Molecular mechanisms of outer segment renewal

外节更新的分子机制

基本信息

批准号：
9184560
负责人：
SILVIA C FINNEMANN
金额：
$ 40.04万
依托单位：
FORDHAM UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-12-01 至 2020-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9184560
关键词：
Age Age related macular degeneration Alpha Cell Animal Model Animals Apoptosis Attenuated Biochemical Cell Death Cell membrane Cells Circadian Rhythms Collaborations Data Defect Development Dimerization Distal Eating Ensure Enzymes Epithelial Excision Eye Eye diseases Frequencies Functional disorder Galectin 1 Genes Health Human Image Impairment Inherited Integrins Length Life Ligand Binding Light Lipids Lipofuscin Maintenance Membrane Membrane Lipids Molecular Mus Neurons Patients Periodicity Phagocytes Phagocytosis Phenocopy Phosphatidylserines Phospholipids Photoreceptors Physiological Processes Process Proteins Rattus Recruitment Activity Regulation Research Retina Retinal Retinal Degeneration Retinal Pigments Retinitis Pigmentosa Rod Outer Segments Rodent Model Role Side Signal Transduction Source Specific qualifier value Structure of retinal pigment epithelium Techniques Time Vision carbohydrate binding protein cell type design dimer extracellular innovation lipid transport mutant novel oxidative damage phospholipid scramblase photoreceptor degeneration prevent public health relevance receptor response retinal rods uptake

项目摘要

DESCRIPTION (provided by applicant): The proposed research aims to elucidate the cellular-molecular mechanisms in the mammalian retina that continuously rejuvenate the light-sensitive outer segment portions of photoreceptor neurons. This fundamental retinal process, known as outer segment renewal, is essential for life-long visual function: Experimental deletion of any gene known thus far to encode a protein of the conserved mammalian outer segment renewal machinery results in progressive loss of photoreceptor function and, eventually, photoreceptor degeneration in rodent models. Furthermore, inherited aberration in outer segment renewal causes forms of retinitis pigmentosa in human patients that are characterized by severe retinal degeneration. Finally, inefficiency of outer segment renewal with age causes lipofuscin accumulation in the RPE in the human eye, which impairs a number of RPE functions contributing to dysfunction associated with age-related macular degeneration. Mammalian outer segment renewal is a synchronized circadian process that involves collaboration of both photoreceptors and neighboring retinal pigment epithelial (RPE) cells. Diurnal exposure of the anionic membrane lipid phosphatidylserine (PS) at distal tips of rod outer segments triggers their shedding and simultaneous clearance phagocytosis by RPE cells. To date, PS exposure is the only confirmed molecular "eat-me" signal of spent cells (e.g. of any cell undergoing apoptosis) and of shed outer segment tips. Despite its universality, the molecular mechanisms triggering PS exposure are still largely obscure in any cell type. We propose two complementary but independent specific aims to identify molecules, activities and their functional interactions that control decorating distal rod outer segment tips with PS in a circadia rhythm in the mammalian retina. Employing innovative live and fixed imaging and quantitative biochemical approaches will allow tracking the entire outer segment renewal process in mutant animals that lack candidate proteins we hypothesize to govern rod PS exposure. In specific aim 1, focus will be on the role of the transmembrane phospholipid scramblase TMEM16F, which localizes to rod outer segment plasma membranes where we propose it to control membrane asymmetry to yield diurnal PS-marked tips. In support, we show: -TMEM16F is elevated in the retina at light onset; -inhibition of TMEM16F prevents ex vivo triggering of PS exposure at rod tips; -mice lacking TMEM16F phenocopy RCS rats and MerTK-/- mice defective in RPE phagocytosis, with normal outer segment development but abnormal RPE phagocytosis followed by progressive photoreceptor loss. In specific aim 2, focus will be on the role of the secreted carbohydrate binding protein galectin-1, which resides in the subretinal space where we propose it to contribute to PS exposure at rod outer segment tips from the extracellular side. In support, we show: -RPE cells are a source of galectin-1; -the PS-externalizing form of galectin-1 is elevated in the retina at light onset; -adding galectin-1 triggers PS externalizationat rod tips; -outer segment renewal in mice lacking galectin-1 is abnormal.

描述（由申请人提供）：拟议的研究旨在阐明哺乳动物视网膜中不断使感光神经元的光敏感外节部分恢复活力的细胞分子机制，这种基本的视网膜过程（称为外节更新）对于视网膜的再生至关重要。终生视觉功能：实验性删除迄今为止已知编码保守哺乳动物外节更新机制蛋白质的任何基因，都会导致感光功能逐渐丧失，并最终导致此外，外节更新的遗传性畸变会导致人类患者出现色素性视网膜炎，其特征是严重的视网膜变性。它损害了许多 RPE 功能，导致与年龄相关的黄斑变性相关的功能障碍，这是一个同步的昼夜节律过程，涉及两个光感受器的协作。和邻近的视网膜色素上皮（RPE）细胞，每日暴露于视杆外节远端的阴离子膜脂质磷脂酰丝氨酸（PS）会触发它们的脱落并同时被RPE细胞清除吞噬作用，迄今为止，PS暴露是唯一被证实的分子“。消耗细胞（例如任何经历凋亡的细胞）和脱落的外节尖端的“吃我”信号。尽管具有普遍性，但触发 PS 暴露的分子机制在很大程度上仍然是未知的。我们提出了两个互补但独立的具体目标，以识别利用创新的实时和固定成像和定量技术在哺乳动物视网膜的昼夜节律中控制装饰远端杆外节尖端的分子、活动及其功能相互作用。生化方法将允许追踪突变动物的整个外节更新过程，这些动物缺乏我们所追求的控制视杆 PS 暴露的候选蛋白。在具体目标 1 中，重点将放在跨膜磷脂扰乱酶 TMEM16F 的作用上。其定位于视杆外节质膜，我们建议它控制膜不对称性以产生昼夜 PS 标记尖端。杆尖PS暴露；-缺乏TMEM16F表型的RCS大鼠和RPE吞噬功能缺陷的MerTK-/-小鼠，具有正常的外部在具体目标 2 中，重点将放在分泌型碳水化合物结合蛋白半乳糖凝集素 1 的作用上，该蛋白位于视网膜下腔，我们认为它有助于视杆细胞的 PS 暴露。作为支持，我们显示： -RPE 细胞是半乳糖凝集素-1 的来源； -半乳糖凝集素-1 的 PS 外化形式在光照下在视网膜中升高。开始； - 添加半乳糖凝集素 1 触发杆尖 PS 外化； - 缺乏半乳糖凝集素 1 的小鼠外节更新异常。