【摘要】 目的本研究旨在研究Annexin A11蛋白(ANXA11)N端突变p.G38R和p.D40G对该蛋白亚细胞定位的影响,探索其引起肌萎缩侧索硬化症(ALS)的机制。方法构建表达ANXA11野生型蛋白(wt)及p.G38R和p.D40G突变蛋白的真核表达载体,采用激光共聚焦显微镜检测3种蛋白在HEK293细胞中的亚定位。结果 ANXA11wt、p.G38R及p.D40 G蛋白均可在HEK 293细胞中形成囊泡状结构,其中p.G38 R组囊泡面积大于wt组(P=0.002)和p.D40 G组(P=0.006),后两组间无明显差异(P=0.791)。在单个细胞中,ANXA11 wt形成的囊泡数目大于p.G38 R组(P=0.002)和p.D40 G组(P<0.001),而p.G38 R组和p.D40 G组差异不明显(P=0.516)。与ANXA11 wt相比,p.G38 R蛋白(P<0.001)和p.D40 G蛋白(P<0.001)在细胞核内的分布显著减少,后两者间无明显差异(P=0.519)。结论 p.G38 R和p.D40 G突变减少了ANXA11蛋白相关囊泡状结构的数目和ANXA11蛋白在细胞核的分布,这可能是引起ALS发病的机制之一。更多还原

【Abstract】 Objective To investigate the influence of the N-terminal mutations of Annexin A11( ANXA11) protein,p. G38 R and p.D40 G,on the subcellular localization of this protein and their mechanism of action in inducing amyotrophic lateral sclerosis( ALS).Methods Eukaryotic expression vectors which expressed ANXA11 wild-type protein,ANXA11 p. G38 R protein,or ANXA11 p. D40 G protein were constructed,and a laser scanning confocal microscope was used to investigate the subcellular localization of these three proteins in HEK293 cells. Results ANXA11 wild-type protein,ANXA11 p. G38 R protein,and ANXA11 p. D40 G protein formed a vesicular structure in HEK293 cells,and the p. G38 R group had significantly larger vesicles than the ANXA11 wild-type group( P =0. 002) and the p. D40 G group( P = 0. 006),while there was no significant difference between the latter two groups( P = 0. 791). In a single cell,the ANXA11 wild-type group had significantly more vesicles than the p. G38 R group( P = 0. 002) and the p. D40 G group( P < 0. 001),and there was no significant difference between the p. G38 R group and the p. D40 G group( P = 0. 516). Compared with ANXA11 wild-type protein,p. G38 R protein and p. D40 G protein had significantly reduced distribution in the nucleus( both P <0. 001),and there was no significant difference between p. G38 R protein and p. D40 G protein( P = 0. 519). Conclusions The p.G38 R and p. D40 G mutations reduce the number of vesicles of ANXA11 protein and the distribution of ANXA11 protein in the nucleus,which may be one of the mechanism of ALS.更多还原