利用酵母双杂交技术筛选与AtbZIP1相互作用的蛋白质

孙晓丽, 段小红, 才华, 李勇, 柏锡, 纪巍, 季佐军, 朱延明

中国生物化学与分子生物学报 ›› 2010, Vol. 26 ›› Issue (11) : 1050-1058.

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中国生物化学与分子生物学报 ›› 2010, Vol. 26 ›› Issue (11) : 1050-1058.
研究论文

利用酵母双杂交技术筛选与AtbZIP1相互作用的蛋白质

  • 孙晓丽,   段小红,   才   华,   李   勇,   柏   锡,   纪   巍,   季佐军,   朱延明
作者信息 +

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

  • SUN Xiao-Li, DUAN Xiao-Hong, CAI Hua, LI Yong, BAI Xi, JI Wei, JI Zuo-Jun, ZHU Yan-Ming
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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.

Abstract

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

/ font-size: 10.5pt / mso-ascii-font-family: 'Times New Roman' / mso-hansi-font-family: 'Times New Roman' / mso-bidi-font-size: 12.0pt / mso-bidi-font-family: 'Times New Roman' / mso-font-kerning: 1.0pt / mso-ansi-language: EN-US / mso-fareast-language: ZH-CN / mso-bidi-language: AR-SA">碱性亮氨酸拉链 / / font-size: 10.5pt / mso-ascii-font-family: 'Times New Roman' / mso-hansi-font-family: 'Times New Roman' / mso-bidi-font-size: 12.0pt / mso-bidi-font-family: 'Times New Roman' / mso-font-kerning: 1.0pt / mso-ansi-language: EN-US / mso-fareast-language: ZH-CN / mso-bidi-language: AR-SA">酵母双杂交系统 / 相互作用蛋白 / 叶绿体定位

Key words

bZIP / yeast two-hybrid / protein interaction / chloroplast subcellular localization

引用本文

导出引用
孙晓丽, 段小红, 才华, 李勇, 柏锡, 纪巍, 季佐军, 朱延明. 利用酵母双杂交技术筛选与AtbZIP1相互作用的蛋白质[J]. 中国生物化学与分子生物学报, 2010, 26(11): 1050-1058
SUN Xiao-Li, DUAN Xiao-Hong, CAI Hua, LI Yong, BAI Xi, JI Wei, JI Zuo-Jun, Zhu-Yan-Ming.  Identification of Transcription Factor AtbZIP1-Interactive Proteins Using Yeast Two-hybrid System[J]. Chinese Journal of Biochemistry and Molecular Biology, 2010, 26(11): 1050-1058
中图分类号:      Q789   

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基金

 

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