A small calcium-binding protein may be key to stabilizing the sensory hair cell stereocilia Elongation Complex

项目总结/摘要 我们的听觉能力依赖于内耳中一小群专门的感觉“毛”细胞， 不能在损坏后再生。超过100个人类基因的变异与耳聋有关， 其中许多改变了毛细胞中的感觉器的结构，称为毛束。发束 由毛细胞表面排列整齐的高度渐变的膜突起或静纤毛组成。 发育过程中的毛束形成及其在生命中的精确结构容易受到 各种遗传、环境和年龄相关的损伤，所有这些都会导致衰退或完全丧失 听觉能力。我的首要目标是描述蛋白质的相互作用，这对头发束的生长至关重要。 发展和终身维护。在这个建议中，我确定了一个新的作用， 在小鼠毛束中富集的蛋白质，特别是在最高的静纤毛的尖端。一组五 蛋白质，被称为延伸复合物，以前报道在这个相同的隔室。损失 该复合体的任何成员阻止了静纤毛的适当伸长，模糊了静纤毛的独特性 并导致人类和小鼠的深度耳聋。有趣的是，类似的缺陷 在缺乏机械电转导通道的关键组分的小鼠突变体中报道。这 表明，通过转导，活跃的毛束以某种方式影响伸长因子，因此， 静纤毛尺寸我提出，在这个建议中研究的钙结合蛋白是一个新的结合 MYO 15 A的伴侣和调节剂，MYO 15 A是肌球蛋白马达，将其他延伸复合物蛋白运送到 静纤毛尖端。MYO 15 A是最早与听力损失相关的蛋白质之一，MYO 15 A突变 是人类遗传性耳聋的第三大常见原因。本提案的目标1将确定 MYO 15 A和我们的钙结合蛋白之间的关系，并确认初步结果表明， 这种新蛋白质对听觉功能至关重要。目标2将研究钙水平的变化 在毛束中的表达影响我们的蛋白质的动态定位及其延伸 复杂的合作伙伴。总之，这些目标确定和研究这种新的静纤毛蛋白作为一种新的蛋白质。 关键的延长复合体的额外成员，并作为读取毛细胞转导的介体 活动影响静纤毛生长。这个项目采用先进的鼠标模型， 培养听觉器官的技术，以及保存的和活的毛细胞的高分辨率成像。 该项目的完成将有助于我的长期目标，即为治疗策略提供信息， 毛细胞的正常发育和终身保存。

PROJECT SUMMARY/ABSTRACT Our ability to hear relies on a small population of specialized sensory ‘hair’ cells in the inner ear that cannot regenerate upon damage. Variants of over 100 human genes have been associated with deafness, many of which alter the structure of the sensory organelle in hair cells, called the hair bundle. The hair bundle consists of organized rows of graded-height membrane protrusions, or stereocilia, on the surface of hair cells. Hair bundle formation during development and its precise architecture during life are vulnerable to a variety of genetic, environmental, and age-related insults, all of which result in the decline or complete loss of hearing ability. My overarching goal is to characterize protein interactions that are vital to the hair bundle’s development and lifelong maintenance. In this proposal, I identify a new role for a calcium-binding protein enriched in the mouse hair bundle, specifically, at the tip of the tallest stereocilia. A group of five proteins, referred to as the Elongation Complex, was previously reported at this same compartment. Loss of any member of this complex prevents proper stereocilia elongation, blurs the distinct identity of stereocilia across rows, and results in profound deafness in both humans and mice. Interestingly, similar defects have been reported in mouse mutants lacking key components of the mechanoelectrical transduction channel. This suggests that, via transduction, active hair bundles somehow influence elongation factors, and thus stereocilia dimensions. I propose that the calcium-binding protein studied in this proposal is a new binding partner and regulator for MYO15A, the myosin motor that transports other Elongation Complex proteins to stereocilia tips. MYO15A was one of the first proteins associated with hearing loss, and MYO15A mutations are the third most common origin of heredity deafness in humans. Aim 1 of this proposal will determine the relationship between MYO15A and our calcium-binding protein, and confirm preliminary results suggesting that this new protein is essential for auditory function. Aim 2 will investigate how changes in calcium levels in the hair bundle upon transduction affect the dynamic localization of our protein and its Elongation Complex partners. Together, these aims identify and investigate this new stereocilia protein as an additional member of the crucial Elongation Complex, and as a mediator that reads hair cell transduction activity to influence stereocilia growth. This project employs cutting- edge mouse models, advanced techniques to culture the auditory organ, and high-resolution imaging of preserved and live hair cells. Completion of the project will contribute to my long-term goal of informing therapeutic strategies to ensure the proper development and lifelong preservation of hair cells.