Photoreceptor Phosphodiesterase Regulation

光感受器磷酸二酯酶调节

• 批准号: 10346165
• 负责人: Rick H Cote
• 金额: $ 37.23万
• 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/10346165
• 关键词: Allosteric Regulation; Architecture; Binding; Blindness; Catalytic Domain; Code; Communication; Complex; Cone dystrophy; Coupled; DNA Sequence Alteration; Defect; Development; Disease; Docking; Enzyme Activation; Enzymes; Etiology; Event; GTP Binding; 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; 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; base; clinically significant; dimer; disease-causing mutation; enzyme activity; next generation sequencing; personalized therapeutic; phosphoric diester hydrolase; photoactivation; programs; protein B; 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.

项目摘要/摘要 光感受器磷酸二酯酶（PDE6）是视觉信号通路的中心酶。精确的 调节其激活和失活对于杆和锥的速度，灵敏度和恢复至关重要 光感受器具有照明。杆和锥PDE6基因中的遗传突变已连接 视网膜疾病，包括色素性视网膜炎，先天性固定夜失明和锥体营养不良。 下一代测序是确定越来越多的PDE6基因突变，这是绝大多数 这仍然是不确定的临床意义。关于视网膜的分子病因，甚至更少 引起疾病的突变。 ROD PDE6由两个催化亚基组成，它们的活性在黑暗中被抑制 通过两种相同的γ-亚基（Pγ）结合来适应状态。视觉信号的光诱导激活后 途径，PDE6活性是通过异三聚体G蛋白透射蛋白的结合而刺激的。光的寿命 激活的PDE6精确地由跨多丁素水解其结合GTP的速率控制，这是一个过程 由RGS9-1控制（G蛋白信号9-1的调节剂）。而参与PDE6调节的蛋白质 在光转导期间，已经确定了PDE6动态的事件的分子序列 与其结合伙伴互动 - 以及其变构调节 - 干燥的PDE6激活和停用 保持不当理解。直到我们了解正常情况下PDE6调节的机理基础 光转导，我们将受到阻碍，以开发针对这些疾病的治疗干预措施 PDE6或其结合伴侣的缺陷导致永久性退化性疾病和视觉疾病。 该应用程序的总体目的是了解参与PDE6的事件的顺序 转霉素及其随后在恢复过程中被RGS9-1和其他蛋白质灭活 pde6至其黑暗适应状态。我们的实验计划是基于抑制性Pγ亚基的假设 PDE6的“主调节器”负责介导在此内发生的多种变构相互作用 PDE6催化二聚体，以及带状蛋白α-亚基和RGS9-1。我们提出了两个具体目标 将（1）描述跨核素α-亚基与PDE6催化和PDE6之间的结合相互作用的序列 抑制性Pγ亚基提供了ROD PDE6激活的综合模型，（2）确定 RGS9-1结合和结构的PDE6灭活复合物的分子结构 加速活化PDE6的Pγ亚基的重排。结果的结果 研究通过增强我们的国家眼睛研究所的视网膜疾病计划的目标 能够预测光转导蛋白中突变的致病性，从而实现 视觉信号失调引起的永久性疾病的个性化热干预措施 视网膜的杆和锥形感光细胞中的途径。