The TRPML1 Role in Lysosomes

项目总结。 IV型粘脂沉积症(MLIV)是一种由基因突变引起的溶酶体储存障碍 编码溶酶体离子通道粘蛋白1(TRPML1)。MLIV导致严重的发育 迟缓、精神运动迟缓、认知障碍、角膜混浊和变性 视网膜。在细胞水平上，检测到磷脂和神经节苷脂的积累，以及 几乎所有患者的细胞都含有来源不明的包裹膜结构。自.以来 TRPML1定位于溶酶体，其下调导致严重的溶酶体储存 表型TRPML1被认为是调节溶酶体功能的基因。目前，还没有 解释溶酶体中TRPML1功能的完整模型。建议TRPML1 通过调节细胞内膜的融合和/或分裂来直接调节细胞膜的运输 内吞途径下部的细胞器。另一种模型表明 TRPML1通过调节溶酶体的酸化作用而影响溶酶体的水解。现在 该项目旨在直接回答MLIV发病机制中的两个主要问题：TRPML1是否 直接调节内吞途径中的膜运输以及TRPML1的哪个方面 通透性使得它对于内吞途径的正常功能是不可或缺的。这将是 通过分子、生物化学和生理技术的结合来完成。 具体地说，将使用siRNA驱动的TRPML1下调系统来建立 TRPML1下调对膜运输和溶酶体的直接和即时影响 脂肪水解液。TRPML1及其近邻离子渗透的结构函数分析 将执行相对TRPML3，以确定独特方面的结构决定因素 TRPML1离子渗透。通过TRPML1的离子渗透将被特定地调制 为了确定TRPML1渗透性使TRPML1对于 内吞途径的正常功能。这项提案中计划的实验将 确定MLIV治疗研究的方向，确定细胞内治疗的新角色 离子通道在膜转运、溶酶体代谢和细胞器离子调节中的作用 并提供了有关离子基本机制的新的结构信息 渠道渗透、整顿、封堵。

Project summary. Mucolipidosis type IV (MLIV) is a lysosomal storage disorder caused by mutations in the gene coding for a lysosomal ion channel Mucolipin 1 (TRPML1). MLIV causes severe developmental delays, psychomotor retardation, cognitive disabilities, corneal cloudiness and degeneration of the retina. At the cellular level, accumulation of phospholipids and gangliosides is detected, and virtually all patients’ cells contain enfolded membranous structures of unknown origin. Since TRPML1 localizes in lysosomes and its downregulation causes severe lysosomal storage phenotype, TRPML1 was suggested to regulate lysosomal function. At present, there is no complete model that explains TRPML1 function in the lysosomes. TRPML1 was suggested to directly regulate intracellular membrane traffic by modulating fusion and/or fission of the organelles within the lower portions of the endocytic pathway. An alternative model suggests that TRPML1 affects lysosomal hydrolysis by regulating lysosomal acidification. The present project aims to directly answer the two main question in MLIV pathogenesis: whether TRPML1 directly regulates membrane traffic in the endocytic pathway and which aspect of TRPML1 permeability makes it indispensable for the proper function of the endocytic pathway. This will be accomplished by a combination of molecular, biochemical and physiological techniques. Specifically, an siRNA driven TRPML1 downregulation system will be used to establish the direct and immediate effects of TRPML1 downregulation on membrane traffic and lysosomal lipid hydrolysis. A structure function analysis of ion permeation through TRPML1 and its close relative TRPML3 will be performed to identify the structural determinants of the unique aspects of TRPML1 ion permeation. The ion permeation through TRPML1 will be specifically modulated in order to establish the aspects of TRPML1 permeability that make TRPML1 indispensable for the proper function of the endocytic pathway. The experiments planned in this proposal will establish the direction for the search for MLIV treatments, identify the new roles for intracellular ion channels in regulation of membrane traffic, lysosomal metabolism and organellar ion homeostasis and provide novel structural information regarding the basic mechanisms of ion channel permeation, rectification and block.