Isolation of a strong Arabidopsis guard cell promoter and its potential as a research tool.
Isolation of a strong Arabidopsis guard cell promoter and its potential as a research tool.
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DOI：
10.1186/1746-4811-4-6
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发表时间：
2008-02-19
期刊：
影响因子：
5.1
通讯作者：
Schroeder, Julian I.
中科院分区：
文献类型：
--
作者：
Yang, Yingzhen;Costa, Alex;Leonhardt, Nathalie;Siegel, Robert S.;Schroeder, Julian I.
保卫细胞信号传导研究中的一个常见限制是，使用已知的保卫细胞启动子或组成型35 S花椰菜花叶病毒启动子难以在拟南芥保卫细胞中获得感兴趣的转基因的一致高表达。35 S启动子的另一个缺点是在整个生物体中异位表达基因可能导致多效性效应。为了改进现有的靶向基因表达的方法，在保卫细胞，我们分离出强大的保卫细胞启动子候选人的基础上，新的保卫细胞特异性微阵列分析的23,000个基因，可与本报告。启动子pGC 1（At 1g 22690）在保卫细胞中驱动强且相对特异的报告基因表达，包括GUS（β-葡萄糖醛酸酶）和黄色骆驼YC3.60（基于GFP的钙FRET报告基因）。未成熟保卫细胞中报告基因表达较弱。YC 3.60的表达足够强，以成像完整植物的保卫细胞中的细胞内Ca 2+动态，并解决保卫细胞中的自发钙瞬变。GC 1启动子还介导了气孔发育突变体too-many-mouths（tmm）中气孔簇的强报告基因表达。此外，相同的启动子：：报告基因构建体也驱动烟草中的保卫细胞特异性报告基因表达，说明该启动子作为保卫细胞中高水平表达的方法的潜力。启动子的连续缺失定义了保卫细胞表达启动子区。此外，使用pGC 1的反义阻遏对于减少保卫细胞中的特异性GFP基因表达是有力的，而叶表皮细胞中的表达不被阻遏，证明了强的细胞型优先基因阻遏。这里描述的pGC 1启动子驱动拟南芥和烟草植物的保卫细胞中的强报告基因表达。它为靶向保卫细胞表达或基因沉默提供了有力的研究工具。它也适用于减少保卫细胞中的特异性基因表达，提供了一种规避遗传冗余和致死性引起的限制的方法。这些进展对于操纵保卫细胞中的信号传导途径和改变植物在胁迫条件下的表现非常有用。此外，新的保卫细胞和叶肉细胞特异性23，000基因微阵列数据在这里公开。
A common limitation in guard cell signaling research is that it is difficult to obtain consistent high expression of transgenes of interest in Arabidopsis guard cells using known guard cell promoters or the constitutive 35S cauliflower mosaic virus promoter. An additional drawback of the 35S promoter is that ectopically expressing a gene throughout the organism could cause pleiotropic effects. To improve available methods for targeted gene expression in guard cells, we isolated strong guard cell promoter candidates based on new guard cell-specific microarray analyses of 23,000 genes that are made available together with this report. A promoter, pGC1(At1g22690), drove strong and relatively specific reporter gene expression in guard cells including GUS (beta-glucuronidase) and yellow cameleon YC3.60 (GFP-based calcium FRET reporter). Reporter gene expression was weaker in immature guard cells. The expression of YC3.60 was sufficiently strong to image intracellular Ca2+ dynamics in guard cells of intact plants and resolved spontaneous calcium transients in guard cells. The GC1 promoter also mediated strong reporter expression in clustered stomata in the stomatal development mutant too-many-mouths (tmm). Furthermore, the same promoter::reporter constructs also drove guard cell specific reporter expression in tobacco, illustrating the potential of this promoter as a method for high level expression in guard cells. A serial deletion of the promoter defined a guard cell expression promoter region. In addition, anti-sense repression using pGC1 was powerful for reducing specific GFP gene expression in guard cells while expression in leaf epidermal cells was not repressed, demonstrating strong cell-type preferential gene repression. The pGC1 promoter described here drives strong reporter expression in guard cells of Arabidopsis and tobacco plants. It provides a potent research tool for targeted guard cell expression or gene silencing. It is also applicable to reduce specific gene expression in guard cells, providing a method for circumvention of limitations arising from genetic redundancy and lethality. These advances could be very useful for manipulating signaling pathways in guard cells and modifying plant performance under stress conditions. In addition, new guard cell and mesophyll cell-specific 23,000 gene microarray data are made publicly available here.
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7.4
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