畜牧兽医学报2017，Vol.48Issue(11)：2034-2045,12.DOI:10.11843/j.issn.0366-6964.2017.11.004
猪FUT2基因沉默对其所在通路基因表达及小肠上皮细胞E.coli F18黏附能力的影响
The Effects of FUT2 Gene Silencing on the Expression of Genes Involved in Glycosphingolipid Biosynthesis-globo Series Pathway and the Adhesion of E.coli F18 to Small Intestinal Epithelial Cells
摘要
Abstract
The purpose of this study was to further explore the function of FUT2 and its regulation mechanisms in swine epithelial cells infected by E.coli F18 at the cellular level,which provide theoretical basis for molecular breeding of improving the resistance of piglets to E.coli F18.The expression differences of FUT2 gene in intestinal epithelial cells before and after E.coli F18ab,F18ac infection and lipopolysaccharide(LPS) induction were detected by Real-time PCR.Then,the Western blotting was performed to identify the protein expression differences of FUT2 in intestinal epithelial cells before and after E.coli F18ab and F18ac infection.Meanwhile the lentiviral interference vectors of porcine FUT2 were designed and constructed,and the intestinal epithelial cells line with FUT2 gene silencing was obtained.The effects of FUT2 gene silencing on the mRNA expression of key genes (FUT1,ST3GAL1,HEXA,HEXB,B3GALNT1,NAGA)involved in glycosphingolipid biosynthesis-globo series pathway and on E.coli F18 adhesion to intestinal epithelial cells were further analyzed.The results showed that the FUT2 expression levels in intestinal epithelial cells were significantly increased after E.coli infection and LPS induction (P＜0.01),and the FUT2 expression level in intestinal epithelial cells was also increased after E.coli infection.Moreover,the lentiviral interference vectors of porcine FUT2 were suc cessfully constructed,and the intestinal epithelial cells line with FUT2 gene silencing were obtained.After FUT2 gene silencing,the expression level of key genes in glycosphingolipid biosynthesis-globo series pathway were decreased at different degrees.The expression level of FUT1 after FUT2 silencing was significantly reduced (P＜0.05),and the expression levels of ST3GAL1,HEXA,HEXB were extremely significantly reduced (P＜0.01),while the expression levels of B3GALNT1 and NAGA genes showed no significant changes.In addition,the adhesion ability of E.coli F18 to swine epithelial cells was decreased significantly (P＜0.05).These results show that the lower expression level of FUT2 may hinder the formation of E.coli F18 receptor,and thus elevate the resistance capability of porcine small intestine epithelial cells to E.coli F18 infection.Our findings can also provide theoretical foundations and evidence for further investigating on the mechanisms of glycosphingolipid biosynthesis-globo series pathway in resistance of pigs to E.coli infection.关键词
猪/FUT2基因/RNAi/大肠杆菌/球系列鞘糖脂生物合成通路Key words
swine/FUT2 gene/RNAi/Escherichia coli/glycosphingolipid biosynthesis-globo series pathway分类
农业科技引用本文复制引用
孙丽,宗秋芳,吴森,吴嘉韵,吴圣龙,包文斌..猪FUT2基因沉默对其所在通路基因表达及小肠上皮细胞E.coli F18黏附能力的影响[J].畜牧兽医学报,2017,48(11):2034-2045,12.基金项目
国家自然科学基金(31572360 （）
31372285) （）
江苏省重点研发计划(现代农业)(BE2015329 （现代农业）
BE2016315) （）
江苏高校优势学科建设工程资助项目(PAPD) （PAPD）
