Microtubule-mediated mechanisms underlying hair cell development and deafness

毛细胞发育和耳聋的微管介导机制

• 批准号: 9232136
• 负责人: Xiaowei Lu
• 金额: $ 33.29万
• 依托单位: UNIVERSITY OF VIRGINIA
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-03-01 至 2019-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9232136
• 关键词: Actins; Affect; Age; Americas; Apical; Auditory; Behavior; Biological Assay; Cell Polarity; Cell division; Cochlear duct; DNA Sequence Alteration; Data; Defect; Development; Disease; Dominant-Negative Mutation; Dynein ATPase; Ear; Economic Burden; Feedback; GPSM2 gene; Genes; Genetic; Goals; Guanosine Triphosphate Phosphohydrolases; Hair; Hair Cells; Hearing; Human; Image; In Situ; Inherited; Injury; Investigation; KRP protein; Kinocilium; Knowledge; Lateral; Ligation; Light; Mechanics; Mediating; Microtubule Stabilization; Microtubules; Mitotic spindle; Modeling; Molecular; Molecular Motors; Morphogenesis; Motor; Mutant Strains Mice; Mutation; Names; National Institute on Deafness and Other Communication Disorders; Natural regeneration; Organ of Corti; Organelles; Pathway interactions; Pattern; Plus End of the Microtubule; Positioning Attribute; Proteins; Recruitment Activity; Regulation; Research; Role; Sensorineural Hearing Loss; Shapes; Signal Transduction; Stimulus; Structure; Surface; Syndrome; Testing; Tissues; Work; base; body position; cell cortex; deafness; design; disability; gain of function; hearing impairment; innovation; insight; interest; kinetosome; mechanotransduction; novel; programs; protease-activated receptor 3; public health relevance; repaired; sensor; social; sound; stem cell technology; targeted delivery

DESCRIPTION (provided by applicant): The actin-based stereociliary bundle (or hair bundle) on the apex of auditory hair cells serves the critical function of converting sound energy to electric signals. Its V-shaped staircase structure renders the bundle directionally sensitive to mechanical stimuli. As such, auditory hair bundles must be uniformly oriented for correct sound transduction. Abnormalities in hair bundle polarity or orientation cause deafness and hearing impairment. A long-term objective of this work is to gain a detailed understanding of the hair bundle morphogenesis programs and how genetic mutations that disrupt these programs cause sensorineural deafness. In particular, mutations in a gene named GPSM2 cause the human hereditary deafness DFNB82 and Chudley-McCullough Syndrome. However, the underlying disease mechanisms remain completely unknown. Our recent insights about a microtubule-mediated pathway in hair cells for hair bundle polarity and orientation suggest a novel testable hypothesis about GPSM2's role in this pathway. Specifically, we have uncovered a critical and previously unappreciated function of the hair cell microtubules and microtubule-based molecular motors in basal body positioning, which is critical for both hair bundle polarity and orientation. We found that hair cells deficient in either the kinesin-II subunit Kif3a or the dynein regulator Lis1 have basal body positioning defects. Consequently, both the polarized V-shape and orientation of the hair bundles are disrupted. We further demonstrate that these microtubule motors regulate an asymmetric domain of Rac GTPase-PAK signaling on the hair cell cortex to mediate basal body positioning. The major goal of this research is to further dissect the molecular components of this microtubule-mediated pathway, including the deafness gene GPSM2, and gain mechanistic insights into microtubule regulation of hair bundle polarity. Our goal will be pursued through the following specific aims. Aim 1 will test the hypothesis that the cell polarity proteins Par 3 and GPSM2 serve as cortical landmarks to tether dynein at the cortex to pull on microtubules and orient the basal body, similar to mechanisms that orient the mitotic spindle during asymmetric cell division. Aim 2 will test the hypothesis that kinesin-II mediated targeted delivery of Par3 and the Rac activator Tiam1 to the cell cortex is critical for spatial regulation of Rac signaling and basal body positioning. Aim 3 will use innovative live imaging to test the hypothesis that PAK signaling regulates both microtubule stability and cortical proteins to stabilize microtubule plus-end attachment at the cell cortex. This research will provide new avenues of investigation into hair cell development and elucidate the function of poorly understood human deafness genes. Gaining a deeper understanding of the hair bundle morphogenesis program will be essential for devising rational therapies to stimulate hair bundle repair following injury, to treat hereditary human deafness and to regenerate auditory hair cells through stem cell technologies.

描述(申请人提供)：听觉毛细胞顶端基于肌动蛋白的立体纤毛束(或毛束)起着将声能转化为电信号的关键功能。它的V形楼梯结构使束状物对机械刺激具有方向性的敏感性。因此，听觉发束必须均匀定向，才能正确地进行声音传导。发束、极性或方向的异常会导致耳聋和听力障碍。这项工作的一个长期目标是详细了解毛束形态发生程序，以及破坏这些程序的基因突变如何导致感音神经性耳聋。特别是，名为GPSM2的基因突变会导致人类遗传性耳聋DFNB82和Chudley-McCullough综合征。然而，潜在的疾病机制仍然完全未知。我们最近对毛细胞中微管介导的毛束极性和方向的通路的洞察提出了一个新的可检验的假说，即GPSM2的S在这一通路中扮演了角色。具体地说，我们发现了毛细胞微管和基于微管的分子马达在基础身体定位中的一个关键和以前未被认识的功能，这对毛束的极性和方向都是关键的。我们发现，缺乏Kinesin-II亚单位Kif3a或动力蛋白调节因子Lis1的毛细胞存在基础身体定位缺陷。结果，发束的偏振V形和取向都被破坏了。我们进一步证明，这些微管马达调节毛细胞皮质上Rac GTPase-PAK信号的不对称区域，以调节基底定位。这项研究的主要目的是进一步剖析这一微管介导的通路的分子组成，包括耳聋基因GPSM2，并从机制上深入了解微管对毛束极性的调节。我们的目标将通过以下具体目标来实现。目的1验证细胞极性蛋白PAR 3和GPSM2作为皮质标志物的假设，将dynein拴在皮质上以拉出微管并定位基底体，类似于在细胞不对称分裂期间定位有丝分裂纺锤体的机制。目的2将验证一个假设，即Kinesin-II介导的Par3和RAC激活剂Tiam1的靶向传递到细胞皮质对RAC信号的空间调节和基础体位至关重要。Aim 3将使用创新的实时成像来测试这一假设，即PAK信号调节微管稳定性和皮质蛋白以稳定细胞皮质的微管正端附着。这项研究将为毛细胞发育提供新的研究途径，并阐明知之甚少的人类耳聋基因的功能。深入了解毛束形态发生程序对于设计合理的治疗方法以刺激损伤后的毛束修复、治疗遗传性耳聋和通过干细胞技术再生听觉毛细胞将是至关重要的。