Understanding the role of cytosolic NADH production in maintaining aerobic glycolysis in the retina

了解细胞质 NADH 产生在维持视网膜有氧糖酵解中的作用

• 批准号: 10132727
• 负责人: Celia Bisbach
• 金额: $ 3.61万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-03-01 至 2021-12-17
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10132727
• 关键词: Address; Affect; Automobile Driving; Blindness; Cerebellum; Citric Acid Cycle; Complex; Cultured Cells; Cytosol; Data Collection; Disease; Ecosystem; Etiology; Eye; Genes; Glucose; Glycolysis; Health; Investigation; Lead; Link; Malignant Neoplasms; Measurement; Measures; Metabolic; Metabolism; Methods; Mitochondria; Mus; Mutate; NADH; Natural regeneration; Neurons; Oxidative Phosphorylation; Oxides; Photoreceptors; Play; Production; Publishing; Pyruvate; Respiration; Retina; Retinal Cone; Role; Source; Structure of retinal pigment epithelium; Testing; Tissues; Vision; Zebrafish; aerobic glycolysis; alternative oxidase; cell type; comparative; experience; human fetal retinal pigment epithelial cell

ABSTRACT Vision loss affects 1.3 billion people worldwide, with 253 million of those people experiencing vision loss which ranges from severe to total blindness. The etiology of diseases which can cause vision loss are highly complex, as more genes have been identified which can be mutated to cause blindness compared to any other disease. However, many of these genes are involved in metabolic processes. This suggests that it is imperative to have a complete understanding of metabolism in a healthy retina so we can better understand how to treat it when it becomes dysregulated in disease. One aspect of retinal metabolism which seems to be very important for maintaining vision is a high level of aerobic glycolysis performed by photoreceptors. However, we are still far from understanding how photoreceptors maintain this high rate pyruvate to lactate conversion since they also contain functional and underused mitochondria which could use pyruvate to more efficiently make ATP. In our preliminary investigations into this question, we have observed that photoreceptors use GAPDH to maintain a uniquely high cytoplasmic NADH/NAD+ ratio and that high levels of NADH could be sufficient to drive lactate production. However, previously published results have shown that GAPDH is not overexpressed in photoreceptors, so it is still unclear what is driving its high activity. In aim 1 of this proposal, I will address how retinas maintain a uniquely high cytoplasmic NADH/NAD+ ratio by performing a comparative analysis of flux through each step in glycolysis between retinas and oxidative tissues. I predict that I will identify steps early in glycolysis which are rate-limiting for flux through GAPDH and thus cytosolic NADH production in oxidative tissues, but not in retinas. In aim 2 of this proposal, I will use an alternative oxidase to specifically lower the cytoplasmic NADH/NAD+ ratio in cone photoreceptors, which will allow us to address both the importance of cytoplasmic NADH for maintaining aerobic glycolysis, as well as the role of photoreceptor lactate production on the health of the retinal ecosystem as a whole.

摘要 全球有13亿人患有失明，其中2.53亿人患有失明 视力丧失，从严重失明到完全失明。疾病的病因学可以 导致视力丧失的原因非常复杂，因为已经发现了更多可能导致视力丧失的基因 与任何其他疾病相比，突变导致失明。然而，这些基因中的许多是 参与代谢过程的。这表明，有一个完整的 了解健康视网膜的新陈代谢，以便更好地了解如何治疗它 当它在疾病中变得失调时。视网膜代谢的一个方面似乎是 对维持视力非常重要的是高水平的有氧糖酵解 光感受器。然而，我们仍远未了解光感受器如何维持这一点。 丙酮酸转化为乳酸的速率很高，因为它们也含有功能性和未充分利用的成分 线粒体可以利用丙酮酸来更有效地制造ATP。在我们的预赛中 对这一问题的研究，我们观察到光感受器利用GAPDH来维持 一个独特的高细胞质NADH/NAD+比率和高水平的NADH可能就足够了 来推动乳酸盐的生产。然而，先前发表的结果表明，GAPDH是 在光感受器中没有过度表达，所以目前还不清楚是什么推动了它的高活性。 在这项提案的目标1中，我将阐述视网膜如何保持独特的高细胞质 通过对糖酵解每一步的通量进行比较分析，得出NADH/NAD+比率 视网膜和氧化组织之间。我预测我会找出糖酵解的早期步骤 在氧化过程中对通过GAPDH的通量以及细胞内NADH的产生是限速的 组织，但不在视网膜。在这项提议的目标2中，我将使用一种替代的氧化物酶 特别是降低视锥细胞感光细胞胞浆NADH/NAD+比率，这将使我们能够 为了解决细胞质NADH对维持有氧糖酵解的重要性， 由于光感受器乳酸的产生对视网膜生态系统的健康的作用 完整的。