RACK1与CLIC1的相互作用鉴定  

The interaction of RACK1 with CLIC1 in cells

作　　者：李彩红 朱亮亮 王蓓华 周恒 李强 朱昉修 范礼斌 Li Caihong;Zhu Liangliang;Wang Beihua;Zhou Heng;Li Qiang;Zhu Fangxiu;Fan Libin(Dept of Biology,School of Life Science,Anhui Medical University,Hefei 230032)

机构地区：[1]安徽医科大学生命科学学院生物系,合肥230032

出　　处：《安徽医科大学学报》2022年第10期1518-1522,共5页Acta Universitatis Medicinalis Anhui

基　　金：安徽省自然科学基金项目(编号:11040606M170);安徽医科大学双肩挑干部学术恢复科研专项(编号:0112011109)。

摘　　要：目的探究胞内氯离子通道蛋白1(CLIC1)与活化蛋白激酶C受体1(RACK1)在细胞内的相互作用。方法构建重组质粒pcDNA3.1-RACK1-HA,转染质粒pcDNA3.1-RACK1-HA和(或)pcDNA3.1-CLIC1-FLAG到HEK 293T细胞中,转染pcDNA3.1-RACK1-HA和(或)pcDNA3.1-CLIC1-FLAG到COS7中。GST-pulldown和免疫共沉淀实验确定CLIC1和RACK1在体内及体外的相互作用。间接免疫荧光实验检测RACK1与CLIC1在细胞中的定位。结果成功构建pcDNA3.1-RACK1-HA,Western blot检测结果证明了RACK1和CLIC1可在HEK 293T细胞中表达,GST-pulldown结果显示RACK1与CLIC1在体外可直接结合,免疫共沉淀结果显示RACK1和CLIC1可在细胞内结合。间接免疫荧光实验表明RACK1与CLIC1可共定位于细胞质。结论RACK1与CLIC1可在细胞中结合。Objective To investigate the interaction between intracellular chloride ion protein 1(CLIC1)and activated protein kinase C receptor 1(RACK1).Methods Plasmids pcDNA3.1-RACK1-HA and/or pcDNA3.1-CLIC1-FLAG were transfected into HEK 293T cells,and pcDNA3.1-RACK1-HA and/or pcDNA3.1-CLIC1-FLAG were transfected into COS7.GST-pulldown and immunoprecipitation assays were performed to determine the interaction between CLIC1 and RACK1 in vivo and in vitro.The co-localization of CLIC1 and RACK1 was observed by indirect immunofluorescence assay.Results Western blot confirmed that CLIC1 and RACK1 could be highly expressed in HEK 293T cells.GST-pulldown showed that RACK1 bound CLIC1 in vitro,and immunoprecipitation showed that CLIC1 and RACK1 interacted in vivo.Furthermore,indirect immunofluorescence assay showed CLIC1 co-localized with RACK1.Conclusion Human CLIC1 protein interacts with RACK1 in vitro and in vivo.

关 键 词：RACK1 CLIC1 相互作用 

分 类 号：R34[医药卫生—基础医学] Q28[生物学—细胞生物学]