摘要
为探究含DENN结构域蛋白1B (DENND1B)对网格蛋白依赖性内吞接头蛋白复合体(AP-2)的β亚基(AP2B1)的影响,本实验利用si RNA干扰技术以及慢病毒包装过表达系统改变DENND1B蛋白的表达水平,通过荧光定量PCR(qPCR)、western blot以及激光共聚焦技术研究DENND1B蛋白对AP2B1蛋白的表达以及细胞内定位的影响。结果显示:DENND1B蛋白沉默后能够下调AP2B1蛋白的表达,而过表达DENND1B后能够上调AP2B1蛋白的表达,并且干扰DENND1B蛋白表达后能够显著性地改变AP2B1在细胞内的定位(p<0.05),使AP2B1蛋白由核周聚集改变为胞质弥散分布。上述结果表明DENND1B蛋白能够调控AP2B1蛋白的表达以及细胞定位。本实验为研究接头蛋白以及网格蛋白依赖性的质膜内吞途径提供了新的切入点。
To investigate the effect of DENND1B protein on AP-2 complex beta subunit(AP2B1), si RNA interference and lentiviral packaging mediated overexpression were used to down or up-regulate the expression of DENND1 B. The expression and localization of AP2B1 were detected by quantitative real-time fluorescence PCR, western blot and laser scanning confocal microscopy. The results showed that the expression and the lacalization of AP2B1 in the cytoplasm were significantly changed in DENND1B specific si RNA-treated A549 cells. By contrast, the expression of AP2B1 was increased in DENND1B-overexpressing A549 cells. The results demonstrate that DENND1B can regulate the expression and localization of AP2B1. Our findings provide a new clue to better understand and study cargo adaptor complex and clathrin-mediated endocytosis.
作者
李奇兵
赵玉辉
王广文
温霞
梁立滨
李俊平
黄山雨
孙楠
姜丽
陈化兰
李呈军
LI Qi-bing;ZHAO Yu-hui;WANG Guang-wen;WEN Xia;LIANG Li-bin;LI Jun-ping;HUANG Shan-yu;SUN Nan;JIANG Li;CHEN Hua-lan;LI Cheng-jun(Animal Influenza Key Laboratory of the Ministry of Agriculture and Rural Affairs,State Key Laboratory of veterinary Biotechnology,Harbin Veterinary Research Institute,Chinese Academy of Agricultural Sciences,Harbin 150069,China)
出处
《中国预防兽医学报》
CAS
CSCD
北大核心
2019年第9期940-944,共5页
Chinese Journal of Preventive Veterinary Medicine
基金
国家重点研发计划项目(2016YFD0500205)
