Role of XIRP2 in hair cell function and degeneration

XIRP2 在毛细胞功能和退化中的作用

基本信息

批准号：
8957404
负责人：
Jung-Bum Shin
金额：
$ 33.6万
依托单位：
UNIVERSITY OF VIRGINIA
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：
2014-12-01 至 2019-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8957404
关键词：
Actinin Actins Address Adherens Junction Age Binding Cell Maintenance Cell physiology Cellular Structures Clustered Regularly Interspaced Short Palindromic Repeats Complement Complex Coupling Defect Development Environmental Risk Factor Epithelial Exhibits Exons F-Actin Genes Genetic Goals Hair Hair Cells Health Hearing Heart High-Frequency Hearing Loss Human Intercellular Junctions Knockout Mice Labyrinth Light Location Maintenance Mass Spectrum Analysis Mechanical Stress Mechanics Mediating Membrane Microfilaments Mus Mutation Myocardium Neoplasms Noise Noise-Induced Hearing Loss Overlapping Genes Pathway interactions Peripheral Phenotype Play Protein Isoforms Proteins Proteome Proteomics RNA Splicing Receptor Cell Regulation Role Sensory Hair Sequence Homology Side Stereocilium Stress Tests Structural defect Structure Supporting Cell Technology Tertiary Protein Structure Testing Time Transgenic Mice base deafness depolymerization hearing impairment novel null mutation prevent research study

项目摘要

DESCRIPTION (provided by applicant): Our long-term goal is to understand the mechanisms that underlie the development, maintenance and degeneration of the mechanosensory hair bundle. Motivated by the fact that a third of all known deafness genes encode hair bundle proteins, we previously used a mass spectrometry based strategy to characterize the hair bundle proteome. This approach proved to be a powerful complement to traditional genetics strategies, leading to the discovery of novel deafness genes previously undetected using genetic strategies alone. For further analysis, we focused on proteins for which the corresponding gene locations overlap with unresolved human deafness loci. One such protein is XIRP2 (for xin actin-binding repeat containing protein 2). In this project, we are testing the hypothesis that XIRP2 is a novel hair cell protein required for long-term stability of stereocilia and the hair cell/supporting cell junctions, with the consequence that XIRP2 deficiency causes hair cell degeneration and progressive hearing loss. Preliminary studies demonstrated that different XIRP2 splice forms are present in the hair cell, displaying distinct localizations in the stereocilia and pericuticular adherens junctions, respectively. To explore the role of XIRP2 for hearing function, we used the CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats)/Cas technology to generate transgenic mice with a functional null mutation in the Xirp2 gene and found evidence for high-frequency hearing loss. In Specific Aim 1, we will investigate the hearing loss phenotype in Xirp2 null mice in detail. Furthermore, we will test whether XIRP2 deficiency renders the hair cell more sensitive to mechanical stress, by testing the sensitivity of Xirp2 null mice to noise-induced hearing loss. Preliminary studies suggested that the stereocilia and the adherens junctions harbor distinct XIRP2 isoforms, implying that defects in either hair cell structure could underlie the observed hearing loss in Xirp2 null mice. n Aim 2 and 3, we will therefore use transgenic mice with isoform specific deletions in the Xirp2 gene, to specifically address their contribution to hair cell degeneration and hearing loss. Our hypothesis that XIRP2 is required for long-term maintenance of hair cell structures dovetails well with its known role in cardiac muscle, where it is involved in the maintenance of the sarcomeric Z-line. XIRP2's role in the hair cell is likely to be distinct from and more complex than its functon in the heart: the hair bundle harbors a novel isoform with vastly different protein domain structure, rendering it practically an unknown protein. Significant for human hearing health, understanding the role of XIRP2 in hair cell degeneration is expected to shed light on mechanisms by which subtle defects caused by genetic and environmental factors can compromise the structural integrity of hair cell structures, significant for understanding the mechanistic basis of age and noise-induced hearing loss. Finally, the chromosomal locus of the XIRP2 gene overlaps with the human deafness loci DFNB27 and DFNA16, opening up the prospect of identifying the causative mutation for two human deafness phenotypes.

描述（由申请人提供）：我们的长期目标是了解机械感觉毛束发育、维持和退化的机制。由于所有已知耳聋基因中有三分之一编码发束蛋白，我们之前使用基于质谱的策略来表征发束蛋白质组。事实证明，这种方法是对传统遗传学策略的有力补充，从而发现了以前仅使用遗传策略无法检测到的新型耳聋基因。为了进一步分析，我们重点关注相应基因位置与未解决的人类耳聋基因座重叠的蛋白质。其中一种蛋白质是 XIRP2（代表含有 xin 肌动蛋白结合重复序列的蛋白质 2）。在这个项目中，我们正在测试这样的假设：XIRP2 是静纤毛和毛细胞/支持细胞连接长期稳定性所需的一种新型毛细胞蛋白，因此 XIRP2 缺陷会导致毛细胞退化和进行性听力损失。初步研究表明，毛细胞中存在不同的 XIRP2 剪接形式，在毛细胞中表现出不同的定位。分别是静纤毛和角质周粘附连接。为了探索 XIRP2 对听力功能的作用，我们使用 CRISPR（成簇规则间隔短回文重复序列）/Cas 技术来产生 Xirp2 基因功能性无效突变的转基因小鼠，并发现了高频听力损失的证据。在具体目标 1 中，我们将详细研究 Xirp2 缺失小鼠的听力损失表型。此外，我们将通过测试 Xirp2 缺失小鼠对噪声引起的听力损失的敏感性来测试 XIRP2 缺陷是否会使毛细胞对机械应力更加敏感。初步研究表明，静纤毛和粘附连接具有不同的 XIRP2 同工型，这意味着任一毛细胞结构的缺陷可能是 Xirp2 缺失小鼠中观察到的听力损失的基础。因此，在目标 2 和 3 中，我们将使用 Xirp2 基因中具有亚型特异性缺失的转基因小鼠，以专门解决它们对毛细胞变性和听力损失的影响。我们的假设是，XIRP2 是长期维持毛细胞结构所必需的，这与其在心肌中的已知作用非常吻合，在心肌中，XIRP2 参与维持肌节 Z 线。 XIRP2 在毛细胞中的作用可能与其在心脏中的功能不同，并且比其在心脏中的功能更复杂：毛束含有一种具有截然不同的蛋白质结构域结构的新型亚型，使其实际上成为一种未知的蛋白质。对人类听力健康具有重要意义，了解 XIRP2 在毛细胞变性中的作用有望揭示遗传和环境因素引起的细微缺陷可能损害毛细胞结构完整性的机制，这对于理解年龄的机制基础具有重要意义和噪音引起的听力损失。最后，XIRP2基因的染色体位点与人类耳聋位点DFNB27和DFNA16重叠，为鉴定两种人类耳聋表型的致病突变开辟了前景。