Mechanism and Functional Significance of Polarity Reversal in Mechanosensory Organs

项目摘要/摘要 在40岁以上的成年人中，有高达35%的人患有前庭疾病。对前庭内耳的研究有 对我们如何处理和补偿头部运动产生了重要的见解，包括 传入神经中的平行信息通道。例如，在黄斑器官中，有两组毛发 细胞采用相反的平面方向的发束，因此对头部运动的反应相反。 这种高度保守的双向组织首先在神经肥大，即侧线器官中被描述。 感知鱼体内的水分运动，但在发育过程中实现这种逆转的遗传程序是 只是刚刚开始被破译。因此，消融研究揭示了反转的重要性 直到最近，前庭功能才得以实现。在这里，我们建议通过以下方式来解决这个问题 研究孤儿G蛋白偶联受体(GPCRx)失活的后果，涉及 我们在小鼠毛细胞上皮细胞取向逆转方面的初步数据。根据我们的初步数据，我们 提示小鼠GPCRx在转录因子EMX2下游和转录因子EMX2上游发挥作用 异三聚体G蛋白GI逆转由平面细胞极性蛋白建立的基态极性。 我们将验证这一假设，并使用GPCRx突变体作为动物模型来精确定位极性 反转的形状，黄斑器官的反应和对前庭行为的下游影响。要达到这些目标 目标，我们将：1)使用遗传学来确定GPCRx如何在分子水平上指示逆转，解决其 与EMX2、GI和平面细胞极性蛋白的上位性关系，并用斑马鱼检验 GPCRx-GI是一条保守的逆转效应通路。2)使用分子标记、电生理学和 钙成像解决毛细胞的成熟和功能，在没有反转的情况下。3)确定如何 极性反转影响传入的组织和功能，包括传入记录和整体 前庭功能采用行为学测试。我们连贯的初步证据确保了可行性 和对该项目的高度兴趣，以及我们对几乎没有研究过的受体蛋白的关注 创新。多PI团队非常适合解决鼠标和 斑马鱼的侧声系统。我们预计，这一合作努力将对以下方面起决定性作用 解决毛细胞取向逆转的机制及其在脊椎动物中的保守性及其意义 用于哺乳动物前庭生理学。透彻地理解极性反转将有助于解释和设计 前庭功能障碍的治疗。

PROJECT SUMMARY/ABSTRACT Vestibular disorders affect as many as 35% of adults past age 40. Studies of the vestibular inner ear have yielded important insights into how we process and compensate for head motion including the existence of parallel channels of information in the afferent nerve. In macular organs, for example, two populations of hair cells adopt opposite planar orientations of their hair bundles and thus opposite responses to head movements. This highly conserved bidirectional organization was first described in neuromasts, the lateral line organs sensing water movements in fish, but the genetic program implementing this reversal during development is only starting to be deciphered. Consequently, ablation studies to reveal the importance of reversal for vestibular function have not been possible until recently. Here we propose to address this question by investigating the consequences of inactivating an orphan G protein coupled receptor (GPCRx), implicated by our preliminary data in orientation reversal in mouse hair cell epithelia. Based on our preliminary data, we suggest that mouse GPCRx functions downstream of the transcription factor EMX2 and upstream of the heterotrimeric G protein Gi to reverse a ground state of polarity established by planar cell polarity proteins. We will test this hypothesis and also use the GPCRx mutant as an animal model to pinpoint how polarity reversal shapes macular organ responses and downstream effects on vestibular behaviors. To reach these goals, we will: 1) Use genetics to determine how GPCRx instructs reversal at the molecular level, solving its epistatic relationship to EMX2, Gi and planar cell polarity proteins in mice, and use zebrafish to test whether GPCRx-Gi is a conserved effector pathway for reversal. 2) Use molecular markers, electrophysiology and calcium imaging to resolve hair cell maturation and function in absence of polarity reversal. 3) Determine how polarity reversal affects afferents' organization and function, with afferent recordings, as well as overall vestibular function using behavioral tests. Our coherent body of preliminary evidence ensures the feasibility and the high interest of the project, and our focus on a virtually unstudied receptor protein guarantees innovation. The multi-PI team is ideally suited to address complementary questions in both the mouse and zebrafish acoustico-lateralis systems. We anticipate that this collaborative effort will be decisive towards solving the mechanism of hair cell orientation reversal, its conservation across vertebrates and its significance for mammalian vestibular physiology. Thorough understanding of polarity reversal will help interpret and design treatments for vestibular dysfunctions.