Regulation of proteasome assembly in auxin signaling

激素生长素对植物的大量细胞和发育过程是必不可少的。生长素的许多作用是通过诱导大量基因的表达来实现必要的功能。AUX/IAA蛋白家族通常抑制生长素反应基因的表达。生长素诱导基因表达的中间步骤是通过泛素化/26S蛋白酶体调节过程降解这些蛋白质，该过程依赖于几个多组分蛋白质复合体的相互作用。这种生长素反应途径的干预步骤还远未被了解。先前的研究揭示了RAC/Rop GTP酶在生长素刺激与AUX/IAA降解的细胞内机制之间的作用。生长素诱导的AUX/IAA蛋白降解过程包括响应信号而即兴组装降解装置，并受AUX/IAA底物的存在调节。细胞培养和全植物研究表明，生长素和底物共同诱导蛋白质复合体的形成，这些复合体在细胞核中以蛋白体的形式出现(称为核蛋白体，NPB)。SCFTIR1、CSN和26S蛋白酶体的底物和组分被募集到这些蛋白复合体中并在其中共存，但它们被募集到NPB中和离开NPB的动力学仍有待阐明。这些NPB具有生物活性，因为已观察到这些结构内的底物水平因生长素处理而下降。这项研究的主要重点是检查这些NPB的生化性质、组成和活性，以及是否存在与生长素信号有关的高分子量蛋白质复合体，或其他已知受泛素化和内源条件下26S蛋白酶体介导的蛋白分解调节的更顺从的信号通路。泛素化和26S蛋白酶体调控的蛋白水解酶参与了真核生物中大量的细胞和发育过程，包括细胞的分裂、生长和分化，以及从胚胎发生到生殖、衰老和细胞程序性死亡的发育途径。该系统的故障会导致严重的发育问题，包括使人衰弱的神经系统疾病和人类的癌症，以及植物应对压力和病原体的能力受损。已经推测了一种受调控的动态过程，用于组装和拆解分子机制，以实现对选定蛋白质的降解。所获得的信息对于理解生长素信号将是重要的，生长素信号本身是植物生物学的一个重要方面，并且代表了植物细胞信号的一个范例。这些知识也应该适用于其他泛素化/26S蛋白酶体调节过程。拟议的研究将为博士后、研究生和本科生提供培训机会。PIS实验室目前的本科生包括四名女性和两名男性(包括美国公民，但来自阿富汗、尼泊尔和乌克兰)；其中三人参与了与拟议研究相关的研究。今年夏天，全校范围内针对少数族裔学生的NSF支持项目(SPUR)将招收新生。

The hormone auxin is essential for a large number of cellular and developmental processes in plants. Auxin achieves many of its actions by inducing the expression of a large number of genes to carry out the necessary functions. The Aux/IAA protein family normally suppresses auxin-responsive gene expression. An intermediate step in auxin induced gene expression is the degradation of these proteins through a ubiquitination/ 26S proteasome regulated process that depends on the interaction of several multi-component protein complexes. The intervening steps for this auxin response pathway are far from being understood. Prior studies revealed the contribution of the Rac/Rop GTPases in relating the auxin stimulus to the intracellular machinery for Aux/IAA degradation. The auxin induced Aux/IAA protein degradation process involves impromptu assembly of the degradation apparatus in response to the signal and is regulated by the presence of the Aux/IAA substrates. Observations from cell culture and whole plant studies suggest that auxin and substrates together induce formation of protein complexes that appear as protein bodies in the nucleus (referred to as nuclear protein bodies, NPBs). Substrates and components of SCFTIR1, CSN and 26S proteasome are recruited into and co-exist in these protein complexes, although the dynamics of their recruitment into and exit out of the NPBs remain to be elucidated. These NPBs are biologically active since the level of substrates has been observed to decline from within these structures in response to auxin treatment. The main focus of the proposed research is to examine the biochemical property of these NPBs, their composition and activity, and to examine the presence of high molecular weight protein complexes involved in auxin signaling, or other more amenable signaling pathways that are known to be regulated by ubiquitination and 26S proteasome-mediated proteolysis under endogenous conditions.Broader Impact. Ubiquitination and 26S proteasome-regulated proteolysis underlies a large number of cellular and developmental processes in all eukaryotes, including cell division, growth and differentiation, and developmental pathways from embryogenesis to reproduction, senescence and programmed cell death. Malfunctioning of this system results in severe developmental problems, including debilitating neurological diseases and cancer in human and compromised ability to cope with stress and pathogens in plants. A regulated and dynamic process for assembly and disassembly of the molecular machinery to carry out degradation of selected proteins has been speculated. Information obtained will be important toward understanding auxin signaling, which in itself is a significant aspect of plant biology and represents a paradigm in plant cell signaling. The knowledge should also be applicable to other ubiquitination/26S proteasome regulated processes. The proposed research will provide training opportunities for postdocs, graduate students and undergraduates. The current group of undergraduates in the PIs laboratories includes four women and two men (including US citizens but natives of Afghanistan, Nepal and Ukraine); three of them are participating in the research related to the proposed studies. New students from a campus-wide NSF supported program for minority students (SPUR), will be recruited to start this summer.