 miRNAs and 2 proteins were related to lipid deposition.Comprehensive analysis indicated that differences in muscle growth and muscle fibre between DSP-TP and LL-YY groups were regulated by ALDOC,EN03,PGK1,PGK2,TNNT1,SETX,SLC9A3R1,miR-152 and miR-339,etc.whereas differences in lipid deposition ability were regulated by LPL,APOA1,APOC3,ACADM,FABP3,GPD2,UQCRC1,miR-122 and miR-29b,etc.Twelve DEPs were analysed using parallel reaction monitoring(PRM)to confirm the reliability and accuracy of the iTRAQ analysis.We selected pig FRZB as a candidate gene for growth traits and identified three polymorphic sites,an insertion(A-532B)and two SNPs(G636A and C650T)in its 5'-UTR.The genotype distributions of G636A and C650T were significantly different among indigenous(DSP and TP)and introduced(LL and YY)breeds.The G636A and C650T sites of FRZB gene had significant effects on growth of new-line Huai pig,and the favorable allele were G and C and the favorable genotypes were GG and CC of the locus of G636A and C650T respectively in new-line Huai pig.In semi-quantitative PCR expression analysis,expression of FRZB mRNA was abundant in tissues of hypophysis,longissimus dorsi muscle,and adipose tissues,and low in the heart,hypothalamus,and brain.Quantitative determination of mRNA level and protein expression analysis were corresponding.The results demonstrated that FRZB gene expression in longissimus dorsi muscle and liver tissue was significantly higher in DSP and TP than in the YY breed(p<0.05);however,in back fat tissue,the expression was significantly higher in the DSP than in TP or YY breeds(p<0.05).These results indicate that FRZB expression has a negative association with muscle growth and a positive association with fat deposition.In conclusion,FRZB may be a major candidate gene for growth traits in pigs.We used the RNA-seq,miRNA-seq and iTRAQ to identify the differentially expressed genes and miRNAs related to muscle growth and lipid deposition of pig.Our findings provide new insights into key proteins involved in muscle growth and lipid deposition in the pig.The results provided the valuable data for the use of modern biotechnology to cultivate pig breed with high growth rate and lipid deposition.