Posttranslational Regulation of Cell Growth and Stress Responses

细胞生长和应激反应的翻译后调节

• 批准号: 10676253
• 负责人: Yonghao Yu
• 金额: $ 55.91万
• 依托单位: COLUMBIA UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-05-01 至 2025-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10676253
• 关键词: ADP ribosylation; Animal Experiments; Atlases; Biochemical; Biology; Cells; Cellular Stress; Cellular biology; Chemicals; Collection; Complex; DNA; DNA sequencing; Disease; Elements; Event; FRAP1 gene; Feedback; Genetic Transcription; Genome; Goals; Human; IGFBP5 gene; Link; Mass Spectrum Analysis; Measures; Mediator; Methods; Mission; Molecular; Pathologic; Pathway interactions; Performance; Phosphorylation; Physiology; Post-Transcriptional Regulation; Post-Translational Protein Processing; Post-Translational Regulation; Process; Protein Analysis; Proteins; Proteome; Proteomics; RNA; Reagent; Role; Signal Transduction; Site; Systems Biology; Technology; Therapeutic Intervention; Tissues; biological adaptation to stress; cell growth; cell growth regulation; cellular targeting; extracellular; human disease; improved; innovation; novel; programs; stoichiometry; trait; transcriptome sequencing; treatment strategy

Project Summary A central goal in disease biology is to describe the molecular processes responsible for transformation of a cell from a normal state into a pathological one. Compared to the rapid progresses in DNA and RNA sequencing technologies, characterization of the final and arguably most actionable element of the central dogma, protein, has lagged behind. The dynamic relationship between the genome of a cell and its proteome is poorly understood, reflecting multiple layers of transcriptional/post-transcriptional regulation. In particular, the complexity of the human proteome is greatly expanded by the ~400 different types of protein posttranslational modifications (PTMs). The various PTM events, either alone or in combination (i.e., “cross-talk”), represent powerful mechanisms to modulate the function of a protein (e.g., activity, stability and localization), the collection of which convey information within the signaling network that underlies the complex traits in various pathophysiological conditions. However, because of many inherent technical difficulties associated with the analysis of protein PTMs (e.g., chemically diverse, unstable and low abundance), a complete description of the posttranslationally modified proteome of any given cells remains a daunting task. The overarching mission of our program is to: (1) develop cutting-edge quantitative proteomic approaches to systematically identify and characterize novel PTMs, (2) comprehensively interrogate the signaling events regulated by phosphorylation (mTOR pathways) and ADP-ribosylation (PARP pathways), and (3) combine these systems biology approaches with classical biochemical, cell biology and animal experiments to decipher the molecular underpinnings of cell growth and stress responses that are controlled by these two important pathways. To accomplish these goals, we will leverage our preliminary results (including a large set of unique hits, reagents and methods), and center our efforts on the following six goals. First, we will develop innovative mass spectrometric technologies with dramatically improved performance for global, quantitative and site-specific analysis of novel PTMs. Second, we will investigate the role of IGFBP5 (a recently identified extracellular target of mTORC1) as a mediator of the “non-cell autonomous” function of mTORC1. Third, we will determine the role of EGR1 (a novel hit identified from our previous MS screen) as a master regulator of the mTORC1-dependent feedback loops. Fourth, we will generate a tissue-specific atlas of mTORC1 phosphorylation substrates, and in doing so, interrogate non-uniform effects of this important pathway on the physiology of different tissues. Fifth, we will develop a large-scale MS approach to site-specific characterization of the D/E-mono-ADP-ribosylated proteome, and finally we will develop a large-scale method to measure absolute protein PARylation stoichiometries. Together, these studies provide a comprehensive framework for the MS identification and functional characterization of PTMs events linked to cell growth control and stress responses.

项目概要 疾病生物学的一个中心目标是描述负责转化的分子过程 细胞从正常状态转变为病理状态。与DNA和RNA的快速进展相比 测序技术，最终的、可以说是最可行的核心元素的表征 教条，蛋白质，已经落后了。细胞基因组与其蛋白质组之间的动态关系 人们对此知之甚少，反映了转录/转录后调控的多层。特别是， 约 400 种不同类型的翻译后蛋白质大大扩展了人类蛋白质组的复杂性 修饰（PTM）。各种 PTM 事件，无论是单独的还是组合的（即“串扰”），代表 调节蛋白质功能（例如活性、稳定性和定位）的强大机制， 其集合在信号网络内传递信息，这些信息是各种复杂特征的基础 病理生理条件。然而，由于许多固有的技术困难 蛋白质 PTM 分析（例如，化学多样性、不稳定和低丰度），完整的描述 任何给定细胞的翻译后修饰蛋白质组仍然是一项艰巨的任务。的总体使命 我们的计划是：（1）开发尖端的定量蛋白质组学方法来系统地识别和 表征新型 PTM，(2) 全面探究磷酸化调节的信号事件 （mTOR 途径）和 ADP-核糖基化（PARP 途径），以及 (3) 将这些系统生物学结合起来 采用经典生化、细胞生物学和动物实验的方法来破译分子 由这两个重要途径控制的细胞生长和应激反应的基础。到 为了实现这些目标，我们将利用我们的初步结果（包括大量独特的命中、试剂 和方法），并集中努力实现以下六个目标。一是发展创新大众 光谱分析技术显着提高了全局、定量和特定地点的性能 分析新的 PTM。其次，我们将研究 IGFBP5（最近发现的细胞外靶标）的作用 mTORC1）作为 mTORC1“非细胞自主”功能的介体。三、我们要确定角色 EGR1（我们之前的 MS 筛选中发现的一个新发现）作为 mTORC1 依赖的主调节因子 反馈循环。第四，我们将生成 mTORC1 磷酸化底物的组织特异性图谱，并在 这样做可以探究这一重要途径对不同组织生理学的非均匀影响。第五， 我们将开发一种大规模 MS 方法来对 D/E-mono-ADP-ribosylated 进行位点特异性表征 蛋白质组，最后我们将开发一种大规模的方法来测量绝对蛋白质PARylation 化学计量。这些研究共同为 MS 识别和识别提供了一个全面的框架。 与细胞生长控制和应激反应相关的 PTM 事件的功能表征。