Yang Li, Liu Lili, Cheng Jinhua, Wu Zhengchang, Bao Wenbin, Wu Shenglong
Key Laboratory for Animal Genetics, Breeding, Reproduction and Molecular Design of Jiangsu Province, College of Animal Science and Technology, Yangzhou University, Yangzhou 225009, China.
Institute of Animal Science, Jiangsu Academy of Agricultural Sciences, China.
Gene. 2023 Jan 30;851:147016. doi: 10.1016/j.gene.2022.147016. Epub 2022 Oct 29.
Porcine α-1,3-fucosyltransferase (FUT3), as a member of the fucosyltransferase family, plays an important role in the resistance of the piglet intestine to pathogenic microbial infection. To further investigate the tissue/developmental expression of FUT3 and its regulatory mechanism, we analyzed changes in the expression of FUT3 in the duodenal tissues of Meishan pigs at different ages and found that the expression of FUT3 showed a decreasing trend with increasing age. In addition, bisulfite sequencing identified nine methylated CpG sites in the FUT3 core promoter (-500 ∼ -206) region. Therein, the methylation level at the mC-9 site located in FUT3 showed a significantly negative association with mRNA expression (P < 0.05). A further dual-luciferase assay demonstrated that methylation at the mC-9 site of the FUT3 promoter inhibited its transcriptional activity. Then, we confirmed the binding of Sp1 to the FUT3 promoter using RNA knockdown and a ChIP-qPCR assay. Our findings indicate that DNA methylation at the mC-9 site may inhibit the binding of the transcription factor Sp1, thus regulating the developmental expression of the FUT3 gene in the duodenum, providing some theoretical basis for the FUT3 gene as an important candidate marker of disease resistance in Meishan pigs.
猪α-1,3-岩藻糖基转移酶（FUT3）作为岩藻糖基转移酶家族的一员，在仔猪肠道抵抗病原微生物感染中发挥重要作用。为进一步研究FUT3的组织/发育表达及其调控机制，我们分析了不同年龄梅山猪十二指肠组织中FUT3的表达变化，发现FUT3的表达随年龄增长呈下降趋势。此外，亚硫酸氢盐测序在FUT3核心启动子（-500 ∼ -206）区域鉴定出9个甲基化的CpG位点。其中，FUT3中位于mC-9位点的甲基化水平与mRNA表达呈显著负相关（P < 0.05）。进一步的双荧光素酶检测表明，FUT3启动子mC-9位点的甲基化抑制了其转录活性。然后，我们通过RNA敲低和染色质免疫沉淀定量PCR（ChIP-qPCR）检测证实了Sp1与FUT3启动子的结合。我们的研究结果表明，mC-9位点的DNA甲基化可能抑制转录因子Sp1的结合，从而调节十二指肠中FUT3基因的发育表达，为FUT3基因作为梅山猪抗病性重要候选标记提供了一些理论依据。
