Possible Involvement of a bZIP Protein in the Repression of ABI3/VP1-mediated Chymotrypsin Inhibitor Gene Expression at the Late-seed Maturation in Winged Bean
Possible Involvement of a bZIP Protein in the Repression of ABI3/VP1-mediated Chymotrypsin Inhibitor Gene Expression at the Late-seed Maturation in Winged Bean
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发表时间：
2006-12
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通讯作者：
Y. Sakata;Kuwano Tomomasa;Takaoka Sachiko;Takashima Noriko;Taji Teruaki;Takenaga Hiroshi;Tanaka Shigeo
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作者：
Y. Sakata;Kuwano Tomomasa;Takaoka Sachiko;Takashima Noriko;Taji Teruaki;Takenaga Hiroshi;Tanaka Shigeo
ABI-/VP+是一种重要的转录因子，通过ABA反应元件或保守的RY重复序列调控植物胚胎成熟过程中多种基因的表达。RY重复序列经常存在于种子特异基因的上游区域。通过对四季豆胰凝乳酶抑制因子(WCI)基因的时间和空间调控表达的广泛研究，我们证明了ry重复是种子特异表达所必需的，但不是必需的。在本研究中，我们从四季豆中克隆了编码ABI-/VP+样因子(WbABI-)和bZIP DNA结合蛋白的cDNAs，以研究ABI-/VP+和bZIP型转录因子在WCI基因表达中的作用。推导的bZIP蛋白序列(WbZIP+)与ROM3高度同源，在菜豆种子成熟后期抑制ABI-/VP+激活的MAT类基因转录。制备了细菌重组WbZIP+蛋白，并用凝胶迁移率改变实验验证了其与WCI-基因启动子区域的结合，WCI-基因编码四季豆主要的WCI蛋白。重组WbZIP+蛋白对含有WCI启动子/ACGT-序列的特定片段具有较高的亲和力。WCI-和WbAB基因在种子成熟中期瞬时表达后，WbZIP+基因在种子成熟后期的表达增强。这些结果表明，WbABI和WbZIP+可能相互拮抗地调节四季豆种子成熟中后期WCI基因的表达水平。
ABI-/VP+ is an important transcription factor, which regulates the expression of many kinds of genes during plant embryo maturation via ABA responsive elements or conserved RY repeats. The RY repeats are frequently found in / upstream regions of seed-specific genes. Through an extensive study of temporallyand spatially-regulated expression of a winged bean chymotrypsin inhibitor (WCI) gene, we have demonstrated that the RY repeat is necessary but not su$cient for the seed-specific expression. In this study, we have cloned cDNAs encoding an ABI-/VP+ like factor (WbABI-) and a bZIP DNA binding protein from winged bean to investigate the participation of ABI-/ VP+ and bZIP-type transcription factors in WCI gene expression. The deduced protein sequence of the bZIP protein (WbZIP+) was highly homologous to ROM,, which is shown to be a repressor against ABI-/ VP+-activated transcription of MAT class genes during late-seed maturation in French bean. Bacterial recombinant WbZIP+ protein was prepared and tested in a gel mobility shift assay to verify its binding to the promoter region of the WCI-gene, which encodes a major WCI protein of winged bean. The recombinant WbZIP+ protein proved to show a high a$nity for specific fragments containing / ACGT-sequences from the WCI-promoter. Enhanced expression of the WbZIP+ gene was observed during late-stage seed maturation after the transient expression of WCI-and WbABIin mid-stage seed maturation. These results suggest that WbABIand WbZIP+ may function antagonisitically to tune the level of WCI gene expression from midthrough late-stage seed maturation in winged bean.