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SIRT7对奶牛乳腺上皮细胞乳蛋白、乳脂和乳糖合成关键基因表达的调控 预览

Effects of SIRT7 on the expression of key genes involved in lactoprotein,milk fat and lactose synthesis in dairy cow mammary epithelial cells
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摘要 [目的]本试验旨在探讨去乙酰化蛋白7(sirtuin 7,SIRT7)对奶牛乳腺上皮细胞(DCMEC)合成乳蛋白、乳脂和乳糖关键基因表达的影响。[方法]以体外原代培养的中国荷斯坦奶牛DCMEC为试验材料,纯化后转染质粒pCDNA3.1-SIRT7,继续培养48或72h后收集细胞。采用RT-qPCR和Western blot法检测乳蛋白、乳脂和乳糖合成关键基因mRNA和蛋白的表达;通过特异性试剂盒检测胞内和培养基中甘油三脂(TG)含量。[结果]1)乳蛋白:与对照组相比,过表达SIRT7可显著提高αs1-酪蛋白基因(CSN1S1)、β-酪蛋白基因(CSN2)和ETS转录因子基因(ELF5)mRNA水平,促进CSN2蛋白合成,激活雷帕霉素靶蛋白(mTOR)磷酸化;2)乳脂:与对照组相比,过表达SIRT7可显著提高乳脂合成关键因子过氧化物酶增殖体激活受体γ(PPARγ)、固醇调节元件结合蛋白1(SREBP1)mRNA和蛋白水平,显著增加细胞内甘油三酯(TG)含量;3)乳糖:与对照组相比,过表达SIRT7可显著提高乳糖合成关键因子葡萄糖转运蛋白1(GLUT1)mRNA表达。[结论]在体外培养条件下,过表达SIRT7可促进DCMEC中乳蛋白、乳脂和乳糖合成关键基因的表达。 [Objectives] The purpose of this study was to explore the roles of sirtuin7(SIRT7)in the expression of lactoprotein,milk fat and lactose synthesis-related genes in dairy cow mammary epithelial cells(DCMEC).[Methods] DCMEC were collected from Chinese Holstein cows and cultured in vitro.The cells were purified and transfected with pcDNA3.1- SIRT 7 by Lipofectamine 3000,and cultured 48 or 72 h for experiments.RT-qPCR and Western blot were used to detect the relative genes and protein expression about lactoprotein,milk fat,and lactose synthesis.The content of triglyceride(TG)in cells and culture medium were measured by a special kit.[Results] 1)Lactoprotein:overexpression of SIRT7 significantly increased the mRNA level of casein alpha s1( CSN1S1 ),β-casein ( CSN2 )and E74-like factor 5( ELF5 ),promoted the protein expression of CSN2,and increased the phosphorylation of mTOR compared to the control group;2)Milk fat:overexpression of SIRT7 significantly increased the milk fat synthesis-related genes and protein expression( PPARγ and SREBP1 ),compared to the control group.Meanwhile,overexpression of SIRT7 significantly increased intracellular TG content;3)Lactose:comparing to the control group,overexpression of SIRT7 significantly increased the mRNA level of glucose transporter 1( GLUT1 ).[Conclusions] Overexpression of SIRT7 could significantly promote the relative genes and protein levels that involved in lactoprotein,milk fat and lactose synthesis in DCMEC.
作者 陈坤琳 钱勇 蒋临正 王慧利 邢光东 李倩倩 仲跻峰 CHEN Kunlin;QIAN Yong;JIANG Linzheng;WANG Huili;XING Guangdong;LI Qianqian;ZHONG Jifeng(Institute of Animal Science/Key Laboratory of Crop and Animal Integrated Farming,Ministry of Agriculture and Rural Affairs,Jiangsu Academy of Agricultural Sciences,Nanjing 210014,China;Youyuan Research Institute of Dairy Industry Co.Ltd.,Nanjing 211100,China;Bright Farming Co.Ltd.,Shanghai 200436,China)
出处 《南京农业大学学报》 CAS CSCD 北大核心 2019年第5期917-923,共7页 Journal of Nanjing Agricultural University
基金 国家自然科学基金项目(31871248) 江苏省自然科学基金项目(BK20161377) 中国博士后基金项目(2019M651755).
关键词 奶牛乳腺上皮细胞 去乙酰化蛋白7 乳蛋白 乳脂 乳糖 dairy cow mammary epithelial cells sirtuin 7 lactoprotein milk fat lactose
作者简介 陈坤琳,助理研究员,主要从事奶牛生理调控方向研究,E-mail:chenkunlin@jaas.ac.cn;通信作者:仲跻峰,研究员,主要从事奶牛育种与繁殖研究,E-mail:zhongjifeng@jaas.ac.cn。
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