Molecular Genetic Analysis of Ethylene Insensitive Loci in Arabidopsis

植物如何感知和转导激素信号以影响形态和功能的巨大变化是生物学中的一个基本问题，同时也具有重要的实际应用价值。对乙烯气体的反应继续作为理解植物激素信号转导机制的范例。乙烯在植物的发育过程中起着至关重要的作用，例如果实的成熟，以及对各种物理和生物胁迫（如病原体攻击）的反应。为了了解这些生物过程背后的详细分子机制，正在使用参考植物拟南芥采用遗传，分子，生化和基因组方法。特别是，本项目的目标是表征乙烯反应的主要调节蛋白EIN3/EIL家族的功能。EIN3编码一种dna结合转录激活因子，是一个由5个EIN3样蛋白组成的小基因家族的成员。这些蛋白参与转录调控级联反应，控制近1000个乙烯反应基因的表达。将采用生化和遗传学方法鉴定EIN3/EIL直接靶基因。基因表达的全基因组研究和体内靶标鉴定将使用全基因组平铺阵列进行。乙烯反应表型的系统突变筛选将通过纯合基因敲除的全基因组“单突变”收集和化学基因组学筛选方法启动。新的乙烯信号通路基因的鉴定和表征及其与已知信号通路组分相互作用的分析将为简单碳氢化合物乙烯的生物效应多样性提供新的见解。此外，了解这些蛋白质的功能将使我们有能力改变乙烯对任何植物的有益和/或有害影响，特别是对具有重要经济或社会价值的作物。(2)拟议活动所产生的更广泛影响。这项研究的长期目标是了解乙烯气体如何在非常详细的机械水平上促进植物发育和抗逆性的无数变化。与探索植物激素信号传导过程基础知识相关的教育活动包括对四个层次学生的培训：高中、本科、研究生和博士后。使用参考植物拟南芥的激素信号研究计划的一个方面是假设驱动的，但这些研究越来越多地采用大规模，数据驱动或基于发现的科学。实际上，传统的假设驱动的“逐基因”研究以及大规模收集“部分列表”信息（如转录组、相互作用组、表型组）在现代生物学研究中都发挥着重要作用，特别是在系统水平上理解这些复杂信号通路的相互作用。这些学生将学习这些新思想/概念，并将应用基因组学和计算生物学的工具和技术，开始解决植物科学中长期存在的问题，如乙烯对生长的调节，将他们的学习经验扩展到传统的课堂/实验室环境之外。

(1) Intellectual merit of the proposed activity How plants perceive and transduce hormone signals to affect dramatic changes in form and function is a fundamental question in biology that also has important practical applications. The response to ethylene gas continues to serve as a paradigm for understanding the mechanisms of plant hormone signal transduction. Ethylene plays critical roles in development, such as in the ripening of fruits and in responses to a variety of physical and biological stresses, such as pathogen attack. In order to understand the detailed molecular mechanisms that underlie these biological processes, genetic, molecular, biochemical and genomic approaches are being employed using the reference plant Arabidopsis. In particular, the goal of this project is to characterize the functions of the EIN3/EIL family of master regulatory proteins of the ethylene response. EIN3 encodes a DNA-binding transcriptional activator and is a member of a small gene family of five EIN3-LIKE proteins. These proteins participate in a transcriptional regulatory cascade that controls the expression of nearly 1,000 ethylene response genes. Biochemical and genetic approaches will be employed to identify direct EIN3/EIL target genes. Genome-wide studies of gene expression and in vivo target identification will be conducted using whole genome tiling arrays. Systematic mutant screens for ethylene response phenotypes will be initiated using a whole genome "uni-mutant" collection of homozygous gene knockouts and through the use of a chemical genomics screening approach. Identification and characterization of novel ethylene signaling pathway genes and analysis of their interactions with the known signaling pathway components will provide new insights into the diversity of biological effects of the simple hydrocarbon, ethylene. Moreover, understanding of the functions of these proteins will allow an ability to modify the beneficial and/or detrimental effects of ethylene in any plant, in particular, crops with important economic or social value.(2) Broader impacts resulting from the proposed activity.The long-term goal of this research is to understand how ethylene gas promotes myriad changes in plant development and stress resistance at a very detailed, mechanistic level. The educational activities associated with this quest for basic knowledge about plant hormone signaling processes includes the training of students at four levels: high school, undergraduate, graduate and postdoctoral. One aspect of this research program on hormone signaling using the reference plant Arabidopsis is hypothesis driven but a growing part of these studies employs large-scale, data-driven or discovery-based science. In reality, both conventional hypothesis-driven, "gene-by-gene" studies as well as large-scale gathering of "part-lists" information (e.g. transcriptome, interactome, phenome) play important roles in modern biological research, in particular, to understand the interactions of these complex signaling pathways at a systems level. These students will learn these new ideas/concepts and will apply the tools and technologies of genomic and computational biology to begin to solve longstanding problems in plant science such as the regulation of growth by ethylene, extending their learning experiences beyond the traditional classroom/laboratory settings.