Effects Of FABP4, IGFBP3and SPP1Variations On Fat,Growth And Carcass Traits In Sheep

In this study, polymorphisms of the FABP4, IGFBP3and SPP1genes in1853sheepsourced from ten breeds （NZ Romney, Poll Dorset, Corriedal, Merino, Perendale, Dorper,Dorset Down, Suffolk, Coopworth, Ola and Ganjia） in New Zealand and China werestudied using PCR-SSCP technology. The aim was to indentify polymorphic sits,haplotypes and recombination event in candidate genes, as well as to assess the effect ofthe putative cSNP on structure and function of the protein. The diversity of geneticcharacter at FABP4locus among various population were also analysed. Given that theFABP4protein plays potential role in fat deposition and growth in the livestock, the fatand lean sheep from coopworth breed were used to analyse the association of FABP4variations with the fat/lean phenotype. To search for molecular markers, the association ofproduction traits for the fat trait （GR value）, growth traits （birth weight, tailing weight,weaning weight and pre-weaning growth rate） and carcass traits （hot weight, leg yield,loin yield, shoulder yield, total yield, proportion leg yield, proportion loin yield andproportion shoulder yield） were also analysed in Romney sheep. The main results show asfollows:1. Polymorphism of ovine FABP4, IGFBP3and SPP1A total of1469samples were typed in two polymorphic regions of FABP4gene. In theexon2-intron2region, eleven different SSCP patterns were observed （A₁A₁，B₁B₁，C₁C₁，D1D1，A1B1，B1C1，A1C1，A1D1，C1D1，B1D1and B1E1）, SSCP patterns （A1A1，B1B1，C1C1，D1D1） represent homozygous and SSCP patterns （A1B1，B1C1，A1C1， A₁D1，C1D1，B1D₁and B₁E₁） represent heterozygous. In the exon3-intron3region, ten elevendifferent SSCP patterns were observed （A₂A₂，B₂B₂，C₂C₂，D2D₂，A2B₂，A2C₂，B₂C₂，C₂D₂，B₂D₂and A₂D₂）, SSCP patterns （A₂A₂，B₂B₂，C₂C₂，D₂D₂） represent homozygousand SSCP patterns （A2B2，A2C2，B2C2，C2D2，B2D2and A2D2） represent heterozygous.In384samples, no different SSCP patterns were observed in IGFBP3and SPP1generespectively.In1469sheep, different nine sequence variants （A1，B1，C1，D1，E1，A2，B2，C2，D₂） were identified（Genbank accesson number: JX290313-JX290317; JX409931-JX409934）. Eight polymorphic sites including c.246+33C deletion, c.246+37A>Gsubstitution, c.246+46C>T substitution, c.246+47G>A substitution, c.317A>Gsubstitution, c.348+166T>C substitution, c.348+298T>C substitution, c.348+356C>Tsubstitution were identified. c.317A>G substitution results in the amino acid change:Lys（AAG）106Arg（AGG）.2. The analysis of the linkage disequilibrium, haplotypes and recombination ofFABP4geneThe linkage disequilibrium of SNPs spanning both regions was analysed in483sheep.Theresults indicated that the relationship of SNPs in two regions was not disequilibriumcompletely （D′=0.548,r2=0.119）. Fourteen different haplotypes were identified includingA₁A₂, A₁B₂, A₁C₂, A₁D₂, B₁A₂, B₁B₂, B₁C₂, C₁A₂, C₁B₂, C₁C₂, D₁A₂, D₁B₂, D₁C₂,E₁A₂.Two chi-like sequences in both polymorphic regions were detected respectivelyincluding5′GTTGGTGA3′and5′GCTGGTGA3′, as well as several reverse repeatsequences comprising of eight bases. This results indicated that above specific sequencesmay facilitate recombination in the ovine FABP4gene. Different haplotypes wereobserved in the progeny from five sires, suggesting that recombination occurred in theFABP4gene. To ensure the truth, DQA₂, ADRB3and PRNP gene located at differentchromosomes were used to confirm the genetic relationship between the sire and theirprogeny.3. The reconstruction of three-dimensional model of the FABP4protein and theeffect of Lys106Arg change on the structure and founctionThe homology modeling method was used to construct the predicted structuresuccessfully, which was composed of two α helixes and ten β chains. The score of modelparameter was0.73（0-1）, indicating a high quality for the predicted model. It is predictedthat the Lys106Arg change will not affect the structure and function of the FABP4proteinobviously, but it has the possibility for c.317A>G substitution linking with other variationsto the FABP4protein indirectly.4. The difference of genetic characters at FABP4locus among variouspopulationsIn five hundred and seventy five sheep from ten populations, the allele A₁, B₁and C₁were common. The allele D₁was observed in eight populations except the Poll Dorset andGanjia, the allele E₁was only observed in the Poll Dorset and Corriedal. The allele A₁with the frequency of38.4%, and genotype A₁B₁was the most common. The allele A₂, B₂and C₂were observed in ten populations and the allele B₂with the frequency of48.3%and genotype A₂B₂was the most common, but the allele D₂was not observed inCorriedale, Merino, Ola and Ganjia.2test indicated that variations were deviated to the Hardy-Weinberg equilibrium, but variations were in the Hardy-Weinberg equilibrium inTibetan sheep （P>0.05）.There was not a difference for genotype between the Romney andPerendale population （P>0.05）, but there was a difference for genotype between theCorriedale and Merino （P<0.05）.At two polymorphic locus of FABP4, the highheterozygosity and PIC （PIC>0.5） were observed in NZ populations, but in two bitetpopulations the high homozygosity and moderate PIC （0.25<PIC <0.5） were observed.Cluster analysis in ten populations showed that Ola and Ganjia, Romney and Perendle, aswell as Corriedale and Romney were clustered respectively.5. The difference of FABP4variations in fat and lean line from Coopworth breedin NZThere was a difference （P<0.001） for sequence variants in both regions between twolines. In the eoxn2-intron2, only the allele B₁, C₁and D₁were observed and C₁was themost common （a frequency of89%） in the lean line, whereas in the fat line, five sequencevariants were observed and sequence A₁with a frequency of51%was the most common.In the exon3-region3, only the sequence A₂, B₂and C₂were observed and C₂was themost common （a frequency of59%） in the lean line. However, the C₂was not observedand B₂with a frequency of83%was the most common in the fat line..8. Effects of FABP4variations on growth trait in Romney sheepThree different models were used to assess the association of sequence and haplotypepresent/absent, copy number, genotype with growth traits （birth weight, tailing weight,weaning weight and pre-weaning growth rate）.The presence of haplotype A₁-A₂and C₁-C₂were trending to be associated with thedecreased birth weight （P<0.1）, but haplotype C₁-A₂were trending to be associated withthe increased birth weight （P<0.1）. The presence of haplotype C₁-A₂was trending to beassociated with the decreased tailing weight （P<0.1）. The presence of haplotype C₁-C₂was trending to be associated with the decreased weaning weight （P<0.1）. Genotype had asignificant effect on tailing weight （P<0.05）, the lambs with genotypep A₂C₂have thelowest weaning weight （28.849±0.452Kg）, tailing weight（12.831±0.273Kg） andpre-weaning growth rate （265.668±4.779g/d）.9. Effects of FABP4variations on fat and carcass traits in Romney sheepThree different models were used to assess the association of sequence and haplotypepresent/absent, copy number, genotype with growth traits （hot weight, leg yield, loin yield,shoulder yield, total yield, proportion leg yield, proportion loin yield and proportion shoulder yield）.The presence of A₁was trending to be associated with the increased GR value, legyield, loin yield and total yield （P<0.05/0.1）. The presence of A₂was trending to beassociatied with the increased hot weight （P<0.1）.The presence of A₁-A₂was trending tobe associated with the decreases hot weight （P<0.1）. The presence of B₁was trending tobe associated with an increase in proportion leg yield（P<0.1）. The presence of D₁wastrending to be associated with the increases shoulder yield （P<0.1）. The presence of A₁-A₂was associated with a decrease in leg yield, hot weight, loin yield, total yield, proportionloin yield and an increase in proportion shoulder yield （P<0.05/0.1）. Genotype had aneffect on shoulder yield （P<0.1）, lambs possessing the genotype A₁B₁have the lowestshoulder yield （16.296±0.133Kg）.