Regulation Of Fatty Acids Metabolism In Inner Mongolia Cashmere Goats

Inner Mongolia cashmere goats are known for their fine cashmere fiber,white color,soft feel and bright luster,which objectively makes people ignore their excellent characteristics as meat livestock.Inner Mongolia Cashmere Goat is the main livestock species,which is regarded as the survival category by local herdsmen.However,up to now,there is no systematic report on the study of Inner Mongolia cashmere goat meat,which restricts the full development and effective use of this breed,and directly affects the overall benefit of cashmere goat breeding.Based on the analysis of physicochemical and nutritional indicators of Inner Mongolia cashmere goat meat(2.5 years old,25-26 kg,wether),using transcription and proteomics methods,we found that there are differences in fatty acid formation genes,microRNAs and proteins in the meat of different parts of cashmere goat.Then we constructed a regulatory network of fatty acid metabolism by using multi-omics combined analysis.These detailed results are as follows:1.Study on the physicochemical properties of cashmere goat meat showed that the intramuscular fat content,brightness(L *)and redness(a *)of LD were higher than that of BF,while protein content,yellowness(b *),shear force and water loss rate in BF were higher than that in LD.Nutritional analysis results of three Inner Mongolia cashmere goats were showed that the intramuscular fat content in LD was significantly higher than that in BF(P< 0.01);A total of 34 fatty acids in its muscles;While the content of unsaturated fatty acids in BF was significantly higher than that in LD(P < 0.05).In conclusion,The selected samples of LD and BF of three goats are representative for studying fatty acid metabolism.2.Compared LD and BF at the mRNA level,we found that: there were 18 differential genes(DEGs)involved in fatty acid metabolism,including ACSL1,LDHB,ACADS and so on;there were 8 signaling pathways related to fatty acid metabolism,including fat digestion and absorption,fatty acid biosynthesis,fatty acid metabolism and PPAR signaling pathways etc.In addition,through gene-pathway network analysis of these genes and pathways,we found that ACSL1,CD36 and PPAR signaling pathway are at the center of the regulatory network and play an important role in the regulation of fatty acid metabolism.The expression of CD36,TECRL,ACADVL and DGAT2 were validated using RT-q PCR,The expression of these selected transcripts was significantly higher in the BF than in the LD(P < 0.01),and the expression patterns were highly consistent with those obtained by theRNA-Seq method.3.At the miRNA level,we are the first one who reported 426 known miRNAs and 30 new miRNAs in meat of Inner Mongolia cashmere goat.KEGG analysis of miRNA target genes in these goats were performed,and it was revealed that the gene was mainly involved in the biosynthesis of fatty acids,PPAR,the degradation and absorption of fatty acids and other signaling pathways.TheRNA-Seq results for chi-let-7c-5p,chi-mi R-365-3p,chi-mi R-191-5p,were validated using.The expression of these selected miRNAs was significantly higher in the LD than in the BF(P < 0.01),and the expression patterns were highly consistent with those obtained by theRNA-Seq method.4.At the protein level,the identification and analysis of BF and LD found that a total of 1209 proteins were identified in Inner Mongolia cashmere goat meat.There were 110 significantly differently expressed proteins in the groups of BF and LD,where the expression of 81 proteins in the BF group was up-regulated,while 29 proteins in LD were up-regulated.The results of KEGG analysis showed that 110 differential proteins were covered in 34 KEGG pathways.there were 3 pathways related to fatty acid metabolism in all pathways,including fatty acid metabolism,fatty acid degradation and fatty acid extension.PPI network analysis showed that HADHA,ACAA2 and ACAT1 might play an important role in regulating IMF anabolism and catabolism in Inner Mongolian cashmere goat.Immunoprecipitation results showed that there was an interaction network around HADHA protein,and the proteins that directly interacted with HADHA were IDH2,GOT2 and ALDOA.Based on this,we speculate that HADHA in Inner Mongolia cashmere goat muscle mainly regulates long-chain fatty acid metabolism through the proteins of IDH2 and ALDOA.Western blot was used to detect the expression levels of HADHA,FABP3 and MYL2,The band intensities of these three proteins were significantly higher in the biceps femoris than in the longissimus dorsi muscle(P < 0.05).5.The joint analysis of mRNA-miRNA-protein showed that in the process of ACSL1 gene regulating the expression of AOA452FP41 protein,1-9 miRNAs acted on ACSL1 alone or together to degrade or inhibit the gene,thereby regulating the expression of AOA452FP41 protein and ultimately affecting metabolism of long-chain fatty acids.Through the fatty acid degradation signaling pathway,Chi-mi R-361-3P＿R +1 acts on ACADS,degrades or inhibits ACADS,reduces the expression of ACADS and ACAT1 proteins,thus affecting the metabolism of short-chain fatty acids,and ultimately leads to a low content of unsaturated fatty acids in the LD.The expressions of key genes(ACSL1 and ACADS),key miRNAs(Chimi R-16b-5p,Chi-mi R-10b-5p and Chi-mi R-16a-5p)and key protein(ACSL1)obtained by multi-omics analysis were verified by fluorescence quantification,Western blot and protein fishing techniques,and it was found that the contents of these parameters in LD and BF muscle were significantly different(P < 0.05).This study discovered key genes that have important contributions to fatty acid metabolism,such as ACSL1,HADHA and ACADS,and constructed a fatty acid regulatory network of ACSL1 at the mRNA,miRNA and protein levels,and laid the foundation for elucidating the mechanism of fatty acid metabolism in Inner Mongolia cashmere goat.