Identification, Tissue Expression And Polymorphism Detection Of Goat PPARγ And FABP Gene Family

Using Nanjiang yellow goat and Tibet goat as experimental materials, we cloned and sequenced FABP1, FABP2, FABP3, FABP4, PPARyl and PPARy2genes. Semi-quantitative RT-PCR was employed to investigate their expression profiles in13tissues incuding subcutaneous fat, longissimus muscle, colorectal, small intestine, ovary, uterus, brain, stomach, kidney, lung, spleen, liver and heart of Tibet goat. Real-time RT-PCR method was used to quantify the mRNA expression of FABP3, FABP4, PPARyl, PPARy2genes in longissimus muscles of Nanjiang yellow goat at different developmental stages, and then the correlative analysis of mRNA level and IMF content was conducted. Additionally, PCR-RFLP method was used to detect the polymorphisms of FABP4gene in Nanjiang yellow goats. The results were showed as follows.(1) The CDS of FABP1-4gene and two isoforms of PPARy were384bp,399bp,402bp,399bp,1428bp and1518bp, which encoded127,132,133,132,475and505amino acid residues respectively, the accession numbers of GenBank were JQ031286, JQ031285, JQ031287, JQ031288, JQ266369and JQ387581respectively. The similarity of goat FABP1-4nucleotide sequences with those FABP1-4genes in sheep, cow, pig, human and mice was more than79%, and the general similarity of amino acid sequences between the same species also showed the same rates. The highest similarity (95%) presented between sheep and cow. The PPARyl genes from goat, sheep, cow, swine, human and mice were appeared to be highly similarity (88%). And the PPARy2gene showed the same similarity between the same species. The amino acid of goat PPARγ1and PPARγ2had more than97%similarity with those in sheep, cow, swine, human and mice.The main modification of carpine FABP1-4and PPARyl-2was the phosphorylation and their secondary structural elements were alpha-helix, random coil and extension fragment. The carpine FABP1-4had a common domain. The two isoforms of PPARy protein contained two highly conserved domains named DBD and LBD, the former showed100%consistency and the latter showed98%consistency among species.(2) The results of semi-quantitative PCR indicated different expression levels of FABP1-4, PPARyl and PPARy2in13tissues of Tibetan goat. FABP1gene had the high expression in colorectal, kidney and liver and moderate expression in small intestine and no expression in other tissues. FABP2gene was only expressed in small intestine and liver,with the highest expressed in small intestine. FABP3gene showed the high expression in longissimus muscle, colorectal, small intestine, uterus, brain, kidney, spleen and heart, while moderate expression in other tissues. The FABP4gene was highly expressed in subcutaneous fat, stomach, lung and spleen, but not expressed in heart. The PPARyl gene had the high expression in spleen and ovary and no expression in heart. In heart, lung and subcutaneous fat the PPARy2gene showed the high expression. And in small intestine the PPARy2gene had no expression.(3) The expression level of FABP3, FABP4, PPARγ1and PPARγ2genes in longissimus muscles of Nanjiang yellow goat had been detected at each developmental stage. The data showed the down-up-down trend in FABP3gene, and the reached the bottom at120th day (P<0.05). The mRNA expression level of FABP4was up-regulated with aging and peaked at120th day which was significantly higher than that at3th day and30th day (P＜0.01). Generally, PPARγ2and PPARγ1genes showed different patterners at different stages. The PPARγ1gene expressed with up-down manner, and peaked at60th day, while the PPARγ2mRNA profile showed the same trend as the FABP4gene, and the expression difference among120th day,3th day and30th day points were significant.The mRNA expression level of FABP3gene showed a positive correlation with IMF content (P<0.05). However, the FABP4gene and PPARy2gene expressions showed a significant negative correlation with IMF content (P＜0.05).(4) There were two mutations found in the intron3(T143C) and exon4(T546C) of FABP4gene in Nanjiang yellow goat, and T546C was synonymous mutation. Three genotypes (CC, TC and CC) were obtained by analyzing T143C and T546C with MunⅠ-RFLP and Ssi Ⅰ-RFLP, respectively. Polymorphic analysis showed that both of them were moderately polymorphic loci, and they were in Hardy-Weinberg equilibrium in goat herd.