Characterization of purified myocilin: glaucoma as a protein misfolding disease

纯化肌纤蛋白的表征：青光眼作为一种蛋白质错误折叠疾病

基本信息

批准号：
10723134
负责人：
Raquel L Lieberman
金额：
$ 9.97万
依托单位：
GEORGIA INSTITUTE OF TECHNOLOGY
依托单位国家：
美国
项目类别：
财政年份：
2011
资助国家：
美国
起止时间：
2011-03-01 至 2026-02-28
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10723134
关键词：
3-Dimensional Academia Acceleration Affect Age of Onset Alleles Alzheimer&apos s Disease Amyloid Amyloid Fibrils Amyloidosis Amyotrophic Lateral Sclerosis Anatomy Anterior eyeball segment structure Aqueous Humor Benign Blindness Cell Death Cells Chemicals Clinic Comprehension Data Databases Defect Deuterium Development Diagnosis Disease Drainage procedure Endoplasmic Reticulum Exhibits Extracellular Matrix Extracellular Matrix Proteins Eye Eye diseases Fingerprint Foundations Functional disorder Funding Glaucoma Glean Goals Grant Growth High temperature of physical object Homeostasis Horns Hydrogen Industry Inherited Knowledge Lead Length Liquid substance Mass Spectrum Analysis Measures Medical Medicine Methods Missense Mutation Modality Molecular Molecular Chaperones Molecular Conformation Molecular Structure Mutagenesis Mutation Nerve Degeneration Nuclear Magnetic Resonance Open-Angle Glaucoma Outcome Pathogenesis Pathogenicity Pathway interactions Patients Peptides Physiologic Intraocular Pressure Physiology Play Process Protein Biochemistry Proteins Relaxation Research Personnel Retina Risk Factors Role Sampling Structure Susceptibility Gene Testing Therapeutic Time Tissues Toxic effect Trabecular meshwork structure Variant Vertebral column amyloid formation biophysical properties blind cytotoxic cytotoxicity early onset experimental study gain of function genome wide association study infancy insight multidisciplinary mutant myocilin non-Native novel novel therapeutics olfactomedin patient population precision medicine pressure protein folding protein misfolding proteostasis rare variant solid state nuclear magnetic resonance structural biology superoxide dismutase 1 symptom treatment timeline

项目摘要

Glaucoma, a leading cause of blindness worldwide (70 million patients), is managed medically by treating the symptom of increased intraocular pressure (IOP), but 10% of patients still go blind. IOP is controlled in the anterior segment of the eye, which contains the trabecular meshwork (TM) extracellular matrix, the anatomical pathway for drainage of aqueous humor fluid. The TM tissue is diseased in most forms of glaucoma; loss of TM homeostasis leads to elevated IOP. Hereditary open angle glaucoma, affecting ~3 million young patients, is caused by mutations in myocilin, a protein highly expressed in the TM. Since 3/2011, studies funded from R01EY021205 have changed the paradigm for anti-glaucoma therapeutics by laying the molecular foundation for approaches that target the disease process, which are now being pursued in academia and industry. We clarified molecular details of the toxic gain-of-function pathogenic mechanism in which mutant myocilin accumulates in the endoplasmic reticulum (ER) of TM cells, leading to TM cell death and an accelerated timeline for vision loss. Studies from R01EY021205 (a) contributed fundamental knowledge of myocilin structure, (b) discovered a counter-productive interaction between myocilin and the ER-resident Hsp90 chaperone Grp94, and (c) characterized myocilin misfolding as amyloid. Wild-type and many different myocilin variants harbor a misfolding propensity; thus, proteostasis issues identified in familial myocilin- associated glaucoma are likely at play in many more patients. Amyloid formation by myocilin places glaucoma alongside more well-studied amyloid diseases like Alzheimer and SOD-1 dependent amyotrophic lateral sclerosis, yet our comprehension of the role of amyloid in glaucoma is in its infancy. Our current objective is to better understand molecular aspects of myocilin fibrilization, focused on the relevant olfactomedin (OLF) domain. Our multidisciplinary team will (a) clarify initiation of aggregation by studying solution structures of wild-type and selected OLF variants, as well as corresponding multi-length scale dynamics, using hydrogen-deuterium exchange mass spectrometry and nuclear magnetic resonance (NMR) structure and relaxation methods (Wade Van Horn, Co-I) (b) compare the end-point structures of selected OLF aggregates to known amyloids by solid state NMR (Anant Paravastu, Co- I) and evaluate cytotoxicity of intermediate aggregates and (c) evaluate common allele full-length myocilin variants for experimental hallmarks of pathogenicity. The expected outcome is a better understanding of the myocilin misfolding process at the molecular level, including molecular determinants of pathogenicity, to enable novel modalities for studying, diagnosing, and treating myocilin-associated glaucoma. More broadly, continued structure/dysfunction studies of myocilin will not only contribute to our understanding of glaucoma and its role in the TM, but will also extend our comprehension of the many other OLF domains, which are implicated broadly in physiology and diseases.

青光眼是全球失明的主要原因（7,000万患者），通过治疗来管理眼内压力增加（IOP）的症状，但10％的患者仍然失明。 IOP在眼睛的前部，其中包含小梁网（TM）细胞外基质，解剖学流体流体排水的途径。 TM组织以大多数形式的青光眼患病。损失 TM稳态导致IOP升高。遗传性开倾角青年瘤，影响约30万年轻患者，是由在TM中高度表达的肌动蛋白突变引起的。自3/2011以来，从 R01EY021205通过奠定分子基础改变了抗云可瘤疗法的范例对于针对疾病过程的方法，该方法现在正在学术界和工业中采用。我们阐明了有毒功能获得的致病机制的分子细节，其中突变体肌动蛋白积聚在TM细胞的内质网（ER）中，导致TM细胞死亡和A 加速视力丧失的时间表。 R01EY021205（a）的研究贡献了基本知识肌动蛋白的结构，（b）发现了肌动蛋白与er-ersentent之间的适得其反相互作用 HSP90伴侣GRP94，（C）将肌动蛋白折叠折叠为淀粉样蛋白。野生型和许多不同的肌动蛋白变体具有错误折叠的倾向；因此，在家族性肌动蛋白 - 相关的青光眼可能会在更多患者中发挥作用。肌动蛋白的淀粉样蛋白形成将青光眼与更精心培养的淀粉样蛋白疾病一样阿尔茨海默氏症和SOD-1依赖性肌萎缩性横向硬化症，但我们对淀粉样蛋白的作用的理解青光眼仍处于起步阶段。我们目前的目标是更好地了解肌动蛋白的分子方面纤维化，重点是相关的嗅觉素（OLF）结构域。我们的多学科团队将（a）澄清通过研究野生型和选定的OLF变体的溶液结构以及使用氢欧交换质谱法和相应的多长度尺度动力学核磁共振（NMR）结构和松弛方法（Wade Van Horn，Co-I）（b）比较固态NMR（Anant Paravastu，co- i）并评估中间聚集体的细胞毒性和（c）评估普通等位基因全长肌动蛋白致病性实验标志的变体。预期的结果是对在分子水平上的肌动蛋白错误折叠过程，包括致病性的分子决定因素，以实现研究，诊断和治疗肌动蛋白相关的青光眼的新型方式。更广泛地，继续肌动蛋白的结构/功能障碍研究不仅有助于我们对青光眼及其作用的理解在TM中，但也将扩展我们对许多其他OLF领域的理解，这与在生理和疾病方面广泛。