Role of Staircase Hair Bundle Morphology in Auditory Mechanotransduction

阶梯毛束形态在听觉机械传导中的作用

• 批准号: 8197185
• 负责人: Gregory I Frolenkov
• 金额: $ 31.59万
• 依托单位: UNIVERSITY OF KENTUCKY
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-12-01 至 2013-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8197185
• 关键词: Affect; Animal Model; Auditory; Cell physiology; Cells; Complex; Data; Development; Disease; Goals; Growth; Hair; Hair Cells; Human; Inherited; Inner Hair Cells; Kinetics; Labyrinth; Lateral; Lead; Link; Mediating; Modification; Molecular; Morphology; Motor; Mus; Mutation; Myosin ATPase; Outer Hair Cells; Point Mutation; Protein Isoforms; Proteins; Public Health; Recovery; Research; Role; Scaffolding Protein; Scanning Electron Microscopy; Scientist; Sensory; Stereocilium; Structure; Technology; Testing; Utricle structure; base; congenital deafness; deafness; human CDH23 protein; human PTPRT protein; insight; postnatal; prevent; repaired; research study; response; restoration

DESCRIPTION (provided by applicant): While number of proteins responsible for structural integrity of the mechanosensory bundle of the inner ear hair cells has been identified, how most of these proteins contribute to mechano-electrical transduction is still unknown. Also unknown is whether the lack or modification of MET contributes to the development of deafness and/or vestibular disorders that result from mutations of hair bundle proteins. The goal of the current project is to determine the role of the myosin XVa-based stereocilia elongation complex in mechanotransduction. In homozygous shaker 2 mice (Myo15sh2/sh2), a recessive point mutation in the motor domain of myosin XVa prevents normal localization of this protein to the tips of stereocilia, resulting in abnormally short stereocilia. According to our preliminary data, cochlear outer hair cells of young postnatal Myo15sh2/sh2 mice possess numerous obliquely oriented "tip links" and apparently normal mechanotransduction. In contrast, Myo15sh2/sh2 inner hair cells have equally short stereocilia without any obliquely oriented tip links, but with numerous "top-to-top" links perpendicular to the core of stereocilia. In spite of their abnormal morphology, Myo15sh2/sh2 inner hair cells have a prominent transduction current with "wild type" nanoampere-scale amplitude but abnormal directional sensitivity and no rapid Ca2+dependent deactivation, known as "fast adaptation". The central hypothesis of the proposal is that the myosin XVa- based stereocilia elongation complex is not required for mechanosensitivity of hair cells but may affect directional sensitivity and adaptation of mechanotransduction. This study will determine: 1) the role of myosin XVa in directional sensitivity of the hair bundle; 2) localization of known molecular components of stereocilia links in the myosin XVa-deficient hair bundles; 3) the role of myosin XVa and its molecular partner, whirlin in the fast adaptation of the transduction current. This study represents a step toward my long-term goal to understand how the hair cells acquire and maintain mechanosensitivity in normal and pathological conditions. Apart from being important for understanding the basic mechanisms of hair cell mechanotransduction, this study will establish how developmental abnormalities of stereocilia growth may affect the transduction machinery. This project will also be the first to study mechanotransduction during restoration of the hair bundle morphology and/or tip links in mammalian cochlear hair cells. Finally, our study will provide a wealth of data on hair cell function in shaker 2 and whirler mice, the animal models for DFNB3 and DFNB31 hereditary deafness in humans. This research is relevant to public health because it investigates exactly how sensory cells of the inner ear acquire and maintain mechanosensitivity in normal and pathological conditions. Our experimental results will help scientists to better understand, prevent, and develop treatments for developmental abnormalities in the sensory cells of the inner ear, which lead to congenital deafness.

描述（由申请人提供）：虽然已经鉴定出负责内耳毛细胞机械感应束的结构完整性的蛋白质数量，但这些蛋白质中的大多数如何促进机械电气转导的大多数仍然未知。也未知的是，MET的缺乏或修饰是否有助于由毛发蛋白突变引起的耳聋和/或前庭疾病的发展。当前项目的目的是确定基于肌球蛋白XVA的立体伸长率在机械传输中的作用。在纯合子振荡器2小鼠（MyO15SH2/SH2）中，肌球蛋白XVA的运动结构域中的隐性点突变可阻止该蛋白正常定位于立体胶质的尖端，从而导致异常短的立体切开素。根据我们的初步数据，年轻的产后Myo15SH2/SH2小鼠的人工耳蜗外毛细胞具有许多倾斜的“尖端链路”，并且显然是正常的机械转导。相比之下，Myo15SH2/SH2内毛细胞具有同样短的立体胶质，而没有任何倾斜的尖端连接，但具有垂直于立体胶体核心的许多“自上而下”链接。尽管它们异常形态，但MyO15SH2/SH2内毛细胞具有突出的转导电流，其“野生型”纳米接管尺寸幅度幅度但异常的定向灵敏度，并且没有快速的Ca2+依赖性依赖性失活，称为“快速适应”。该提案的中心假设是，基于肌球蛋白XVA的立体伸长延伸复合物不需要毛细胞的机械敏感性，而是可能影响机械传导的定向灵敏度和适应性。这项研究将确定：1）肌球蛋白XVA在头发束的定向灵敏度中的作用； 2）在肌球蛋白XVA缺陷束中的立体胶质连接的已知分子成分的定位； 3）肌球蛋白XVA及其分子伴侣Whirlin在转导电流的快速适应中的作用。这项研究代表了我朝着我的长期目标迈出的一步，以了解毛细胞如何在正常和病理条件下如何获得和维持机械敏感性。除了了解毛细胞机械转移的基本机制外，本研究还将确定立体胶体生长的发育异常如何影响转导机械。该项目还将是第一个在恢复哺乳动物耳毛细胞中发束形态和/或尖端链路期间研究机械转导的项目。最后，我们的研究将提供有关Shaker 2和Whirler小鼠的毛细胞功能的大量数据，DFNB3的动物模型和DFNB31人类的遗传性耳聋。这项研究与公共卫生有关，因为它准确地研究了内耳的感觉细胞如何在正常和病理条件下获得机械敏感性。我们的实验结果将有助于科学家更好地理解，预防和发展内耳感觉细胞中发育异常的治疗方法，从而导致先天性耳聋。

项目成果

Nanoscale live-cell imaging using hopping probe ion conductance microscopy.

• DOI: 10.1038/nmeth.1306
• 发表时间: 2009-04
• 期刊: NATURE METHODS
• 影响因子: 48
• 作者: Novak, Pavel;Li, Chao;Shevchuk, Andrew I.;Stepanyan, Ruben;Caldwell, Matthew;Hughes, Simon;Smart, Trevor G.;Gorelik, Julia;Ostanin, Victor P.;Lab, Max J.;Moss, Guy W. J.;Frolenkov, Gregory I.;Klenerman, David;Korchev, Yuri E.
• 通讯作者: Korchev, Yuri E.

Fast adaptation and Ca2+ sensitivity of the mechanotransducer require myosin-XVa in inner but not outer cochlear hair cells.

• DOI: 10.1523/jneurosci.4566-08.2009
• 发表时间: 2009-04-01
• 期刊: The Journal of neuroscience : the official journal of the Society for Neuroscience
• 作者: Stepanyan R;Frolenkov GI
• 通讯作者: Frolenkov GI

Stereocilia Bundle Imaging with Nanoscale Resolution in Live Mammalian Auditory Hair Cells.

活体哺乳动物听觉毛细胞中的立体纤毛束成像具有纳米级分辨率。

• DOI: 10.3791/62104
• 发表时间: 2021
• 期刊: Journal of visualized experiments : JoVE
• 作者: Galeano-Naranjo,Carolina;Veléz-Ortega,ACatalina;Frolenkov,GregoryI
• 通讯作者: Frolenkov,GregoryI

Visualization of Live Cochlear Stereocilia at a Nanoscale Resolution Using Hopping Probe Ion Conductance Microscopy.

使用跳跃探针离子电导显微镜以纳米级分辨率可视化活体耳蜗立体纤毛。

• DOI: 10.1007/978-1-4939-3615-1_12
• 发表时间: 2016
• 期刊: Methods in molecular biology (Clifton, N.J.)
• 作者: Vélez-Ortega,ACatalina;Frolenkov,GregoryI
• 通讯作者: Frolenkov,GregoryI