Mechanism Of RXRA Targeting CD36 To Regulate Lipid Metabolism And Residual Feed Intake Of Meat Duck

The shortage of feed and the soaring price of feed have brought a huge blow to the waterfowl breeding industry,which is in a harsh environment of high cost and low return.The cost of ducks feed accounts for 60%-70% of the total cost,and increasing feed utilization will also indirectly reduce the breeding cost of China’s poultry breeding industry.Improving feed efficiency has become a new breeding goal for the duck industry.Currently,domestic and foreign scholars are at the stage of histological analysis and bioinformatics prediction regarding the feed utilization rate of meat ducks,but have not conducted functional verification and mechanism research on candidate genes.Therefore,this study aims to explore the effect of candidate genes on residual feed intake in meat ducks and their mechanisms.500 Qiangying ducks were selected as experimental materials,and the growth performance and feed efficiency of 21～ 42-day-old ducks were recorded and calculated.The expression of RXRA in breast muscles of 42-day-old meat ducks was detected and sequenced into 25 individuals with high RXRA expression(HRXRA),medium RXRA expression(MRXRA),and low RXRA expression(LRXRA).The growth performance,feed efficiency,slaughter performance,intramuscular fat and serum lipid content of meat ducks were measured.The results showed that the average daily feed intake(ADFI),feed conversion rate(FCR),and residual feed intake(RFI)in the LRXRA group were significantly lower than those in the MRXRA and HRXRA groups(P<0.05),but had no significant impact on growth performance and slaughter performance.The serum triglyceride(TG)level in the LRXRA group was significantly higher than that in the HRXRA group(P<0.05).Primary skeletal muscle cells(CS2)were isolated and cultured from 13-day-old duck embryos.After overexpressing and knocking down the expression of RXRA in CS2,RT-PCR and western blotting were used to detect the expression levels of genes and proteins related to the PPAR signaling pathway.The content of intracellular TG,cholesterol(CHOL),and non-esterified free fatty acids(NEFA)was measured using Elisa’s kit.The fat content was visually analyzed using oil red O staining.The relationship between RXRA and candidate target genes was validated using dual fluorescein reporter genes and chromatin immunoprecipitation(CHIP).The difference in intracellular fatty acid content was analyzed by meteorological mass spectrometry.RNA-seq is used to explore differential genes.The results showed that overexpression of RXRA decreased the content of TG,CHOL,NEFA,and lipid droplets in CS2,and upregulated the m RNA expression of CD36,ACSL1,and PPARG genes,as well as the protein expression of CD36 and PPARG.Knocking down RXRA increased the content of TG,CHOL,NEFA,and lipid droplets in CS2,and downregulated the m RNA and protein expressions of CD36,ACLS1,ELOVL6,and PPARG.The double luciferase reporter gene activity of the wild type CD36 promoter was higher than that of the mutant type due to overexpression of the RXRA gene.RXRA binds to-860/-852 nt,-688/-680 nt,and-165/-157 nt in the promoter region of CD36.In addition,overexpression of CD36 in CS2 cells can inhibit the content of TG,CHOL,NEFA,lipid droplets,and saturated fatty acids(SFA).Knocking down the expression of CD36 can increase the content of TG,CHOL,NEFA,lipid droplets,SFA,and monounsaturated fatty acids(MUFA).CD36 mainly accelerates the outflow of fatty acids from CS2 through phosphonate and hypo-phosphonate metabolism,fatty acid metabolism,and substance metabolism.The whole blood genome of 243 Qiangying ducks was extracted,and the relationship between single nucleotide polymorphisms(SNPs)of RXRA and CD36 genes and growth and feed traits of meat ducks was analyzed by sequencing and enzyme digestion.Results: Four SNPs were detected in the coding region of the RXRA gene,and eight SNPs were found in the non-coding region.The amino acids of g.5,952,703 C>T and g.5,952,667 C>T have changed.g.5,952,667 is related to ADFI,RFI,and FCR.One SNP was detected in CD36noncoding(g.11,233,114 T>G).There was no significant correlation between this locus and growth performance and feed efficiency in meat ducksThe expression of CD36 in the breast muscle of 42-day-old meat ducks was detected and sequenced,and 25 individuals with high CD36 expression(HCD36),medium CD36expression(MCD36),and low CD36 expression(LCD36)were divided.The expression of RXRA gene in breast muscle,growth performance,feed efficiency,slaughter performance,intramuscular fat,and serum lipid content of meat ducks were measured.The results showed that the expression of RXRA gene,ADFI,FCR,and RFI in HCD36 group were significantly higher than those in MCD36 and LCD36 groups(P < 0.05).Compared with HCD36 and MCD36 groups,the content of abdominal fat and serum low density lipoprotein(LDL)in the LCD36 group significantly decreased,while the content of intramuscular fat significantly increased(P < 0.05).The ADFI,RFI,and FCR of meat ducks are influenced by the RXRA gene in the pectoral muscle,and the RXRA gene expression level is significantly correlated with the levels of intramuscular fat and serum lipids.The ADFI,RFI,and FCR of meat ducks are influenced by the RXRA gene in the pectoral muscles,and the expression level of the RXRA gene is significantly correlated with the levels of intramuscular fat and serum lipids.In duck skeletal muscle satellite cells,RXRA promotes the expression of CD36 and inhibits intracellular accumulation.CD36 accelerates fatty acid metabolism through pathways such as fatty acid metabolism,fatty acid elongation,and extracellular flow.The SNPs of RXRA are associated with ADFI,RFI,and FCR in meat ducks.RXRA affects feed efficiency by promoting the expression of CD36 and inhibiting fat accumulation in breast muscles of meat ducks.Our research aims to clarify the impact of candidate genes on feed efficiency and their mechanisms in meat ducks,and provide a theoretical basis for cultivating meat ducks with high feed efficiency.