REGULATION OF AUXIN RESPONSE FACTOR ACTIVITY IN ARABIDOPSIS

 描述（由申请人提供）：激素介导的通过转录因子对基因激活或抑制的调节对所有生物体都至关重要。生长素反应因子（ARF）转录因子是植物生长的重要调节因子，为研究激素对基因激活和阻遏的调控提供了理想的模型。我们最近发现蛋白质多聚化和蛋白酶体降解是调节ARF活性的两种以前未知的机制。本研究项目的长期目标是确定这些ARF调控机制在拟南芥转录控制中的重要性。阐明ARF调控的分子机制可能会揭示其他转录因子共同的控制过程。 抑制-去抑制模式调节ARF活性。在低浓度的激素生长素下，ARF转录活性被Aux/IAA阻遏蛋白抑制。当生长素浓度增加时，由F-box蛋白（TIR 1）和Aux/IAA阻遏蛋白组成的辅助受体复合物直接结合生长素。F-box蛋白参与Skp 1-Cullin-F-box（SCF）E3泛素连接酶，其靶向Aux/IAA蛋白进行降解。这种降解事件解除ARF抑制，从而允许生长素调节的基因转录。虽然这种生长素活性的抑制和去抑制的分子模型似乎相对简单，我们最近的初步数据表明，几个令人兴奋的新的控制组件和翻译后修饰影响ARF转录活性和蛋白质积累。 本项目旨在通过鉴定ARF蛋白活性和积累的调节因子来阐明生长素信号传导的分子机制。为了实现这一目标，我们将使用生物化学，生物物理，细胞生物学，合成生物学，分子和遗传技术的组合，以深入了解影响ARF活动的因素。第一个目的是了解 蛋白质多聚化在ARF转录活性调节中的作用。在合成酵母生长素反应系统和植物中的研究将用于测试这一目标的各个方面，其中包括功能测定和3C分析。第二个目的是了解多聚化在ARF定位调节中的作用。我们将确定是否ARF翻译后修饰影响ARF细胞定位。我们的第三个目标是建立ARF蛋白酶体依赖的降解在调节生长素反应和植物发育的cDNA。我们将使用各种遗传学、生物化学和细胞生物学技术来了解调节ARF稳定性的生物学和发育作用。拟议的研究是创新的，因为我们的方法主要集中在ARF调控的分子理解上，由我们最近关于ARF 7的结构数据指导，它有可能极大地改变生长素信号模型。这项研究具有重要意义，因为它有望以ARF因子为模型，推进和扩展对转录因子调控的理解。

 DESCRIPTION (provided by applicant): Hormone-mediated modulation of gene activation or repression through transcription factors is central to all organisms. Auxin Response Factor (ARF) transcription factors are critical modulators of plant growth and provide an ideal model for exploring hormone control of gene activation and repression. We have recently identified protein multimerization and proteasomal degradation as two previously unknown mechanisms that regulate ARF activity. The long-term goal of this research project is to determine the importance of these ARF regulatory mechanisms in Arabidopsis thaliana transcriptional control. Elucidating the molecular mechanism of ARF regulation likely will uncover control processes common to other transcription factors. A repression-derepression paradigm regulates ARF activity. Under low concentrations of the hormone auxin, ARF transcriptional activity is repressed by Aux/IAA repressor proteins. When auxin concentrations increase, a co-receptor complex, comprised of an F-box protein (TIR1) and an Aux/IAA repressor protein, directly binds auxin. The F-box protein participates in a Skp1-Cullin-F-box (SCF) E3 ubiquitin ligase, which targets the Aux/IAA protein for degradation. This degradation event relieves ARF repression, thereby allowing auxin-regulated gene transcription. Although this molecular model of repression and derepression for auxin activity appears relatively simple, our recent preliminary data suggest several exciting new control components and posttranslational modifiers influence ARF transcriptional activity and protein accumulation. This project aims to elucidate auxin signaling molecular mechanisms by identifying regulators of ARF protein activity and accumulation. To achieve this goal, we will use a combination of biochemical, biophysical, cell biology, synthetic biology, molecular and genetic techniques to gain insight into factors that influence ARF activity. The first aim is to understand the role of protein multimerization in the regulation of ARF transcriptional activity. Studies in both a synthetic yeast auxin response system and in planta will be used to test aspects of this aim, which includes functional assays and 3C analysis. The second aim is understand the role of multimerization in the regulation of ARF localization. We will determine whether ARF posttranslational modification affects ARF cellular localization. Our third aim is to establish rols for ARF proteasome-dependent degradation in regulating auxin response and plant development. We will use a variety of genetic, biochemical, and cell biology techniques to understand the biological and developmental roles for regulated ARF stability. The proposed research is innovative because our approaches focus strongly on the molecular understanding of ARF regulation, guided by our recent structural data on ARF7 and it has the potential to dramatically alter the auxin signaling model. The proposed research is significant because it is expected to advance and expand understanding of transcription factor regulation, using ARF factors as a model.