Photoreceptor Phosphodiesterase Regulation

光感受器磷酸二酯酶调节

• 批准号: 10699959
• 负责人: Rick H Cote
• 金额: $ 37.45万
• 依托单位: UNIVERSITY OF NEW HAMPSHIRE
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-30 至 2026-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10699959
• 关键词: Acceleration; Allosteric Regulation; Architecture; Binding; Blindness; Catalytic Domain; Code; Communication; Complex; Cone dystrophy; Coupled; DNA Sequence Alteration; Darkness; Defect; Development; Disease; Docking; Enzymes; Etiology; Event; GTP Binding; GTP-Binding Protein Regulators; GTP-Binding Proteins; Genes; Goals; Guanosine Triphosphate; Guanosine Triphosphate Phosphohydrolases; Heterotrimeric GTP-Binding Proteins; Inherited; Light; Lighting; Link; Mediating; Mission; Modeling; Molecular; Molecular Conformation; Molecular Target; Multienzyme Complexes; Mutation; National Eye Institute; Night Blindness; Outcomes Research; Pathogenicity; Pathway interactions; Photoreceptors; Phototransduction; Process; Proteins; RGS1 gene; Recovery; Regulation; Research; Retina; Retinal Cone; Retinal Degeneration; Retinal Diseases; Retinal Dystrophy; Retinitis Pigmentosa; Rod; Signal Pathway; Signal Transduction; Site; Speed; Therapeutic Intervention; Transducin; Vertebrate Photoreceptors; Vision Disorders; Visual; Work; achromatopsia; alpha Subunit Transducin; clinically significant; dimer; disease-causing mutation; next generation sequencing; personalized therapeutic; phosphoric diester hydrolase; photoactivation; programs; retinal rods

PROJECT SUMMARY/ABSTRACT Photoreceptor phosphodiesterase (PDE6) is the central enzyme of the visual signaling pathway. Precise regulation of its activation and deactivation is essential for the speed, sensitivity, and recovery of rod and cone photoreceptors to illumination. Inherited mutations in rod and cone PDE6 genes have been linked in a variety of retinal diseases, including retinitis pigmentosa, congenital stationary night blindness, and cone dystrophy. Next-generation sequencing is identifying a growing number of mutations in PDE6 genes, the large majority of which remain of uncertain clinical significance. Even less is known about the molecular etiology of retinal disease-causing mutations. Rod PDE6 consists of two catalytic subunits whose activity is inhibited in the dark- adapted state by binding of two identical γ-subunits (Pγ). Upon light-induced activation of the visual signaling pathway, PDE6 activity is stimulated by binding of the heterotrimeric G-protein, transducin. The lifetime of light- activated PDE6 is precisely controlled by the rate at which the transducin hydrolyzes its bound GTP, a process controlled by RGS9-1 (Regulator of G-protein Signaling9-1). While the proteins involved in regulation of PDE6 during phototransduction have been identified, the molecular sequence of events in which PDE6 dynamically interacts with its binding partners--as well as its allosteric regulation--during PDE6 activation and deactivation remain poorly understood. Until we understand the mechanistic basis of PDE6 regulation during normal phototransduction, we will be hampered in developing therapeutic interventions for those diseases arising from defects in PDE6 or its binding partners that result in retinal degenerative diseases and visual disorders. The overall objective of this application is to understand the sequence of events accompanying PDE6 activation by transducin and its subsequent inactivation by RGS9-1 and other proteins during recovery of PDE6 to its dark-adapted state. Our experimental plan is based on the hypothesis that the inhibitory Pγ subunit of PDE6 is the “master regulator” responsible for mediating multiple allosteric interactions that occur within the PDE6 catalytic dimer, as well as with the transducin α-subunit and RGS9-1. We propose two specific aims that will (1) delineate the sequence of binding interactions between transducin α-subunits and PDE6 catalytic and inhibitory Pγ subunits to provide a comprehensive model of rod PDE6 activation, and (2) determine the molecular architecture of the PDE6 inactivation complex upon RGS9-1 binding and the structural rearrangements of the Pγ subunit that accelerate termination of activated PDE6. The outcomes of this research advance the goals of the Retinal Diseases Program at the National Eye Institute by enhancing our ability to predict the pathogenicity of mutations in phototransduction proteins, thereby enabling development of personalized therapeutic interventions for retinal diseases resulting from dysregulation of the visual signaling pathway in rod and cone photoreceptor cells of the retina.

项目概要/摘要 光感受器磷酸二酯酶（PDE6）是视觉信号通路的核心酶。精确的 其激活和失活的调节对于视杆细胞和视锥细胞的速度、灵敏度和恢复至关重要 光感受器来照明。视杆细胞和视锥细胞 PDE6 基因的遗传突变与多种疾病有关 视网膜疾病，包括色素性视网膜炎、先天性静止性夜盲症和视锥细胞营养不良。 下一代测序正在识别 PDE6 基因中越来越多的突变，其中绝大多数 仍具有不确定的临床意义。关于视网膜的分子病因学知之甚少 引起疾病的突变。 Rod PDE6 由两个催化亚基组成，其活性在黑暗中受到抑制 通过结合两个相同的 γ 亚基 (Pγ) 来适应状态。光诱导激活视觉信号 途径中，PDE6 活性是通过异源三聚体 G 蛋白转导蛋白的结合来刺激的。光的寿命—— 激活的 PDE6 由转导蛋白水解其结合的 GTP 的速率精确控制，这是一个过程 由 RGS9-1（G 蛋白信号传导调节器 9-1）控制。而参与 PDE6 调节的蛋白质 在光转导过程中，PDE6 动态发生的分子事件序列已被确定。 在 PDE6 激活和失活过程中与其结合伙伴及其变构调节相互作用 仍然知之甚少。直到我们了解正常情况下 PDE6 调节的机制基础 光转导，我们将在开发针对由光转导引起的疾病的治疗干预措施方面受到阻碍 PDE6 或其结合伴侣的缺陷会导致视网膜退行性疾病和视觉障碍。 此应用程序的总体目标是了解伴随 PDE6 的事件顺序 在恢复过程中转导蛋白的激活以及随后 RGS9-1 和其他蛋白质的失活 PDE6 达到暗适应状态。我们的实验计划基于以下假设：抑制性 Pγ 亚基 PDE6 的 PDE6 是负责介导体内发生的多种变构相互作用的“主调节器” PDE6 催化二聚体，以及转导蛋白 α 亚基和 RGS9-1。我们提出两个具体目标 将 (1) 描绘转导蛋白 α 亚基与 PDE6 催化和 抑制性 Pγ 亚基以提供杆 PDE6 激活的综合模型，并且 (2) 确定 RGS9-1 结合后 PDE6 失活复合物的分子结构和结构 Pγ 亚基的重排加速了激活的 PDE6 的终止。这样做的结果 研究通过增强我们的能力来推进国家眼科研究所视网膜疾病计划的目标 能够预测光转导蛋白突变的致病性，从而能够开发 针对视觉信号失调引起的视网膜疾病的个性化治疗干预 视网膜视杆细胞和视锥细胞感光细胞中的通路。