Signal-Responsive Transcription of Plant Nuclear Genes

植物核基因的信号响应转录

• 批准号: 9527364
• 负责人: Jonathan Arias
• 金额: $ 4.6万
• 依托单位: University of Maryland Biotechnology Institute
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 1996
• 资助国家: 美国
• 起止时间: 1996-04-01 至 1998-03-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=9527364&HistoricalAwards=false
• 关键词: Signal; Responsive; Transcription; Plant; Nuclear

9527364 Arias The goal of these studies is to better understand how the expression of protein-encoding genes of plants are regulated by environmental stimuli. A central hypothesis to be tested here is that defense gene expression in plants is modulated by a phosphorylation-dependent change in the transcriptional activity of G-box binding factors. Aside from understanding a process of general importance to all higher eukaryotes the proposed studies may ultimately lead to the development of novel and sustainable forms of crop protection. Changes in the transcription activity of plant GBF's in response to defense cues (eg., nonhost bacterial species or biotic elicitors) cDNA's of these factors will be fused to a novel DNA-binding domain and expressed in plant cells containing a reporter gene. Changes in the phosphorylation of these chimeric GBF's will be assessed by phosphopeptide mapping. Protein kinases that phosphorylate GBF's in response to defense stimuli will be biochemically purified to obtain N-terminal sequences for cloning GBF-protein kinase cDNA's. Cloned putative kinases will be tested in vivo for their ability to phosphorylate GBF and act in defense-signaling. To identify potential mediators of plant GBF-dependent transcription a far-western screening of plant cDNA expression libraries with labeled GBF as probe will be performed. Cloned proteins will by functionally characterized by standard protein binding and transfection techniques. %%% Higher plants resist microbial infection in part by synthesizing phytoalexins. The rapid and transient activation of "defense" genes that encode for enzymes in phytoalexin biosynthesis largely determines the timing and magnitude of this response. Recent evidence suggests that protein kinases couple defense signal perception to enhanced gene expression. For example, inhibitors of protein kinases block the expression of plant defense genes, whereas agents that stimulate protein kinase activities in other metazoan systems both enhance pla nt defense gene expression and result in the transient phosphorylation of a few cellular proteins. How plant gene building factors function in signal-dependent transcription is currently unknown but, like in animals, may involve changes in the phosphorylation state of the factor. ***

9527364 Arias这些研究的目标是更好地了解植物蛋白质编码基因的表达是如何受到环境刺激的调控的。这里要检验的一个中心假设是，植物中的防御基因表达是由G-box结合因子转录活性的磷酸化依赖的变化来调节的。除了了解对所有高等真核生物具有普遍重要性的过程外，拟议的研究可能最终导致开发新的和可持续的作物保护形式。植物GBF转录活性的变化响应于这些因子的防御信号(如非寄主细菌或生物激发子)的cDNAs将被融合到一个新的DNA结合域，并在含有报告基因的植物细胞中表达。这些嵌合GBF的磷酸化变化将通过磷酸肽图谱进行评估。对在防御刺激下磷酸化GBF的蛋白激酶进行生化纯化，以获得克隆GBF-蛋白激酶cDNAs的N-端序列。克隆的可能的蛋白激酶将在体内测试它们对GBF的磷酸化和参与防御信号的能力。为了确定植物GBF依赖转录的潜在介体，我们将以标记的GBF为探针，对植物表达文库进行远距离筛选。克隆的蛋白质将通过标准的蛋白质结合和转染技术进行功能表征。高等植物抵抗微生物侵染的部分途径是合成植物保卫素。在植保素生物合成中编码酶的“防御”基因的快速和瞬时激活在很大程度上决定了这种反应的时间和幅度。最近的证据表明，蛋白激酶将防御信号感知与增强的基因表达结合在一起。例如，蛋白激酶的抑制剂阻止了植物防御基因的表达，而在其他后生动物系统中刺激蛋白激酶活性的药物既增强了植物防御基因的表达，又导致了少数细胞蛋白的瞬时磷酸化。植物基因构建因子如何在信号依赖的转录中发挥作用目前尚不清楚，但像动物一样，可能涉及该因子的磷酸化状态的变化。***