Vincentz M, Leite A, Neshich G, Vriend G, Mattar C, Barros L, Weinberg D, de Almeida E R, de Carvalho M P, Aragão F, Gander E S
Laboratório de Biologia Molecular, Centro Nacional de Recursos Genéticos e Biotecnologia, SAIN, Brasilia-DF, Brazil.
Plant Mol Biol. 1997 Aug;34(6):879-89. doi: 10.1023/a:1005874404706.
The expression of Brazil nut storage albumin genes is highly regulated during seed development. Several sequences in the promoter of one of these genes show homologies with the target sites of the maize O2 bZIP regulatory protein. We therefore asked whether the O2 protein would recognize these promoter sequences. We show that the O2 protein binds to three different sequences (F1, F2 and F3). F1 and F3 are hybrid C/G and A/G boxes, respectively, that are homologous to the O2-binding site of a maize alpha-zein gene. F2 is a new O2-binding sequence related to the O2 target sites of the Coix alpha-coxin, the maize b-32 genes and the AP-1 pseudopalindrome. Molecular modelling showed that an Asn and a Ser in the 02 DNA binding domain make different base-specific contacts with each operator. 5' Promoter deletions of the be2S1 gene showed that the domain containing the O2 target sites F1 and F2 is required for detectable reporter gene expression in transgenic tobacco seeds. Moreover, the homologous coix O2 protein was shown to in situ transactivate the promoter region encompassing the three O2-binding sites F1, F2 and F3. Thus, these sites may be in vivo regulatory sequences mediating activation by bZIP regulatory proteins.
巴西坚果贮藏白蛋白基因的表达在种子发育过程中受到高度调控。这些基因之一的启动子中的几个序列与玉米O2 bZIP调控蛋白的靶位点具有同源性。因此，我们询问O2蛋白是否会识别这些启动子序列。我们发现O2蛋白与三个不同的序列（F1、F2和F3）结合。F1和F3分别是杂合的C/G盒和A/G盒，与玉米α-醇溶蛋白基因的O2结合位点同源。F2是一个新的O2结合序列，与薏苡仁α-伴白蛋白、玉米b-32基因的O2靶位点以及AP-1假回文序列相关。分子建模表明，O2 DNA结合结构域中的一个Asn和一个Ser与每个操纵基因形成不同的碱基特异性接触。be2S1基因的5'端启动子缺失表明，在转基因烟草种子中，含有O2靶位点F1和F2的结构域是可检测的报告基因表达所必需的。此外，同源的薏苡仁O2蛋白被证明能原位反式激活包含三个O2结合位点F1、F2和F3的启动子区域。因此，这些位点可能是体内介导bZIP调控蛋白激活的调控序列。
