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ROLE OF MICROTUBULE-BASED TRANSPORT IN NEURONAL POLARITY

基于微管的运输在神经元极性中的作用

基本信息

批准号：
8647011
负责人：
JILL C WILDONGER
金额：
$ 23.94万
依托单位：
UNIVERSITY OF WISCONSIN-MADISON
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-09-01 至 2015-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8647011
关键词：
Acetylation Address Affect Affinity Alleles Animal Model Award Axon Behavior Binding Biological Assay California Cell Polarity Complex Cues Cytoskeleton Defect Dendrites Development Distal Drosophila genus Dynein ATPase Engineering Gene Mutation Genes Genetic Genomics Goals Human In Vitro Kinesin Knock-in Mouse Lead Learning Link Mediating Mentors Microtubules Miller-Dieker Syndrome Modeling Modification Molecular Molecular Motors Morphogenesis Motor Motor Activity Mus Mutation Nervous system structure Neurites Neurodevelopmental Disorder Neuronal Migration Disorder Neurons Organelles Organism Patients Phase Post-Translational Protein Processing Process Proteins Reagent Research Personnel Role San Francisco Shapes Signal Transduction Site-Directed Mutagenesis Specific qualifier value Specificity Staging Structure System Techniques Testing Tubulin Universities Work Yang base cell motility cofactor developmental disease fly human disease in vivo insight migration neurodevelopment neuron development neuron loss novel research study single molecule tool

项目摘要

Shortly after differentiating, neurons establish distinct axonal and dendritic compartments that are specialized to send and receive signals, respectively. Polarity is essential for neurons to function in a neuronal circuit, yet how neurons polarize within a developing organism remains virtually unknown. My long-term goal as an independent biomedical researcher is to identify the mechanisms that create distinct axonal and dendritic compartments within neurons and to understand how this contributes to normal neuronal function in vivo. This proposal is based on our finding in fruit flies that the microtubule-based molecular motor dynein is necessary for two key features of neuronal polarity: the polarized localization of dendritic proteins and organelles and the uniform plus-end distal orientation of axonal microtubules. Two outstanding questions I will address in this proposal are: (1) How is dynein's function in neurons controlled by its interactions with different cofactors? and (2) How is dynein's activity regulated by its interaction with microtubules; more specifically, do microtubule modifications (such as acetylation, detyrosination, and polyglutamylation) provide spatial cues that influence dynein's activity and thereby shape neuronal polarity? The mentored phase of this award will be carried out at the University of California, San Francisco (UCSF), under the guidance of Dr. Yuh Nung Jan. During the mentored phase, I will use a genetic approach to characterize the cofactors that provide functional specificity to dynein developing fruit fly nervous system (Aim 1). Next, I will extend my studies in vitro and develop a new dynein motor construct to determine how dynein motor activity is affected by microtubule modifications (Aim 2). To do so, I will collaborate with Dr. Ronald Vale (UCSF) to learn in vitro techniques to analyze motor- microtubule interactions, including single molecule motility assays. To address how microtubule modifications affect neuronal polarization in vivo (Aim 3), I will use new knock-in technique called "genomic engineering" to build reagents for my independent phase. Dr. Yang Hong (University of Pittsburgh), who pioneered the genomic engineering technique, will serve as a consultant, as will Dr. Anthony Wynshaw-Boris (UCSF), a leader in the study of genes linked to human neurodevelopmental disorders such as classical lissencephaly. During the independent phase, I will address the following questions: Are microtubule modifications necessary for neurons to form distinct axonal and dendritic compartments in vivo? Is any one modification particularly important, or are there combinations of modifications that specify axon or dendrite formation? How do microtubule modifications regulate polarized transport in developing neurons in vivo? To answer these questions, I will use genomic engineering to knock-in multiple tubulin alleles with targeted mutations that block different microtubule modifications, both singly and in combination. Using currently available reagents and new polarity markers that I will generate, I will then characterize the effect of these mutations on neuronal polarity and dynein-mediated polarized transport within developing fruit fly nervous system. Through this combination of in vitro and in vivo approaches, these studies will provide significant new insight into microtubule-based mechanisms that shape neuronal polarity in a developing organism.

分化后不久，神经元就会形成不同的轴突和树突隔室