Hong J C, Cheong Y H, Nagao R T, Bahk J D, Key J L, Cho M J
Plant Molecular Biology and Biotechnology Research Center, Gyeongsang National University, Chinju, Korea.
Plant J. 1995 Aug;8(2):199-211. doi: 10.1046/j.1365-313x.1995.08020199.x.
G-box binding factors (GBFs) constitute a family of plant DNA-binding proteins that bind to the G-box motif, a regulatory cis element present in many plant genes with a palindromic DNA motif of CACGTG. Previously TCCACGTGTC, a G-box motif, from an auxin responsive gene GmAux28 has been identified as a sequence-specific protein-binding site. Here the isolation of two soybean cDNA clones, referred to as SGBF-1 and SGBF-2, encoding proteins which bind to the G-box motif is reported. The primary structure of SGBF-1 and SGBF-2 predicts that these proteins contain a basic leucine zipper (bZIP) DNA-binding domain and an N-terminal proline-rich domain. A dramatic difference in the pattern of protein-DNA complex formation was observed when recombinant SGBF-1 and SGBF-2 proteins were analyzed by electrophoretic mobility shift assays (EMSAs). The SGBF-1 binding pattern obtained with the G-box probe resulted in three major retarded bands while the SGBF-2 formed a single complex. This shows that the characteristically diffuse banding pattern of plant nuclear proteins interacting with the G-box is also observed in a binding assay using only one recombinant GBF. EMSAs were performed with a few selected binding sequences to study the effect of flanking nucleotides to the hexanucleotide G-box core motif. The binding specificity of the SGBF proteins resembles that described for type A cauliflower nuclear G-box binding proteins which bind class I G-box elements [(G/T)(C/A)CACGTG(G/T)(A/C)]. Phylogenetic analysis of 13 GBF-like proteins from various plant species reveals that the SGBF-1 and SGBF-2 proteins belong to different lineages, suggesting that they may have distinct functions in activating transcription.
G盒结合因子（GBF）构成了一类植物DNA结合蛋白家族，它们与G盒基序结合，G盒基序是一种存在于许多植物基因中的调控顺式元件，具有回文DNA基序CACGTG。此前，来自生长素响应基因GmAux28的G盒基序TCCACGTGTC已被鉴定为序列特异性蛋白结合位点。本文报道了两个大豆cDNA克隆SGBF - 1和SGBF - 2的分离，它们编码与G盒基序结合的蛋白质。SGBF - 1和SGBF - 2的一级结构预测这些蛋白质含有一个碱性亮氨酸拉链（bZIP）DNA结合结构域和一个N端富含脯氨酸的结构域。当通过电泳迁移率变动分析（EMSA）分析重组SGBF - 1和SGBF - 2蛋白时，观察到蛋白质 - DNA复合物形成模式存在显著差异。用G盒探针获得的SGBF - 1结合模式产生了三条主要的滞后带，而SGBF - 2形成了一个单一复合物。这表明在仅使用一种重组GBF的结合试验中也观察到了植物核蛋白与G盒相互作用时典型的弥散条带模式。用一些选定的结合序列进行EMSA，以研究侧翼核苷酸对六核苷酸G盒核心基序的影响。SGBF蛋白的结合特异性类似于对结合I类G盒元件[（G/T）（C/A）CACGTG（G/T）（A/C）]的A型花椰菜核G盒结合蛋白所描述的特异性。对来自不同植物物种的13种GBF样蛋白的系统发育分析表明，SGBF - 1和SGBF - 2蛋白属于不同的谱系，这表明它们在激活转录方面可能具有不同的功能。
