Arabidopsis 2010: Systematic Discovery of Ligands for Arabidopsis START proteins

智力上的优点。植物的生长在一定程度上受称为激素的小分子信号控制。这些激素通过与受体蛋白结合来发挥作用，受体蛋白反过来又起到协调基因转录和细胞生长等反应的作用。例如，激素脱落酸(ABA)是植物在逆境时合成的一种小的亲脂化合物。一旦合成，它就会与受体蛋白结合，进而抑制生长并激活适应压力环境条件所需的转录反应。许多激素受体对已经被定义并且已知控制植物生长的许多方面。因此，激素信号及其受体蛋白是现代植物生物学研究的中心问题。有了完整的基因组序列，很明显，高等植物的基因组编码的受体蛋白比已知的激素多数百个；这一点和其他观察表明，存在的激素比我们目前所知的要多。鉴于植物激素的重要性已经得到了充分的证明，发现激素的新方法和工具可能会对我们对植物生物学的理解产生革命性的影响。原则上，识别激素的一种方法是通过它们与受体蛋白的结合来纯化它们。为了开发、探索和实施这一想法，该项目将执行一种简单而一般的策略来识别与受体蛋白结合的化合物。该项目的重点是START蛋白，它构成了一个已知或可疑的激素受体大家族。拟南芥基因组编码89个起始蛋白，其中几个在发育、非生物或生物胁迫生物学中发挥重要作用。尽管它们很重要，但大多数START蛋白的内源性配体尚不清楚。在这个项目中，我们利用蛋白质-配体鉴定方面的最新技术进步来系统地表征拟南芥Start蛋白的配体。利用最近开发的小分子下拉方法，我们将使用START蛋白作为试剂，指导其内源配体的鉴定和随后的纯化。一旦分离出来，这些配体的结构将使用高灵敏度的微探针2D-核磁共振和其他方法相结合来确定。我们的建议建立在不同方法的综合基础上，并为新一轮以蛋白质为目标的配体发现奠定了坚实的基础，这将使发现新的激素成为可能。Broader Impact这一跨学科项目将为学生和博士后提供一个国际上独特的植物化学生物学和遗传学培训环境。此外，通过UCR？S REU计划和其他途径，将对包括未被充分代表的少数民族在内的本科生进行代谢物分析、纯化、分离和一般植物生物学方法方面的培训。该协会维护着一个网站，提供有关植物化学基因组学和化学生物学的信息和技术建议。这个网站是获取实验室化学资源信息的门户，PI已经与世界各地的数十个实验室共享了这些资源。小分子下拉和代谢物纯化的方案将公开提供，并通过PI的网站进行传播。该项目将系统地定义整个蛋白质家族的蛋白质-代谢物相互作用，这在任何生物体中都是史无前例的，并将创造一个令人兴奋的跨学科培训环境，培养出准备迎接与采用新方法解决重要生物学问题相关的挑战的科学家。此外，新激素和生长调节剂的发现预计将对农业产生长期好处。

Intellectual merit. Plant growth is controlled in part by small molecule signals called hormones. These hormones work by binding to receptor proteins, which in turn function to coordinate responses like gene transcription and cellular growth. For example, the hormone abscisic acid (ABA) is a small lipophilic compound that is synthesized by plants in times of stress. Once synthesized, it binds to a receptor protein, which in turn inhibits growth and activates the transcriptional responses needed for acclimation to stressful environmental conditions. Numerous hormone-receptor pairs have been defined and are known to control many aspects of plant growth. Thus, hormone signals and their receptor proteins are a central preoccupation of modern plant biological studies. With complete genome sequences in hand, it is apparent that the genomes of higher plants encode hundreds more receptor proteins than known hormones; this and other observations imply that more hormones exist than we currently know about. Given the well-documented importance of plant hormones, new methods and tools for discovering hormones may have a transformative influence on our understanding of plant biology.In principle, one way to identify hormones is to purify them by virtue of their association with their receptor proteins. To develop, explore and implement this idea, this project will execute a simple and general strategy for identifying compounds that bind to receptor proteins. The project focuses on START proteins, which form a large family of known or suspected hormone receptors. The Arabidopsis genome encodes 89 START proteins, several of which play important roles in developmental, abiotic or biotic stress biology. In spite of their importance, the endogenous ligands for most START proteins are not known. In this project, we leverage recent technological advances in protein-ligand identification to systematically characterize the ligands of Arabidopsis START proteins. Using recently developed small-molecule pull down methods, we will use START proteins as reagents that guide the identification and subsequent purification of their endogenous ligands. Once isolated, the structure of these ligands will be determined using a combination of high-sensitivity microprobe 2D-NMR and other methods. Our proposal rests on an integrative base of diverse approaches and builds a robust foundation for a new wave of protein-targeted ligand discovery, which will enable the discovery of new hormones.Broader impacts This interdisciplinary project will provide students and post-docs an internationally unique training environment for plant chemical biology and genetics. In addition, through UCR?s REU program and other avenues, undergraduates, including underrepresented minorities will be trained in methods for metabolite analysis, purification, isolation and general plant biology. The PI maintains a website that provides information and technical advice about plant chemical genomics and chemical biology. This site is gateway to information about the laboratory's chemical resources that the PI has shared with dozens of laboratories across the world. Protocols for small molecule pull downs and metabolite purification will be made publicly available and disseminated through the PI's website. This project, which will systematically define protein-metabolite interactions for an entire protein family, is unprecedented in any organism and will create an exciting, interdisciplinary training environment that produces scientists ready to embrace the challenges associated with adopting new approaches to important biological questions. The discovery of new hormones and growth regulators is additionally expected to be of long-term benefit to agriculture.