ACTIVITY-DRIVEN PLASTICITY OF THE HAIR CELL CYTOSKELETON

活动驱动的毛细胞细胞骨架的可塑性

• 批准号: 10748106
• 负责人: Alejandra Catalina Velez Ortega
• 金额: $ 44.2万
• 依托单位: UNIVERSITY OF KENTUCKY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-06-07 至 2028-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10748106
• 关键词: Actins; Adult; Affect; Architecture; Auditory; Cells; Cochlea; Crosslinker; Cytoskeleton; Data; Development; Exhibits; F-Actin; Filament; Fluorescence Recovery After Photobleaching; Freeze Substitution; Freezing; Hair; Hair Cells; Hearing; Height; Impairment; In Situ; Ions; Knowledge; Label; Labyrinth; Lead; Length; Link; Maintenance; Measures; Mediating; Microfilaments; Molecular; Morphology; Mus; Myosin ATPase; Natural regeneration; Noise-Induced Hearing Loss; Organelles; Organism; Pattern; Physiological; Polymers; Process; Protein Isoforms; Proteins; Rattus; Reporting; Rest; Sampling; Scanning Electron Microscopy; Sensory; Shapes; Stereocilium; Structure; Supporting Cell; Testing; Transmission Electron Microscopy; beta Actin; cellular microvillus; congenital deafness; crosslink; depolymerization; experimental study; gamma Actin; hearing impairment; mutant mouse model; new therapeutic target; noise exposure; polymerization; postnatal; sound; tomography; vibration

PROJECT SUMMARY/ABSTRACT The mechanosensitivity of the inner ear hair cells depends on cellular projections known as stereocilia, organized in rows of increasing height, with mechano-electrical transduction (MET) channels located at the tips of shorter row stereocilia. The core of stereocilia consists of a highly crosslinked paracrystalline array of actin filaments. While crosslinker proteins are constantly renewed, the renewal of actin is limited to the stereocilia tips. We previously reported that the stereocilia actin core exhibits activity-dependent plasticity (Velez-Ortega, et al., eLife 2017). We showed that the blockage of MET channels or the breakage of the tip links that gate these channels lead to the selective shortening of transducing stereocilia (i.e. the stereocilia that harbor MET channels), while the non-transducing tallest row stereocilia remain unaffected. Once the MET blockage is removed or the tip links regenerate, the stereocilia regrow. Our preliminary data also show that this MET-dependent stereocilia remodeling can affect the resting tension within the MET machinery in seconds. Thus, this process may dynamically regulate the sensitivity of hair cells to sound-induced vibrations and, hence, the sensitivity of our hearing. Yet, the exact mechanisms of MET-dependent stereocilia remodeling are still obscure. It is unknown even whether the activity-dependent plasticity of the stereocilia cytoskeleton is limited to the regions of active actin renewal or can expand beyond this region into the “stable” part of the stereocilia shaft. Here, we hypothesize that the MET activity regulates the extent of the stereocilia cytoskeleton undergoing active actin remodeling. To test this, Aim 1 will evaluate MET-dependent changes in actin dynamics within the stereocilia and the cuticular plate, an actin-rich structure supporting the stereocilia bundle. Aim 2 will evaluate MET-driven changes in the ultrastructural organization of stereocilia actin with transmission electron microscopy tomography. Since the MET-dependent stereocilia remodeling was studied so far only in young postnatal hair cells, Aim 3 will assess whether this phenomenon is present also in the mature adult auditory hair cells. In Aim 4, we begin to explore the molecular players involved in the MET-driven stereocilia remodeling, by evaluating the expression of so-called “stereocilia row identity proteins” in a mutant mouse model that exhibits MET-dependent actin remodeling not only in transducing stereocilia but also, unexpectedly, in non-transducing stereocilia. The study is significant, because it may clarify how exactly a hair cell performs fine adjustments of the architecture of the stereocilia bundle, thereby maintaining the sensitivity of our hearing throughout a lifetime. In addition, stereocilia shortening—and perhaps their eventual disappearance—could occur after noise exposure (when the MET current is reduced due to tip link breakage) or in certain cases of congenital deafness (due to impaired MET current). Therefore, this study will expand our knowledge of the molecular mechanisms of various types of hearing loss.

项目总结/摘要 内耳毛细胞的机械敏感性取决于细胞投射， 静纤毛，排列成行，高度逐渐增加，具有机械-电转换（MET）通道 位于短列静纤毛顶端。静纤毛的核心由高度交联的 肌动蛋白丝的准晶体排列。虽然交联剂蛋白质不断更新， 肌动蛋白仅限于静纤毛尖端。我们先前报道了静纤毛肌动蛋白核心表现出活性依赖性可塑性（Velez-Ortega等人，eLife 2017）。我们发现MET通道的阻塞 或门控这些通道的尖端连接的断裂导致转导的选择性缩短， 静纤毛（即，具有MET通道的静纤毛），而非转导最高行静纤毛 不受影响。一旦MET堵塞被去除或尖端连接再生，静纤毛就会再生。 我们的初步数据还表明，这种MET依赖的静纤毛重塑可以影响静息的 金属爆炸装置的张力因此，该过程可以动态地调节灵敏度 毛细胞对声音引起的振动的敏感性，因此，我们的听觉的敏感性。然而， MET依赖性静纤毛重塑的机制仍然不清楚。目前还不清楚， 静纤毛细胞骨架的活性依赖性可塑性仅限于活跃的肌动蛋白更新区域 或者可以扩张超过该区域进入静纤毛轴的“稳定”部分。在这里，我们假设， MET活性调节静纤毛细胞骨架经历主动肌动蛋白重塑的程度。 为了验证这一点，Aim 1将评估静纤毛内肌动蛋白动力学的MET依赖性变化， 角质板，支持静纤毛束的富含肌动蛋白的结构。目标2将评估MET驱动 静纤毛肌动蛋白超微结构的变化 断层扫描由于MET依赖的静纤毛重塑迄今为止仅在年轻的出生后研究 Aim 3将评估这种现象是否也存在于成熟的成年听毛中 细胞在目标4中，我们开始探索参与MET驱动的静纤毛的分子参与者 重塑，通过评估突变小鼠中所谓的“静纤毛行同一性蛋白”的表达， 模型显示MET依赖的肌动蛋白重塑不仅在转导静纤毛， 出乎意料地，在非转导静纤毛中。这项研究意义重大，因为它可能会阐明 毛细胞对静纤毛束的结构进行精细的调整，从而维持 听觉的灵敏度。此外，静纤毛缩短，也许他们的 最终消失-可能发生在噪声暴露后（当MET电流由于尖端而降低时 连接断裂）或在某些先天性耳聋的情况下（由于受损的MET电流）。因此本 这项研究将扩大我们对各种类型听力损失的分子机制的认识。