Activating mutations in MEK: from molecules to morphologies

激活 MEK 突变：从分子到形态

基本信息

批准号：
9333420
负责人：
REBECCA D. BURDINE
金额：
$ 42.44万
依托单位：
PRINCETON UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2011
资助国家：
美国
起止时间：
2011-09-15 至 2019-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9333420
关键词：
Adult Affect Antineoplastic Agents Binding Biochemical Biological Biological Assay Birth Cardiac Myoblasts Cell Count Cultured Cells DNA Sequence Alteration Defect Development Drosophila genus Embryo Enzymes Event Experimental Models Face Germ-Line Mutation Goals Heart Heart Abnormalities Human Image Joints Kinetics Knock-in Light Literature MAP Kinase Gene MAPK Signaling Pathway Pathway Malignant Neoplasms Maps Mass Spectrum Analysis Modeling Monitor Morphogenesis Morphology Mus Mutate Mutation Neurocognitive Organ Pathway interactions Patients Pattern Phenotype Phosphorylation Predisposition Proteins Publishing Reaction Regulation Research Research Personnel Signal Transduction Testing Tissue imaging Tissues Tube Variant Work Zebrafish base cardiogenesis design experimental study extracellular genome sequencing imaging approach in vivo insight migration postnatal programs protein structure public health relevance quantitative imaging reconstitution three dimensional structure whole genome

项目摘要

DESCRIPTION (provided by applicant): Our work is designed to provide fundamental understanding of RASopathies, a large class of developmental abnormalities caused by the germline mutations in components of the RAS/MAPK pathway. Patients with these conditions display a broad spectrum of phenotypes, including heart defects, short stature, facial dysmorphisms, and neurocognitive delays. Whole genome sequencing of RASopathies provides new sequence variants in the well characterized components of the RAS pathway, but the functional consequences of these sequence variations is poorly understood. We will quantitatively characterize the functional and phenotypic consequences of the activating mutations in MEK, a core component of the RAS pathway. Focusing on the same group of mutations, we will first determine their effects on the regulation and enzymatic activity of MEK (Aim 1). This will be done using mass spectrometry-based kinetic assays with purified proteins. Next, we will quantify the consequences of activating mutations for the kinetics of RAS signaling in vivo (Aim 2). This will be done using a quantitative imaging approach, with which we will monitor RAS signaling in Drosophila embryos. Finally, we will determine how the same mutations influence RAS-dependent tissue morphogenesis (Aim 3). This will be done using live imaging experiments in zebrafish, focusing on heart development as a model of a morphogenetic event that is commonly affected in RASopathies. Our studies should provide mechanistic insights into the biochemical and morphogenetic effects of mutations identified in a large group of human developmental abnormalities and highlight the importance of using multiple models and quantitative approaches for answering a specific biological question.

描述（由申请人提供）：我们的工作旨在提供对 RASopathies 的基本了解，RASopathies 是由 RAS/MAPK 通路成分的种系突变引起的一大类发育异常，患有这些疾病的患者表现出广泛的表型，包括。心脏缺陷、身材矮小、面部畸形和神经认知延迟 RASopathies 的全基因组测序为 RAS 通路的明确组成部分提供了新的序列变异，但这些序列变异的功能后果是我们将定量描述 MEK（RAS 途径的核心组成部分）激活突变的功能和表型后果，我们将首先确定它们对 MEK 调节和酶活性的影响。目标 1)。接下来，我们将量化激活突变对体内 RAS 信号动力学的影响（目标 2）。我们将使用定量成像方法来监测果蝇胚胎中的 RAS 信号传导，最后，我们将确定相同的突变如何影响 RAS 依赖性组织形态发生（目标 3）。，重点关注心脏发育作为 RASopathies 中常见影响的形态发生事件的模型，我们的研究应该为在一大群人类发育中发现的突变的生化和形态发生效应提供机制见解。异常并强调使用多种模型和定量方法来回答特定生物学问题的重要性。