Molecular Mechanisms of Hair Bundle Development and Maintenance

发束发育和维护的分子机制

基本信息

批准号：
10643931
负责人：
Jonathan Edward Bird
金额：
$ 39.38万
依托单位：
UNIVERSITY OF FLORIDA
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-07-01 至 2025-06-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10643931
关键词：
ATP phosphohydrolase ATPase Domain Acceleration Actins Affect Animals Architecture Auditory Binding Biochemical Biological Assay C-terminal Cells Clustered Regularly Interspaced Short Palindromic Repeats Cochlea Complex Critical Pathways Cryoelectron Microscopy Cytoskeleton Data Defect Detection Development Ear Electron Microscopy Epidermal Growth Factor Receptor Pathway Substrate 8 Filament Fluorescence Microscopy Foundations GNAI3 gene GPSM2 gene Genes Genetic Genetic Engineering Goals Growth Hair Hair Cells Hearing Heart Height Human In Vitro Individual Inherited Laboratories Labyrinth Life Life Cycle Stages Longevity Maintenance Measures Mechanics Microfilaments Microscopy Molecular Molecular Motors Motor Mus Mutant Strains Mice Mutation Myosin ATPase N-terminal Organelles Outcome Pathologic Pathology Pathway interactions Patients Phase Polymers Process Property Protein Isoforms Proteins Pyrenes Resolution Sensory Spectrum Analysis Stereocilium Structure Tail Testing WHRN gene Work deafness disability experimental study hearing impairment hereditary hearing loss in vitro activity in vivo light microscopy mechanotransduction monomer mouse model mutant novel particle polymerization postnatal preservation protein complex protein purification single molecule sound synergism trafficking

项目摘要

Project Summary The detection of sound in the cochlea requires hair cells and their mechano-sensitive organelles, called stereocilia. The long-term goal of this laboratory is to study how stereocilia grow and how their integrity is maintained over a lifetime. These are critical processes and are commonly disrupted in hereditary forms of human hearing loss. In this proposal, we investigate a molecular motor called myosin 15 (MYO15A) that sets the size of the actin filament core that is the structural foundation within each stereocilium. Mutations in the MYO15A gene cause human hereditary hearing loss, DFNB3. Our initial experiments have revealed a novel mechanism that allows MYO15A to control the actin core, and we hypothesize that the hair cell regulates stereocilia architecture using different MYO15A isoforms. To test this, we will investigate the molecular properties of MYO15A to understand how it influences growth of the actin core, reveal how these activities are regulated within the hair cell, and examine how mutations cause hearing loss in a mouse model. In Aim 1, we use purified proteins and spectroscopy / single-molecule assays to extensively characterize how MYO15A accelerates actin polymerization. As part of this, we will introduce mutations to explore candidate regions within MYO15A that underlie this activity. In Aim 2, we expand our study to different isoforms of MYO15A and use biochemical assays and cryo-electron microscopy to investigate key differences in their enzymatic activity and how these are regulated. In Aim 3, we characterize a mutant mouse where a novel MYO15A isoform has been removed using CRISPR genetic engineering, and study how these animals lose their hearing using a combination of high- resolution electron and light microscopy. Overall, our proposal will provide critical new information into basic mechanisms of stereocilia plasticity, in addition to revealing the distinct pathologies that cause deafness in patients suffering with DFNB3.

项目概要耳蜗中声音的检测需要毛细胞及其机械敏感细胞器，称为静纤毛。该实验室的长期目标是研究静纤毛如何生长以及其完整性如何维持一生。这些都是关键过程，通常会以遗传形式被破坏。人类听力损失。在本提案中，我们研究了一种名为肌球蛋白 15 (MYO15A) 的分子马达，它可以设置肌动蛋白丝核心的大小是每个静纤毛内的结构基础。突变在 MYO15A基因导致人类遗传性听力损失，DFNB3。我们最初的实验揭示了一种新颖的 MYO15A 控制肌动蛋白核心的机制，我们假设毛细胞调节使用不同 MYO15A 亚型的静纤毛结构。为了测试这一点，我们将研究分子特性 MYO15A 以了解它如何影响肌动蛋白核心的生长，揭示这些活动是如何调节的毛细胞内，并检查突变如何导致小鼠模型听力损失。在目标 1 中，我们使用纯化的蛋白质和光谱学/单分子检测广泛表征 MYO15A 如何加速肌动蛋白聚合。作为其中的一部分，我们将引入突变来探索 MYO15A 内的候选区域是这项活动的基础。在目标 2 中，我们将研究扩展到 MYO15A 的不同亚型并使用生化检测和冷冻电子显微镜来研究它们酶活性的关键差异以及这些差异是如何发生的受监管。在目标 3 中，我们描述了一种突变小鼠，其中一种新的 MYO15A 亚型已被使用去除 CRISPR 基因工程，并通过结合高通量研究这些动物如何失去听力分辨率电子和光学显微镜。总体而言，我们的提案将为基本信息提供重要的新信息静纤毛可塑性机制，除了揭示导致耳聋的独特病理学之外患有 DFNB3 的患者。

项目成果

期刊论文数量（4）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

The actin cytoskeleton in hair bundle development and hearing loss.

发束发育和听力损失中的肌动蛋白细胞骨架。

DOI：
发表时间：
2023-09-01
期刊：
影响因子：
2.8
作者：
Park, Jinho;Bird, Jonathan E
通讯作者：
Bird, Jonathan E

Large-scale annotated dataset for cochlear hair cell detection and classification.

用于耳蜗毛细胞检测和分类的大规模注释数据集。

DOI：
发表时间：
2024-04-23
期刊：
影响因子：
9.8
作者：
Buswinka, Christopher J;Rosenberg, David B;Simikyan, Rubina G;Osgood, Richard T;Fernandez, Katharine;Nitta, Hidetomi;Hayashi, Yushi;Liberman, Leslie W;Nguyen, Emily;Yildiz, Erdem;Kim, Jinkyung;Jarysta, Amandine;Renauld, Justine;Wesson, Ella
通讯作者：
Wesson, Ella

The ATPase mechanism of myosin 15, the molecular motor mutated in DFNB3 human deafness.

肌球蛋白 15 的 ATP 酶机制，分子马达在 DFNB3 人类耳聋中发生突变。

DOI：
发表时间：
2021
期刊：
影响因子：
0
作者：
Jiang, Fangfang;Takagi, Yasuharu;Shams, Arik;Heissler, Sarah M;Friedman, Thomas B;Sellers, James R;Bird, Jonathan E
通讯作者：
Bird, Jonathan E

Myosin motors in sensory hair bundle assembly.

感觉毛束组件中的肌球蛋白马达。