利用酵母双杂交技术筛选与AtbZIP1相互作用的蛋白质  

Identification of Transcription Factor AtbZIP1-Interactive Proteins Using Yeast Two-hybrid System

作　　者：孙晓丽[1] 段小红[1] 才华[1] 李勇[1] 柏锡[1] 纪巍[1] 季佐军[1] 朱延明[1] 

机构地区：[1]东北农业大学生命科学学院植物生物工程研究室,哈尔滨150030

出　　处：《中国生物化学与分子生物学报》2010年第11期1050-1058,共9页Chinese Journal of Biochemistry and Molecular Biology

基　　金：国家自然科学基金项目(No.30940005No.30570990);东北农业大学创新团队(No.CXT004)~~

摘　　要：碱性亮氨酸拉链bZIP类转录因子在植物的生长发育、光形态建成、光信号传导及非生物胁迫反应中发挥重要的作用.为研究AtbZIP1基因的作用机理,本研究首先验证了该基因的自激活转录活性,通过缺失突变确定了该转录因子的转录激活结构域;以AtbZIP1缺失突变体AtbZ3为诱饵蛋白,采用Matchmaker Gold Yeast Two-Hybrid System(Clonetch),共筛选获得5个与诱饵蛋白相互作用的蛋白质;并通过AbA(Aureobasidin A)抗生素标记基因,His营养缺陷和LacZ蓝白斑检测验证了阳性克隆.亚细胞定位分析发现,AtbZIP1蛋白除了定位于细胞核外,还定位于叶绿体细胞.通过分析这些靶蛋白的已知功能,为研究AtbZIP1蛋白的未知生物学功能提供重要信息.bZIP transcription factors play important roles in various biological processes of plant growth and development,photomorphogenesis,light signal transduction and abiotic stress response.Using a yeast two-hybrid system,we isolated and identified proteins interacted with AtbZIP1 to explore the functions and mechanisms of AtbZIP1.With the serial deletion mutants,the transactive domain of AtbZIP1 that activated the gene itself was identified.Then an Arabidopsis cDNA library was screened using the AtbZ3 deletion mutant as the bait.Five target proteins were fished out and were verified using the AbA antibiotic marker from the Y2H yeast line or the His heterotrophia marker and the LacZ marker from the AH109 line.We found that AtbZIP1 located in both nuclei and chloroplasts,which correlated best with one of the target proteins-chloroplast ATP synthase CF0 subunit.Further analyses of these AtbZIP1 interacting proteins may help to understand the functions the AtbZIP1 gene.

关 键 词：碱性亮氨酸拉链 酵母双杂交系统 相互作用蛋白 叶绿体亚细胞定位 

分 类 号：Q789[生物学—分子生物学]