Auxin Response Factor Specificity in Arabidopsis

描述（由申请人提供）：目前生长素初级反应的模型主要基于生长素感知机制的最新发现，生长素反应因子（ARF蛋白）结合DNA的体外研究，以及生长素反应基因表达数据。生长素响应启动子的定义特征知之甚少，并且尚未研究指导体内arf启动子相互作用的特异性决定因素。解决这些问题是理解生长素主要反应和证实生长素在转录中的作用模型的关键下一步。拟议研究的结果将对植物和动物的激素反应的研究具有广泛的效用，并将特别与植物生长和发育的研究有关。本课题的实验目标是：1)通过生长素应答基因表达和染色质免疫沉淀（ChIP）分析，结合转录因子MP/ARF5和NPH4/ARF7，确定ARF靶位点；2)通过整合和比较生物信息学分析新数据集和现有数据集，定义MP/ARF5-和NPH4/ arf7招募启动子；3)利用定量启动子报告分析和与截断和嵌合的ARF蛋白共免疫沉淀的DNA序列分析，确定ARF蛋白-启动子相互作用特异性的决定因素

DESCRIPTION (provided by applicant): The current model of the primary auxin response is based largely on recent findings regarding the mechanism of auxin perception, on in vitro studies of DNA binding by Auxin Response Factors (ARF proteins), and on auxin-responsive gene expression data. The defining features of auxin-responsive promoters are very poorly understood, and the determinants of specificity guiding in vivo ARF-promoter interactions have not been investigated. Addressing these questions is a critical next step in understanding the primary auxin response and substantiating the model of auxin action in transcription. The results of the proposed studies will be of broad utility to studies of hormone responses in plants and animals, and will be of particular relevance to studies of plant growth and development. With the experiments presented in this proposal, I aim to 1) identify ARF target loci through analysis of auxin-responsive gene expression and Chromatin Immunoprecipitation (ChIP) with the transcription factors MP/ARF5 and NPH4/ARF7; 2) define MP/ARF5- and NPH4/ARF7-recruiting promoters through consolidation and comparative bioinformatics analysis of new and existing datasets; 3) define determinants of specificity in ARF protein-promoter interactions using quantitative promoter-reporter assays and analysis of DNA sequences coimmunoprecipitating with truncated and chimeric ARF proteins