TRP-CHANNELS IN LENS DEVELOPMENT AND CATARACT

项目摘要/摘要 瞬时受体电位(Trp)超家族的非选择性阳离子通道在多种疾病中发挥着重要作用。 生理过程，包括对光、温度、压力和疼痛的感知。在以前的基因中 研究发现，MSTR相关色氨酸通道(TRPM3)的一个成员的突变 儿童遗传性白内障与青光眼和眼前段有不同程度的相关性 缺陷。在这项研究中，我们将研究TRPM3在晶状体发育和白内障病理生物学中的作用。 使用基因编辑的小鼠和细胞系，以达到三个特定的目的。在AIM-1中，我们将描述镜头的特征 RNA深度测序法测定Trp通道转录本剪接变异体及其钙通道特性 使用荧光钙成像技术对晶状体TRPM3亚型进行分析。在AIM-2中，我们将描述 TRPM3功能障碍和缺乏对晶状体阳离子状态、水含量和钙内流特性的影响 使用原子吸收光谱和钙成像技术的组合。在AIM-3中，我们将 TRPM3功能障碍和缺陷对晶状体形态发生和基因表达的影响 分别使用差示免疫荧光显微镜和RNA深度测序技术。 这些研究的结果将为Trp通道在晶状体生长、阳离子中的作用提供新的见解。 儿科白内障的动态平衡、钙动力学和病理生理学 终身视力损害的重要危险因素。更广泛地说，这些研究将有助于 了解Trp通道在其他眼组织(如虹膜、睫状体)中的功能和功能障碍 疾病(例如青光眼、前眼缺陷)。

Project Summary/Abstract The transient receptor potential (TRP) superfamily of non-selective cation channels play critical roles in diverse physiological processes including the perception of light, temperature, pressure, and pain. In previous genetic studies, we have discovered that mutation of a member of the melastatin-related TRP-channels (TRPM3) underlies an inherited form of pediatric cataract that is variably associated with glaucoma and anterior segment defects. In this proposal, we will investigate the role of TRPM3 in lens development and cataract pathobiology using gene-edited mice and cell-lines by addressing three specific aims. In aim-1, we will characterize lens TRP-channel transcript splice-variants by means of RNA deep-sequencing and the calcium channel properties of lens TRPM3 isoforms using fluorometric calcium imaging techniques. In aim-2, we will characterize the effects of TRPM3 dysfunction and deficiency on lens cation status, water content, and calcium influx properties using a combination of atomic absorption spectrometry and calcium imaging techniques. In aim-3, we will characterize the effects of TRPM3 dysfunction and deficiency on lens morphogenesis and gene expression using differential immuno-fluorescence microscopy and RNA deep-sequencing techniques, respectively. Results from these studies will provide new insights regarding the role of TRP-channels in lens growth, cation homeostasis, calcium dynamics, and pathophysiology within the context of pediatric cataract – a clinically important risk factor for lifelong visual impairment. More broadly, these studies will contribute to an understanding of TRP-channel function and dysfunction in other eye tissues (e.g., iris, ciliary body) and diseases (e.g., glaucoma, anterior eye defects), respectively.