Involvement of TAAAG elements suggests a role for Dof transcription factors in guard cell-specific gene expression
Involvement of TAAAG elements suggests a role for Dof transcription factors in guard cell-specific gene expression
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DOI：
10.1046/j.1365-313x.2001.01166.x
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发表时间：
2001-11-01
期刊：
影响因子：
7.2
通讯作者：
Mueller-Roeber, B
中科院分区：
文献类型：
--
作者：
Plesch, G;Ehrhardt, T;Mueller-Roeber, B
由于其独特的结构和功能，保护细胞在生理水平上受到了广泛的关注。然而，我们对保护细胞特异性的决定和维持所涉及的分子事件知之甚少。KST1基因编码马铃薯保护细胞的K+内流通道，因此被选为研究保护细胞特异性基因表达调控的模型。携带KST1启动子与大肠杆菌uidA （β -葡萄糖醛酸酶）报告基因融合的转基因马铃薯植株显示，启动子在保护细胞和花中具有活性。对KST1启动子的详细解剖发现了两个独立的小TATA盒近端调控单元，每个单元都足以指导保护细胞特异性基因的转录。这两个片段都包含序列基序5'-TAAAG-3'，它与一类新的锌指转录因子（称为Dof蛋白）的已知靶点有关。在不同启动子结构的背景下，这些Dof靶点的阻断突变显著降低了保护细胞启动子的活性。我们克隆了一个Dof基因StDof1，发现它在保护细胞高度富集的表皮片段中表达。在凝胶阻滞实验中，StDof1蛋白以序列特异性的方式与含有TAAAG基序的KST1启动子片段相互作用。这些结果证明TAAAG元件是反式Dof蛋白控制保护细胞特异性基因表达的靶点。我们的数据将有助于设计量身定制的保护细胞启动子，作为保护细胞功能的分子工程的进一步工具，从而控制作物植物的气孔二氧化碳（CO2）吸收和水分流失。
Due to their unique structure and function, guard cells have attracted much attention at the physiological level. Very little, however, is known about the molecular events involved in the determination and maintenance of guard cell specificity. The KST1 gene encodes a K+ influx channel of guard cells in potato, and was therefore chosen as a model to study regulation of guard cell-specific gene expression. Transgenic potato plants carrying a fusion between the KST1 promoter and the E. coli uidA (beta -glucuronidase) reporter gene revealed promoter activity in guard cells and in flowers. A detailed dissection of the KST1 promoter led to the discovery of two independent small TATA box-proximal regulatory units, each of which was sufficient to direct guard cell-specific gene transcription. Both fragments contain the sequence motif, 5'-TAAAG-3', which is related to known target sites for a novel class of zinc finger transcription factors, called Dof proteins. Block mutagenesis of these Dof target sites in the context of different promoter constructs dramatically reduced guard cell promoter activity. A Dof gene, StDof1, was cloned and shown to be expressed in epidermal fragments highly enriched for guard cells. In gel retardation experiments, the StDof1 protein interacted in a sequence-specific manner with a KST1 promoter fragment containing the TAAAG motif. These results provide evidence that TAAAG elements are target sites for trans-acting Dof proteins controlling guard cell-specific gene expression. Our data will add to the design of tailor-made guard cell promoters as a further tool in molecular engineering of guard cell function and, hence, control of stomatal carbon dioxide (CO2) uptake and water loss in crop plants.