GLUCOSE SIGNAL TRANSDUCTION PATHWAY IN PLANTS

这项研究的长期目标是以拟南芥为模型系统，阐明植物葡萄糖信号转导途径的分子机制。通过光合作用生产糖是植物中最重要的活动，并支持地球上所有的生命。我的实验室最近的研究提供了令人信服的证据来支持这一新概念，即糖作为中央信号分子调节光合作用基因的表达，并协调许多重要过程，包括植物的萌发、幼苗发育、叶和根的分化、开花和衰老。通过利用突变体、瞬时表达和转基因植物，我们提供了新的证据表明，葡萄糖信号可以与葡萄糖代谢解偶联，并且至少有一个特定的己糖激酶(AtHXK1)作为植物中主要的葡萄糖传感器/信号转导因子具有双重催化和调节作用。拟议中的研究采用了一个植物模型A.thaliana，将揭示植物中葡萄糖感应和信号传递的分子机制的非常规和令人兴奋的新见解，并可能对其他真核生物产生影响。将使用分子、生化、遗传和细胞方法的组合。三个具体目标是：1)将AtHXK1介导的葡萄糖反应定义为传感器/信号转导，利用拟南芥AtHXK1基因敲除突变体和HXK水平改变的转基因植株来鉴定HXK介导的葡萄糖反应靶基因。将测试一套全面的葡萄糖依赖生物测定，以确定HXK介导的发育活动。将监测葡萄糖的磷酸化和积累。这些实验将建立分析植物中糖反应的标准，并明确定义特定的HXK作为遗传手段的葡萄糖传感器/信号转导的作用。2)阐明AtHXK1在葡萄糖传感和信号转导中的作用机制综合实验，以确定AtHXK1的表达模式和亚细胞定位，建立AtHXK1功能分析，产生和分析AtHXK1突变体，鉴定AtHXK1蛋白复合体，并确定AtHXK1信号伙伴。3)对葡萄糖不敏感的拟南芥突变体进行表型和分子特征分析，确定葡萄糖信号转导途径的组成成分。我们将分离并分析最有趣的突变基因Gin1。

The long-term objective of this research proposal is to elucidate the molecular mechanisms underlying the glucose signal transduction pathway in plants using Arabidopsis thaliana as a model system. Sugar production through photosynthesis is the most important activity in plants and supports all life on the planet. Recent studies from my laboratory have provided compelling evidence to support the new concept that sugars are acting as central signaling molecules to modulate photosynthetic gene expression and coordinate many vital processes, including germination, seedling development, leaf and root differentiation, flowering, and senescence in plants. By using mutants, transient expression, and transgenic plants, we have provided new evidence that glucose signaling can be uncoupled from glucose metabolism, and at least one specific hexokinase (AtHXK1) has dual catalytic and regulatory roles as a major glucose sensor/signal transducer in plants. The proposed studies employing a plant model A.thaliana will reveal unconventional and exciting new insights into the molecular mechanisms of glucose sensing and signaling in plants with potential implications in other eukaryotes. A combination of molecular, biochemical, genetic, and cellular approaches will be used. Three specific aims are: 1) Define the glucose responses mediated through AtHXK1 as a sensor/signal transducer An Arabidopsis AtHXK1 knockout mutant and transgenic plants with altered HXK levels will be used to identify HXK- mediated glucose-responsive target genes. A comprehensive sets of glucose-dependent bioassays will be tested to define HXK- mediated developmental activities. Glucose phosphorylation and accumulation will be monitored. These experiments will establish the standards for analyzing sugar responses in plants and will unequivocally define the role of specific HXK as a glucose sensor/signal transducer by genetic means. 2) Elucidate the molecular mechanism of AtHXK1 function in glucose sensing and signaling Comprehensive experiments are proposed to determine the expression patterns and subcellular localization of AtHXK1, establish AtHXK1 functional assays, generate and analyze AtHXK1 mutants, characterize the AtHXK1 protein complex, and identify AtHXK1 signaling partners. 3) Identify the components in the glucose signal transduction pathways The Arabidopsis mutants that are glucose insensitive (gin) will be phenotypically and molecularly characterized. The gene for the most interesting mutant gin1 will be isolated and analyzed.