重组人CREG蛋白与溶酶体组织蛋白酶和M6P/IGFⅡR的相互作用  

Interactions between the recombinant human CREG protein and cathepsins and M6P/IGFIIR

作　　者：孙鸣宇[1] 闫承慧[1] 田孝祥[1] 李洋[1] 韩雅玲[1] 

机构地区：[1]解放军沈阳军区总医院全军心血管病研究所心内科,辽宁省沈阳市110016

出　　处：《中国组织工程研究》2015年第37期5961-5965,共5页Chinese Journal of Tissue Engineering Research

基　　金：国家自然科学基金资助项目(81130072;81370243)~~

摘　　要：背景:以往研究证实,CREG是一种与M6P/IGFⅡR直接结合的溶酶体蛋白,并依赖于与M6P受体的相互作用有效转运至溶酶体。目的:分析外源性CREG蛋白与溶酶体组织蛋白酶和M6P/IGFⅡR的相互作用关系及其对M6P/IGFⅡR表达变化及细胞内定位的影响。方法:应用细胞免疫荧光双染和免疫共沉淀方法,观察外源性CREG蛋白与溶酶体组织蛋白酶和M6P/IGFⅡR的相互作用关系,并应用gain-of-function和loss-of-function模型,通过Western blot和细胞免疫荧光双染方法,观察CREG对M6P/IGFⅡR表达变化及细胞内定位的影响。结果与结论:细胞免疫荧光双染和免疫共沉淀方法证实外源性CREG蛋白与溶酶体组织蛋白酶和M6P/IGFⅡR有直接相互作用关系;应用gain-of-function和loss-of-function模型进一步证实,CREG对M6P/IGFⅡR表达变化无影响,而影响其在细胞内的定位。提示外源性CREG蛋白定位于溶酶体,且与溶酶体组织蛋白酶和M6P/IGFⅡR有相互作用关系,并影响M6P/IGFⅡR的细胞内定位。BACKGROUND;It has been found that cellular repressor of E1A-stimulated genes（CREG） is a lysosomal protein binding directly to the mannose-6-phosphate（M6P）/insulin-like growth factor II receptor（IGFIIR） and depends on the interaction with M6 P receptors for efficient delivery to lysosomesOBJECTIVE;To study the interactions between the exogenous CREG protein and cathepsins and M6P/IGFIIR and to confirm the effect of CREG protein on expression and distribution of M6P/IGFIIR.METHODS;Double-stained immunofluorescence and coimmunoprecipitation were applied to observe the interactions between the exogenous CREG protein and cathepsin B,cathepsin L and M6P/IGFIIR.Using gain-of-function and loss-of-function approaches,the effect of CREG on expression and distribution of M6P/IGFIIR were studied by western blot assay and immunofluorescence staining.RESULTS AND CONCLUSION;Double-stained immunofluorescence and coimmunoprecipitation analysesconfirmed the direct interactions between the exogenous CREG protein and cathepsin B,cathepsin L and M6P/IGFIIR.It was verified that CREG plays a critical role not in the expression but in the distribution of M6P/IGFIIR using gain-of-function and loss-of-function approaches.These findings provide evidence that exogenous CREG protein is located in lysosomes and has interactions with cathepsins and M6P/IGFIIR,also CREG plays a critical role in the distribution of M6P/IGFIIR.

关 键 词：组织构建 组织工程 E1A激活基因阻遏子 国家自然科学基金 

分 类 号：R318[医药卫生—生物医学工程]