PROTEOMIC STUDY OF BRASSINOSTEROID SIGNAL TRANSDUCTION

油菜素甾醇信号转导的蛋白质组学研究

• 批准号: 7601819
• 负责人: ZHIYONG WANG
• 金额: $ 1.71万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-09-30 至 2008-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7601819
• 关键词: Affinity Chromatography; Biochemical; Cell Surface Receptors; Cell membrane; Classification; Computer Retrieval of Information on Scientific Projects Database; Coupled; DNA Binding; Funding; Gene Expression; Genetic; Goals; Grant; Institution; Mass Spectrum Analysis; Metals; Methods; Molecular; Nuclear; Phosphoproteins; Phosphorylation; Phosphorylation Site; Phosphotransferases; Plants; Proteins; Proteomics; Research; Research Personnel; Research Project Grants; Resources; Role; Signal Transduction; Site-Directed Mutagenesis; Source; Steroids; Testing; Transgenic Organisms; Two-Dimensional Gel Electrophoresis; United States National Institutes of Health; multicatalytic endopeptidase complex; research study; response; steroid hormone; transcription factor; two-dimensional

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Brassinosteriods (BRs) are steroid hormones with important roles in plants. BR signal is perceived by the cell-surface receptor kinase BRI1, which initiates a cascade of protein phosphorylation leading to nuclear gene expression and cellular responses. Downstream BR signaling involves the cytoplasmic GSK3/SHAGGY-like kinase BIN2. BIN2 phosphorylates the transcription factors BZR1 and BZR2/BES1. Phosphorylation of the transcription factors inhibits their DNA binding and nuclear localization and promotes their degradation by the proteasome. The goal of this study is to understand BR signal transduction at the biochemical and proteomic levels. First, we will identify the BIN2-phosphorylation sites of BZR1 using mass spectrometry (MS). The importance of some of the phosphorylation sites will be further tested by site-directed-mutagenesis and transgenic experiments. Second, in order to understand how BR signaling regulates BIN2 activity, we will purify BIN2-interacting proteins and identify them using MS analysis. Third, we are identifying BR-regulated proteins using two-dimensional difference gel electrophoresis (2-D DIGE) coupled with mass spectrometry. We will identify BR-regulated proteins by performing systematic 2-D DIGE/MS analysis of total, plasma membrane, and phosphoprotein fractions. Forth, we plan to study BR regulated protein phosphorylation using immobilized metal affinity chromatography and the iTRAQ method. The functions of the identified proteins will be studied using genetic and transgenic experiments. This research project will advance our understanding of the molecular mechanism for steroid responses in plants, which will have broad implications in our understanding of steroid actions in general.

该副本是利用众多研究子项目之一 由NIH/NCRR资助的中心赠款提供的资源。子弹和 调查员（PI）可能已经从其他NIH来源获得了主要资金， 因此可以在其他清晰的条目中代表。列出的机构是 对于中心，这不一定是调查员的机构。 铜菜（BRS）是类固醇激素，在植物中起着重要作用。细胞表面受体激酶BRI1感知到BR信号，该细胞表面受体激酶BRI1启动了一系列蛋白质磷酸化，从而导致核基因表达和细胞反应。下游BR信号传导涉及细胞质GSK3/shaggy样激酶BIN2。 BIN2磷酸化转录因子BZR1和BZR2/BES1。转录因子的磷酸化抑制其DNA结合和核定位，并促进蛋白酶体的降解。这项研究的目的是了解生化和蛋白质组学水平的BR信号转导。首先，我们将使用质谱法（MS）确定BZR1的BIN2-磷酸化位点。某些磷酸化位点的重要性将通过位置指导的 - 抗衰测和转基因实验进一步测试。其次，为了了解BR信号如何调节BIN2活性，我们将净化BIN2相互作用的蛋白质并使用MS分析识别它们。第三，我们使用二维差异凝胶电泳（2-D DIGE）与质谱法鉴定BR调节的蛋白质。我们将通过对质膜，质膜和磷蛋白级分的系统分析进行系统的2-D DIGE/MS分析来鉴定BR调节的蛋白质。第四，我们计划使用固定的金属亲和力色谱和ITRAQ方法研究BR调节的蛋白质磷酸化。将使用遗传和转基因实验研究所鉴定蛋白的功能。该研究项目将促进我们对植物中类固醇反应的分子机制的理解，这将对我们对类固醇作用的理解具有广泛的影响。