Dai Lihe, Chu Xiaohong, Lu Fuzeng, Xu Ruhai
Key Laboratory of Animal Genetics and Breeding of Zhejiang Province, Institute of Animal Husbandry and Veterinary Science, Zhejiang Academy of Agricultural Sciences, Hangzhou, 310021, People's Republic of China.
Mol Biol Rep. 2016 Nov;43(11):1305-1313. doi: 10.1007/s11033-016-4068-x. Epub 2016 Aug 26.
As an important triglyceride hydrolase in mammalian cells, patatin-like phospholipase domain-containing 2 (PNPLA2) predominantly performs the first step in triglyceride hydrolysis. The objective of this study was to detect and evaluate the effects of mutations in the 5' upstream region of porcine PNPLA2 gene with fat deposition and carcass traits. Four single nuclear polymorphisms were identified, including g.161969 T>C, g.161962 A>G, g.161953 C>G and g.161904 G>T, and subsequently genotyped in five pure breeds. Three haplotypes were constructed, including H1(CGGT), H2(TACG) and H3(CACT), which were the most abundant haplotypes in Duroc (0.75), Landrace (0.78) and Chinese indigenous breeds (>0.73), respectively. Duroc individuals with the H1H1 diplotype always exhibited the lowest feed conversion ratio (FCR) (P < 0.05), while H2H2 had the thickest backfat thickness (P < 0.05). Landrace individuals with H2H3 had lower backfat thickness (P < 0.05), higher muscle thickness (P < 0.05) and estimated lean meat percentage (P < 0.05) than those with diplotype H2H2 and H3H3. Luciferase assay indicated pGL3-basic-H2 had the highest activity and pGL3-basic-H1 had the lowest activity in driving reporter gene transcription in HEK293 cells in vitro. In H1 haplotype, two GR binding sites and an ERα binding site were predicted to be introduced. While in H2 and H3, there were other transcriptional factor binding sites predicted in H2 and H3, such as Sp1, AP-2 and CAC-binding proteins, which were broadly expressed transcription factors and capable of contributing to basal promoter activity. The reduced basal promoter activity of H1 may be due to the lack of inducement for GR and ERα binding sites in HEK293 cells. The identified functional polymorphisms provide new evidence of PNPLA2 as an important candidate gene for fat deposition and carcass traits in pigs.
作为哺乳动物细胞中一种重要的甘油三酯水解酶，含帕他汀样磷脂酶结构域蛋白2（PNPLA2）主要负责甘油三酯水解的第一步。本研究的目的是检测和评估猪PNPLA2基因5'上游区域突变对脂肪沉积和胴体性状的影响。共鉴定出4个单核苷酸多态性，包括g.161969 T>C、g.161962 A>G、g.161953 C>G和g.161904 G>T，并随后在5个纯品种中进行了基因分型。构建了3种单倍型，包括H1（CGGT）、H2（TACG）和H3（CACT），它们分别是杜洛克猪（0.75）、长白猪（0.78）和中国本土品种（>0.73）中最丰富的单倍型。具有H1H1双倍型的杜洛克个体总是表现出最低的饲料转化率（FCR）（P<0.05），而H2H2个体的背膘最厚（P<0.05）。与具有H2H2和H3H3双倍型的长白猪个体相比，具有H2H3的长白猪个体背膘更薄（P<0.05）、肌肉更厚（P<0.05）且估计瘦肉率更高（P<0.05）。荧光素酶报告基因检测表明，在体外HEK293细胞中，pGL3-basic-H2驱动报告基因转录的活性最高，pGL3-basic-H1的活性最低。在H1单倍型中，预测引入了2个糖皮质激素受体（GR）结合位点和1个雌激素受体α（ERα）结合位点。而在H2和H3单倍型中，预测在H2和H3中有其他转录因子结合位点，如Sp1、AP-2和CAC结合蛋白，它们是广泛表达的转录因子，能够促进基础启动子活性。H1单倍型基础启动子活性降低可能是由于在HEK293细胞中缺乏对GR和ERα结合位点的诱导。所鉴定的功能多态性为PNPLA2作为猪脂肪沉积和胴体性状的重要候选基因提供了新的证据。
