Ohme-Takagi M, Shinshi H
Plant Molecular Biology Laboratory, National Institute of Bioscience and Human Technology, Ibaraki, Japan.
Plant Cell. 1995 Feb;7(2):173-82. doi: 10.1105/tpc.7.2.173.
We demonstrated that the GCC box, which is an 11-bp sequence (TAAGAGCCGCC) conserved in the 5' upstream region of ethylene-inducible pathogenesis-related protein genes in Nicotiana spp and in some other plants, is the sequence that is essential for ethylene responsiveness when incorporated into a heterologous promoter. Competitive gel retardation assays showed DNA binding activities to be specific to the GCC box sequence in tobacco nuclear extracts. Four different cDNAs encoding DNA binding proteins specific for the GCC box sequence were isolated, and their products were designated ethylene-responsive element binding proteins (EREBPs). The deduced amino acid sequences of EREBPs exhibited no homology with those of known DNA binding proteins or transcription factors; neither did the deduced proteins contain a basic leucine zipper or zinc finger motif. The DNA binding domain was identified within a region of 59 amino acid residues that was common to all four deduced EREBPs. Regions highly homologous to the DNA binding domain of EREBPs were found in proteins deduced from the cDNAs of various plants, suggesting that this domain is evolutionarily conserved in plants. RNA gel blot analysis revealed that accumulation of mRNAs for EREBPs was induced by ethylene, but individual EREBPs exhibited different patterns of expression.
我们证明，GCC框是一个在烟草属植物及其他一些植物中乙烯诱导的病程相关蛋白基因5'上游区域保守的11碱基序列（TAAGAGCCGCC），当它被整合到异源启动子时，是乙烯应答所必需的序列。竞争性凝胶阻滞试验表明，烟草核提取物中对GCC框序列具有特异性的DNA结合活性。分离出了4种编码对GCC框序列具有特异性的DNA结合蛋白的不同cDNA，并将其产物命名为乙烯应答元件结合蛋白（EREBPs）。EREBPs的推导氨基酸序列与已知DNA结合蛋白或转录因子的氨基酸序列无同源性；推导的蛋白质也不包含碱性亮氨酸拉链或锌指基序。在所有4种推导的EREBPs共有的59个氨基酸残基区域内鉴定出了DNA结合结构域。在各种植物cDNA推导的蛋白质中发现了与EREBPs的DNA结合结构域高度同源的区域，表明该结构域在植物中是进化保守的。RNA凝胶印迹分析表明，乙烯诱导了EREBPs的mRNA积累，但各个EREBPs表现出不同的表达模式。
