α-TTP互作蛋白MYO1D介导肝细胞α-生育酚的转运  

α-TTP Interacting Protein MYO1D Mediatesα-Tocopherol Transport in Hepatocytes

作　　者：薛瑛 简路洋 罗海玲[1] XUE Ying;JIAN Lu-Yang;LUO Hai-Ling(College of Animal Science and Technology/State Key Laboratory of Animal Nutrition,China Agricultural University,Beijing 100193,China)

机构地区：[1]中国农业大学动物科学技术学院/动物营养学国家重点实验室,北京100193

出　　处：《农业生物技术学报》2022年第11期2163-2173,共11页Journal of Agricultural Biotechnology

基　　金：国家自然科学基金(31872990)。

摘　　要：维生素E在提高畜禽繁殖机能、提高免疫力及保护肝脏中起着非常重要的作用,其储存和代谢主要发生在肝脏中。α-生育酚转移蛋白(α-tocopherol transfer protein,α-TTP)是一种主要在肝脏中表达的蛋白质,可以特异性结合α-生育酚并促进其从肝脏转运到外周组织吸收利用。但有研究显示α-TTP在转运α-生育酚的过程中可能还需要其他互作因子的作用。本研究通过免疫共沉淀(co-immunoprecipitation,Co-IP)和Pull-down筛选获得α-TTP的互作蛋白—肌球蛋白1D(myosin 1d,MYO1D),免疫荧光观察显示MYO1D与α-TTP存在明显共定位;利用Myo1d-shRNA慢病毒质粒,构建Myo1d基因的干扰载体,筛选获得稳定感染慢病毒干扰载体的肝细胞系;qRT-PCR检测结果显示,干扰组Myo1d基因表达极显著降低(P<0.01),干扰效率为46.83%,干扰组α-ttp基因表达无显著变化;试剂盒检测结果显示,Myo1d干扰组细胞内α-生育酚含量显著升高(P<0.05)。上述结果提示,MYO1D可以与α-TTP相互作用介导肝细胞中α-生育酚转运,可能在运输α-生育酚-TTP复合物的过程中提供动力。本研究为筛选α-TTP转运α-生育酚过程中的互作因子提供了实验基础,为进一步研究肝脏内α-TTP转运α-生育酚的机制提供了参考依据。Vitamin E plays a very important role in improving the reproductive function of livestock and poultry,improving immunity and protecting the liver,and its storage and metabolism mainly occurs in the liver.α-tocopherol transfer protein(α-TTP)is mainly expressed in the liver,which can specifically bindα-tocopherol and promote its transport from the liver to peripheral tissues for absorption and utilization.However,some studies have shown thatα-TTP may also require other interacting factors in the process of transportingα-tocopherol.In this study,using co-immunoprecipitation(Co-IP)and Pull-down screening,theα-TTP-interacting protein-myosin 1d(MYO1D)was obtained.Immunofluorescence observation showed that MYO1D andα-TTP co-localized significantly.The Myo1d-shRNA lentivirus plasmid was used to construct the interference vector of the Myo1d gene,and the hepatocytes line stably infected with the lentiviral interference vector was obtained by screening.The results of qRT-PCR showed that the Myo1d gene expression in the interference group decreased significantly(P<0.01),and the interference efficiency was 46.83%,while the expression ofα-ttp gene in the interference group had no significant change.The results of the kit test showed that the content ofα-tocopherol in the Myo1d interference group increased significantly(P<0.05).The above results suggest that MYO1D could interact withα-TTP to mediateα-tocopherol transport in hepatocytes,and might provide power in the process of transportingα-tocopherol-TTP complexes.These findings provide experimental bases for screening the interaction factors in the process ofα-TTP transportingα-tocopherol,and provide a reference basis for further research on the mechanism ofα-TTP transportingα-tocopherol in the liver.

关 键 词：生育酚转移蛋白(α-TTP)互作蛋白 肌球蛋白1D(MYO1D) SHRNA慢病毒 α-生育酚转运 

分 类 号：S826[农业科学—畜牧学]