Hyperglycemia-induced translational control of gene expression in the retina

项目概要/摘要 糖尿病视网膜病变是美国工作年龄段失明的主要原因， 美国每年新增12,000例。增殖性肿瘤循证治疗原则 糖尿病性视网膜病变涉及激光介导的消融，其不能改变糖尿病性视网膜病变的分子病理学。 因此，近一半的患者需要未来的治疗。因此，我们的总体目标是确定新的 在分子水平上干预的目标，这将导致创新的，非破坏性的 糖尿病视网膜病变的病因是什么？糖尿病视网膜病变的病因是什么？发病机制 是由高血糖症和胰岛素介导的信号传导减少的组合引起的， 其通过诱导结构性和生理性的糖尿病神经血管并发症而导致糖尿病神经血管并发症。 视网膜的变化。本申请中提出的研究是创新的，因为它代表了一种 一种完全不同的方法来解决糖尿病视网膜病变的分子基础，即高血糖诱导 基因表达的翻译控制的改变。核心假设是，添加O- 连接N-乙酰葡糖胺（O-GlcNAc化）至翻译起始因子的丝氨酸或苏氨酸残基 介导从帽依赖性到帽非依赖性mRNA翻译的转变，导致改变的基因 在糖尿病视网膜病变的病理生理学过程中，假设得到支持 通过发现通过己糖胺生物合成途径的葡萄糖通量增加和 mRNA帽结合复合物的关键组分，包括eIF4E结合蛋白1、eIF4G、eIF4A和 在糖尿病诱导的高血糖症条件下，聚A结合蛋白。此外，我们还提供 初步证据表明高血糖症有利于具有内部核糖体进入位点的mRNA的翻译， 例如编码关键血管生长因子的那些，其方式依赖于eIF4F的破坏 复杂的装配。在指导阶段，PI将从以下实验室获得技术专业知识： 博士Gerald哈特关于用于鉴定控制mRNA的蛋白质中的O-GlcNAc化位点的方法 翻译.一旦确定了修饰位点， 将定义损伤eIF4F复合体组装。辅导阶段还将为候选人提供时间 接受托马斯加德纳博士的指导，以评估是否防止eIF4F复合物组装的破坏 足以抑制糖尿病小鼠模型中该疾病发病机制的早期临床前阶段。 关于成果，预计该项目不仅将扩大PI的技能和分析系统， 而且还将确定与糖尿病相关的代谢异常联系起来的新机制， 视网膜中血管生长因子表达增强。确定这种机制意义重大 因为它有望验证用于开发预防和/或治疗药物的新靶点， 这些干预措施旨在解决糖尿病视网膜病变的分子基础并促进健康视力。

Project Summary/Abstract Diabetic retinopathy is the leading cause of blindness in working age Americans, accounting for more than 12,000 new cases in the United States each year. The principle evidenced based treatment for proliferative diabetic retinopathy involves laser-mediated ablation, which fails to alter the molecular pathology of the disease, and as such, nearly half of patients require future treatments. Thus, our overall goal is to identify new targets for intervention at the molecular level that will lead to development of innovative, nondestructive therapies that address treatment of the cause of diabetic retinopathy, rather than the effect. The pathogenesis of this disease is caused by a combination of hyperglycemia and a reduction in insulin mediated signaling, which results in diabetic neurovascular complications through the induction of structural and physiological changes in the retina. The research proposed in this application is innovative, because it represents an entirely different approach to address the molecular basis of diabetic retinopathy, i.e. hyperglycemia-induced alterations in the translational control of gene expression. The central hypothesis is that the addition of O- linked N-Acetylglucosamine (O-GlcNAcylation) to serine or threonine residues of translation initiation factors mediates a shift from cap-dependent to cap-independent mRNA translation, resulting in an altered gene expression pattern that contributes to the pathophysiology of diabetic retinopathy. The hypothesis is supported by findings of elevated flux of glucose through the hexosamine biosynthetic pathway and O-GlcNAcylation of key components of the mRNA cap-binding complex, including eIF4E binding protein 1, eIF4G, eIF4A, and poly(A)-binding protein, under conditions of diabetes-induced hyperglycemia. Furthermore, herein we provide preliminary evidence that hyperglycemia favors the translation of mRNAs with internal ribosome entry sites, such as those encoding key vascular growth factors, in a manner that is dependent on the disruption of eIF4F complex assembly. During the mentored phase, the PI will acquire technical expertise from the laboratory of Dr. Gerald Hart on the methodology used to identify O-GlcNAcylation sites in proteins that control mRNA translation. Once the modified sites have been identified, the mechanisms through which hyperglycemia impairs eIF4F complex assembly will be defined. The mentored phase will also provide time for the candidate to receive guidance from Dr. Thomas Gardner to evaluate if preventing disruption of eIF4F complex assembly is sufficient to inhibit early preclinical phases of the pathogenesis of this disease in a mouse model of diabetes. With respect to outcomes, this project is expected to not only expand the PI's skills and systems of analysis, but will also identify novel mechanisms that link the metabolic abnormalities associated with diabetes to enhanced vascular growth factor expression in the retina. Identification of such mechanisms is significant because it is expected to validate new targets for the development of preventive and/or therapeutic interventions aimed at addressing the molecular basis of diabetic retinopathy and promoting healthy vision.