Effects Of THRSP Gene Polymorphism On Fat Deposition And Its Molecular Mechanisms In Pigs

China is the largest pork consumption country in the world.People pay more and more attention to the quality and flavor of pork.The research shows that the increase of fat content will contribute to improve the quality and flavor of pork.As an important transcription factor,THRSP gene regulates the secretion of enzymes related and cytokines in lipid metabolism,and affects the fat deposition in pigs.Therefore,THRSP gene is the key factor to regulate the expression of genes related to fat synthesis and decomposition.In this study,we selected 1078 pigs in 8 pig populations,including 3 local pig breeds,3introduced pig breeds and 2 hybrid populations to detect the polymorphism in 5’ proximal regulatory region(PRR)of THRSP gene and backfat thickness.The relationships between the PRR haplotypes and backfat thickness and between the PRR polymorphisms and transcription factor binding sites were analyzed.The regulatory effect of the PRR promoter-types on the transcription level of THRSP gene was studied by dual luciferase reporter assay.Some candidate genes related to the fat deposition were screened based on the PRR promotertypes using RNA-seq,and their biological functions were studied by GO enrichment and KEGG pathway analysis.The main research work and achievements are summarized as follows:1)After sequencing,it was found that there were 5 SNP sites in PRR fragment sequence,A-95 T,G-98 A,C-376 A,T-400 C and C-459 T upstream from the CDS start of the THRSP gene.The distribution of 5 SNP sites was specific among the ability of fat deposition in different pig breeds.And five combined genotypes were detected: CCTCGAGAAT(frequency 0.2078),CCTCGAGGAA(frequency 0.1364),CCTTGGGGAA(frequency0.2403),CTTCGAGAAT(frequency0.1558)and CTTCGAGGAA(frequency 0.2597),which showed that CAA was dominant haplotype in local pig breeds and TGG in introduced pig breeds.By comparative analysis,the PRR homology of THRSP gene was 100% between pig and cattle,but the homology between pig and sheep or chickens was lower.2)The expression level of THRSP gene was significantly higher in local pig breeds(72.717)than that in introduced pig breeds(8.718).The ability of fat deposition in different pig breeds was significantly correlated with the expression level of THRSP gene and PRR haplotypes.3)In dual luciferase reporter assay in vitro,the expression of luciferase gene with different PRR promoter showed an increasing trend from TTCCAAGGAA,CCCCCCAAGGAA,CCTTAAGGAA,CCTTGGAATT to CTTGGGAA of PRR promoter-types,and there weresignificant differences among them(p < 0.01).And the expression level of THRSP gene in pig’s liver tissues with TTCCAAGGAA,CCCCAAGGAA and CTTGGGAA genotypes showed similar differences(P < 0.01),which suggested that the PRR polymorphism of THRSP gene plays an important role in regulating its expression efficiency.4)It showed by RNA-seq analysis that 130 genes differentilly expressed in pig’s liver tissues between CCTTGGGGAA and CCCCAAAATT,42 genes differentilly expressed between CCTTGGGGAA and CCTCGAGAAT,and 56 genes differentilly expressed between CCCCAAAATT and CCTCGAGAAT.And FDPS,CYP7A1,FADS2 and FADS1,which were associated with fat metabolism,were regulated through PPAR pathway.