Structure and function in Notch Signaling

Notch 信号传导的结构和功能

基本信息

批准号：
8927540
负责人：
Stephen C. Blacklow
金额：
$ 40.24万
依托单位：
HARVARD MEDICAL SCHOOL
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-09-16 至 2019-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8927540
关键词：
Acute Acute T Cell Leukemia Address Adult Award Binding Biochemical Biological Assay Blocking Antibodies CADASIL Cardiovascular Diseases Cell Nucleus Cell surface Cells Cleaved cell Complex Credentialing DNA Defect Development Disease Enzymes Event Family Foundations Functional disorder Gene Targeting Goals Health Homeostasis Human Human Biology Human Pathology Immune response Individual Inherited Integral Membrane Protein Knowledge Ligand Binding Ligands Malignant Neoplasms Membrane Metalloproteases Molecular Mutation Nerve Degeneration Neurodegenerative Disorders Notch Signaling Pathway Nucleic Acid Regulatory Sequences Oncogenic Organism Peptide Hydrolases Play Process Protease Domain Protein Isoforms Proteolysis Receptor Activation Recurrence Regulation Research Project Grants Resistance Roentgen Rays Role Signal Pathway Signal Transduction Site Specificity Stroke Structure Syndrome T-Lymphocyte Therapeutic Tissues Transcriptional Activation Work angiogenesis base cell growth developmental disease fly gamma secretase human disease leukemia leukemia/lymphoma loss of function mutation malignant breast neoplasm neoplastic cell notch protein novel therapeutics premature receptor receptor binding response therapeutic target tumor

项目摘要

DESCRIPTION (provided by applicant): The overarching goal of this research project is to elucidate at a structural and mechanistic level critical features of both normal and pathogenic Notch signal transduction. Prior to activation, Notch is quiescent and resistant to proteolysis, bu ligand binding triggers the "on" state by stimulating proteolytic cleavage of Notch by the ADAM10 (or ADAM17) metalloprotease at a juxtamembrane site immediately external to the membrane. The work proposed here will address key unresolved questions in the Notch field, including: i) how Notch receptors specifically recognize their ligands, and ii) how ADAM10 proteolysis of Notch is achieved and regulated. We propose to address these key questions in signaling by pursuing the following specific aims: 1. Determine the basis for specific binding between Notch receptors and their canonical ligands. In this aim, we will combine structural, biochemical, and cell-based assays to address one of the central unresolved questions in Notch signal transduction: how do Notch receptors recognize their ligands? The top priority will be to solve an X-ray structure of a receptor-ligand complex. I the process of pursuing this highest-impact, long-term objective, we will determine the intrinsic selectivity of various Notch receptors for different Delta-like ligands, as well as determine structures of individual receptor and ligand fragments in isolation and/or in combination with blocking antibodies of potential therapeutic relevance. 2. Determine the basis for catalytic specificity of ADAM-family metalloproteases in ligand- dependent proteolysis of Notch receptors. Top priorities of this aim will be to identify the processing sites of the different Notch isoforms, to elucidate the structure of the full ADAM10 ectodomain, and to determine the role of the ADAM regulatory region on Notch proteolysis by comparing the enzymatic activities of the isolated protease domain and the full ectodomain on Notch-derived substrates. Successful completion of these aims will constitute a major breakthrough in our understanding of this fundamental signaling pathway and the role it plays during normal development and in the pathophysiology of diseases associated with aberrant Notch signaling. By deepening our knowledge about the structural and biochemical foundations underlying ligand engagement and regulated proteolysis, our studies will identify new therapeutic strategies for management of Notch-related human pathologies, which include developmental disorders, neurodegenerative diseases, cardiovascular diseases, and cancer.

描述（由申请人提供）：该研究项目的总体目标是在结构和机制水平上阐明正常和致病性Notch信号转导的关键特征。在激活之前，Notch 处于静止状态并且对蛋白水解有抵抗力，bu 配体结合通过在紧邻膜外部的近膜位点刺激 ADAM10（或 ADAM17）金属蛋白酶对 Notch 进行蛋白水解切割，从而触发“开启”状态。这里提出的工作将解决 Notch 领域中未解决的关键问题，包括：i) Notch 受体如何特异性识别其配体，以及 ii) 如何实现和调节 Notch 的 ADAM10 蛋白水解。我们建议通过追求以下具体目标来解决信令中的这些关键问题： 1. 确定Notch受体与其典型配体之间特异性结合的基础。为此，我们将结合结构、生化和细胞检测来解决 Notch 信号转导中未解决的核心问题之一：Notch 受体如何识别其配体？首要任务是解析受体-配体复合物的 X 射线结构。在追求这一最具影响力的长期目标的过程中，我们将确定各种Notch受体对不同Delta样配体的内在选择性，并确定单独和/或组合的单个受体和配体片段的结构具有潜在治疗相关性的阻断抗体。 2. 确定 ADAM 家族金属蛋白酶在配体依赖性 Notch 受体蛋白水解中的催化特异性的基础。该目标的首要任务是确定不同 Notch 亚型的加工位点，阐明完整 ADAM10 胞外域的结构，并通过比较分离的蛋白酶的酶活性来确定 ADAM 调节区在 Notch 蛋白水解中的作用Notch 衍生底物上的结构域和完整胞外域。成功完成这些目标将是我们对这一基本信号通路及其在正常发育和与异常Notch信号相关疾病的病理生理学中所发挥的作用的理解的重大突破。通过加深我们对配体结合和调节蛋白水解的结构和生化基础的了解，我们的研究将确定管理Notch相关人类病理的新治疗策略，其中包括发育障碍、神经退行性疾病、心血管疾病和癌症。