The role of the let-7 microRNA gradient in the development of tonotopic specialization in the cochlea

let-7 microRNA 梯度在耳蜗音调专业化发展中的作用

• 批准号: 9483543
• 负责人: Meenakshi Prajapati
• 金额: $ 4.45万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-05-01 至 2020-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9483543
• 关键词: ARHGEF5 gene; Adult; Antibodies; Apical; Auditory; Auditory Brainstem Responses; Auditory system; Cell Cycle; Cell Differentiation process; Cell Size; Cellular Morphology; Cochlea; Computer software; Cues; Data; Detection; Development; Discrimination; Doxycycline; Electrophysiology (science); Epithelium; Family; Frequencies; Gene Expression; Gene Targeting; Genes; Growth; Hair Cells; Harvest; Hearing; Hearing problem; Human; Image; Intrinsic factor; Label; Length; Life; Measures; Membrane; Metabolism; MicroRNAs; Molecular; Morphology; Mus; Music; Neurons; Organ; Outer Hair Cells; Patients; Perception; Preparation; Production; Property; RNA-Binding Proteins; Reporting; Resolution; Rodent; Role; Sensory; Signal Transduction; Speech; Stem cells; System; Tetanus Helper Peptide; Transgenic Mice; base; biophysical properties; cell dimension; cell growth; cellular transduction; experimental study; fluorescence imaging; hair cell regeneration; insight; molecular marker; neuronal cell body; neurotransmission; overexpression; postnatal; ranpirnase; reconstruction; sound; sound frequency

Project Summary The cochlea is the organ in the auditory system responsible for the detection of sound. It houses a spiral- shaped sensory epithelium containing mechanosensory hair cells that transduce sound waves into neuronal signals. This sensory epithelium is tonotopically organized such that it detects high frequency sounds at the base of the spiral and low frequency sounds at the apex. The tonotopic specializations include a graded increase in hair cell size from the base to the apex, as well as graded changes in the electrophysiological properties of hair cells and neurons. However, the extrinsic and intrinsic factors controlling cochlear tonotopic specialization are largely unknown. The main aim of this proposal is to characterize the role of the LIN28B/let-7 axis in tonotopic specialization of auditory hair cells. The pro-growth RNA-binding protein LIN28B and the functionally opposing let-7 family of miRNAs regulate the expression of hundreds of genes, including genes involved in metabolism and cell growth. We have previously shown that LIN28B and let-7 miRNAs are expressed in opposing gradients that inform sensory progenitor cell cycle exit and hair cell differentiation. Our preliminary data show that during cochlear maturation, let-7 is highly expressed in basal outer hair cells, whereas pro-growth let-7 target genes are expressed in apical outer hair cells. Based on these findings we hypothesize that the basal-apical gradient of let-7 miRNA instructs the development of cochlear tonotopic specialization, specifically tonotopic differences in hair cell size and hair cell- specific gene expression. The development of tonotopic specialization in the mouse cochlea has not been well characterized, but it is thought that tonotopic changes start to become evident around the onset of hearing (P11). In aim 1, we propose to establish when tonotopic differences in hair cell/stereocilia size arise in the murine cochlea, by analyzing these features prior (P7), during (P14) and after the onset of hearing (P21, P30 and P60). In addition, we will characterize markers for tonotopic specialization in the murine cochlea. In aim 2, we will use existing transgenic mouse lines to abolish the gradient of let-7 miRNA in the maturing cochlea (P1- P12). At P30, we will assess the effects of these manipulations on frequency-specific hearing, as well as the tonotopic specializations of hair cells. We predict that LIN28B overexpression will produce a more `apical' identity of hair cells and functional deficits in high frequency hearing. Conversely, let-7g overexpression will lead to loss of `apical' hair cell identity and deficits in low frequency hearing. Our findings will provide insight into the molecular mechanisms regulating tonotopic identity of hair cells. This will benefit efforts in hair cell regeneration therapies, as generating appropriately specialized hair cells along the cochlea will be critical to the perception of meaningful sound.

项目摘要 耳蜗是听觉系统中负责检测声音的器官。它有一个螺旋- 包含机械感觉毛细胞的成形感觉上皮，其将声波传导到神经元 信号.这种感觉上皮是按音调组织的，使得它在听觉区检测高频声音。 螺旋的底部和低频率的声音在顶点。音调拓扑特化包括分级的 毛细胞大小从基部到顶端的增加，以及电生理学的分级变化， 毛细胞和神经元的特性。然而，控制耳蜗音调的外在和内在因素， 专业化在很大程度上是未知的。这项建议的主要目的是确定联合国的作用， LIN 28 B/let-7轴与听毛细胞的音调定位特化促生长RNA结合蛋白 LIN 28 B和功能上相反的let-7家族的miRNA调节数百个基因的表达， 包括参与新陈代谢和细胞生长的基因。我们之前已经证明，LIN 28 B和let-7 miRNA以相反的梯度表达，其告知感觉祖细胞周期退出和毛细胞 分化我们的初步数据表明，在耳蜗成熟过程中，let-7在基底膜中高表达， 外毛细胞，而促生长let-7靶基因在顶端外毛细胞中表达。基于这些 我们假设let-7 miRNA的基底-顶端梯度指导了 耳蜗音调定位特化，特别是毛细胞大小和毛细胞的音调定位差异， 特异性基因表达在小鼠耳蜗中音调定位特化的发展还没有被证实。 很好地描述了，但人们认为，音调的变化开始变得明显周围的听力发作 （P11）。在目的1中，我们提出建立在毛细胞/静纤毛大小的张力定位差异何时出现在 通过分析小鼠耳蜗在听力开始前（P7）、听力开始时（P14）和听力开始后（P21，P30）的这些特征， P60）。此外，我们将在小鼠耳蜗tonotopic专业化的标志物的特点。在目标2中， 我们将使用现有的转基因小鼠系来消除成熟耳蜗中let-7 miRNA的梯度（P1-P10）。 P12）。在P30，我们将评估这些操作对频率特异性听力的影响，以及 毛细胞的色调特化。我们预测，LIN 28 B过表达将产生更多的“顶端”细胞。 毛细胞的同一性和高频听力的功能缺陷。相反，let-7 g过表达将 导致“顶端”毛细胞特性丧失和低频听力缺陷。我们的发现将提供 进入调节毛细胞的张力定位特性的分子机制。这将有利于毛细胞的努力 再生疗法，因为沿着耳蜗沿着产生适当特化的毛细胞将是至关重要的， 对有意义的声音的感知。