Niu X, Guiltinan M J
Department of Horticulture, Pennsylvania State University, University Park 16802.
Nucleic Acids Res. 1994 Nov 25;22(23):4969-78. doi: 10.1093/nar/22.23.4969.
EmBP-1 is a wheat DNA binding protein of the basic leucine zipper (bZIP) class of transcription factors implicated in the mechanisms of abscisic acid mediated gene activation. Understanding the role of EmBP-1 in regulating gene transcription requires elucidation of its DNA binding specificity. The binding of EmBP-1 was studied using gel shift selection of DNA from random sequence pools. DNA binding sites were identified by sequencing of a selected pool and by cloning and sequencing individual sites. The binding sites were compared by mobility shift assay and DNase I footprinting, which show that EmBP-1 binds to a family of sequences with varying degrees of affinity. The highest affinity site bound by EmBP-1 is the palindrome GCCACGTGGC. EmBP-1 also binds several other sequences with high affinity, however most of these are asymmetric. While nearly all sequences bound by EmBP-1 contain an ACGT core sequence, EmBP-1 can also bind at least two sites with altered cores. These results provide a basis for comparing the DNA binding specificity of EmBP-1 with those of other plant bZIP proteins and provide insight into the possible target sites which EmBP-1 might bind in vivo.
EmBP-1是一种小麦DNA结合蛋白，属于碱性亮氨酸拉链（bZIP）类转录因子，与脱落酸介导的基因激活机制有关。了解EmBP-1在调节基因转录中的作用需要阐明其DNA结合特异性。使用从随机序列库中进行凝胶迁移选择的方法研究了EmBP-1的结合情况。通过对选定库进行测序以及对单个位点进行克隆和测序来鉴定DNA结合位点。通过迁移率变动分析和DNase I足迹分析对结合位点进行了比较，结果表明EmBP-1与一系列具有不同亲和力的序列结合。EmBP-1结合的亲和力最高的位点是回文序列GCCACGTGGC。EmBP-1还与其他几个序列具有高亲和力结合，然而其中大多数是不对称的。虽然几乎所有被EmBP-1结合的序列都包含一个ACGT核心序列，但EmBP-1也可以与至少两个核心序列改变的位点结合。这些结果为比较EmBP-1与其他植物bZIP蛋白的DNA结合特异性提供了基础，并为EmBP-1在体内可能结合的靶位点提供了见解。
