New Proteomic Technologies for the Analysis of Tyrosine Kinase Signaling Pathways

用于分析酪氨酸激酶信号通路的蛋白质组学新技术

• 批准号: 8136442
• 负责人: W. Andy Tao
• 金额: $ 29.77万
• 依托单位: PURDUE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-09-01 至 2015-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8136442
• 关键词: Affinity; Biochemical; Biological Models; Breast Cancer Cell; Cancer Biology; Cell Proliferation; Cells; Characteristics; Chemicals; Clinical Data; Complex Mixtures; Coupled; Data; Development; Disease; Event; Goals; Growth; Immune; Individual; Knowledge; Lead; Link; Malignant Neoplasms; Maps; Mass Spectrum Analysis; Medicine; Methodology; Methods; Modeling; Modification; Molecular; Monitor; Neoplasm Metastasis; Outcome; Pathway interactions; Phosphopeptides; Phosphoproteins; Phosphorylation; Phosphorylation Site; Phosphotransferases; Physiological; Play; Polymers; Property; Protein Tyrosine Kinase; Proteins; Proteomics; Protocols documentation; Research; Research Personnel; Role; Series; Signal Pathway; Signal Transduction; Site; Squamous cell carcinoma; System; Techniques; Technology; Testing; Therapeutic; Tumor Suppressor Proteins; Tyrosine Phosphorylation; Tyrosine Phosphorylation Site; base; cancer cell; cancer prevention; cancer type; cell growth; cell growth regulation; cell motility; cell type; design; human SYK protein; improved; in vivo; innovation; interest; leukemia/lymphoma; malignant breast neoplasm; malignant phenotype; melanoma; neoplastic cell; novel; outcome forecast; overexpression; pre-clinical; public health relevance; response; tool

DESCRIPTION (provided by applicant): A key objective of phosphoproteomic analyses is to obtain an in-depth understanding of protein phosphorylation and signaling networks in cells. However, despite the great progress that has been made over the past few years, our ability to analyze such signaling pathways is still greatly limited by a number of serious technical challenges. The long term goal is to develop novel proteomics strategies valuable for understanding on the molecular level the role of phosphorylation in regulating cell proliferation. The objective in this application is to devise and optimize effective and efficient methodologies and strategies for the analyses of tyrosine kinase phosphoproteomes, using a dual functional tyrosine kinase Syk as the model system. This approach will utilize functional, soluble polymers for the efficient and inclusive isolation of phosphopeptides coupled with an integrated chemical, mass spectrometric, and computational strategy for the identification of specific tyrosine kinase substrates and for quantitative phosphoproteomics. We will focus on Syk-dependent signaling and establish protocols that will ultimately provide a powerful method to dissect any signaling pathway regulated by protein-tyrosine phosphorylation. Guided by strong preliminary data, this objective will be achieved by pursuing three specific aims: 1) Develop a proteomic platform for the analysis of protein phosphorylation. We will primarily develop, characterize and optimize functionalized soluble polymers designed for the affinity and chemical isolation of phosphopeptides; 2) Identification of specific tyrosine-kinase substrates and their phosphorylation sites; and 3) In-depth analyses of tyrosine kinase phosphoproteomes in several cancer types. We will develop and apply our quantitative proteomic technologies for the identification and quantification of tyrosine-phosphorylation sites in several cancer types. This project is innovative because it will develop a series of novel analytical techniques to directly identify novel tyrosine kinase substrates and phosphorylation sites that will provide investigators with a significantly expanded capability to dissect signaling pathways. The developed techniques will provide detailed knowledge at the molecular level on precisely how tyrosine kinases are involved in cell signaling under different physiological conditions through dynamic changes in protein phosphorylation. It is also expected that, by identifying a kinase's direct substrates, the proposed research will provide a powerful tool to conveniently map kinase networks. PUBLIC HEALTH RELEVANCE: The proposed studies to develop a series of analytical techniques have potential applicability for investigators in the molecular signaling field. The tools will allow them to examine important roles of tyrosine kinase in multiple signaling pathways. The proposed research will provide unique strategies and materials that will strengthen the important contribution that proteomics can make to our understanding of cancer biology.

描述（由申请人提供）：磷酸化蛋白质组学分析的一个关键目标是深入了解细胞中的蛋白质磷酸化和信号网络。然而，尽管过去几年取得了巨大进步，但我们分析此类信号通路的能力仍然受到许多严峻的技术挑战的极大限制。长期目标是开发新的蛋白质组学策略，有助于在分子水平上理解磷酸化在调节细胞增殖中的作用。本申请的目的是使用双功能酪氨酸激酶 Syk 作为模型系统，设计和优化用于分析酪氨酸激酶磷酸化蛋白质组的有效且高效的方法和策略。该方法将利用功能性可溶性聚合物来有效且包容性地分离磷酸肽，并结合集成的化学、质谱和计算策略来识别特定的酪氨酸激酶底物和定量磷酸化蛋白质组学。我们将重点关注 Syk 依赖性信号传导并建立最终提供强大方法来剖析蛋白质酪氨酸磷酸化调节的任何信号传导途径的方案。在强有力的初步数据的指导下，这一目标将通过追求三个具体目标来实现：1）开发用于分析蛋白质磷酸化的蛋白质组学平台。我们将主要开发、表征和优化专为磷酸肽的亲和力和化学分离而设计​​的功能化可溶性聚合物； 2）特定酪氨酸激酶底物及其磷酸化位点的鉴定； 3) 深入分析几种癌症类型中的酪氨酸激酶磷酸化蛋白质组。我们将开发并应用我们的定量蛋白质组技术来识别和定量几种癌症类型中的酪氨酸磷酸化位点。该项目具有创新性，因为它将开发一系列新颖的分析技术来直接识别新颖的酪氨酸激酶底物和磷酸化位点，这将为研究人员提供显着扩展的剖析信号通路的能力。所开发的技术将在分子水平上提供关于酪氨酸激酶如何通过蛋白质磷酸化的动态变化在不同生理条件下精确参与细胞信号传导的详细知识。还预计，通过识别激酶的直接底物，拟议的研究将为方便地绘制激酶网络提供强大的工具。 公共卫生相关性：拟议的开发一系列分析技术的研究对于分子信号传导领域的研究人员具有潜在的适用性。这些工具将使他们能够检查酪氨酸激酶在多种信号传导途径中的重要作用。拟议的研究将提供独特的策略和材料，以加强蛋白质组学对我们对癌症生物学的理解做出的重要贡献。