Bioenergetic and Proteolytic Impact of Ubiquitin-like Pathways in Metabolically Stressed Neurons

代谢应激神经元中泛素样途径的生物能和蛋白水解影响

基本信息

批准号：
9761342
负责人：
Neal Bennett
金额：
$ 6.16万
依托单位：
J. DAVID GLADSTONE INSTITUTES
依托单位国家：
美国
项目类别：
财政年份：
2019
资助国家：
美国
起止时间：
2019-09-01 至 2022-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9761342
关键词：
Acute Address Affect Aging Bioenergetics Cell Respiration Cell physiology Cells Cellular Stress Clustered Regularly Interspaced Short Palindromic Repeats Consumption Data Disease Exposure to Failure Functional disorder Genes Genetic Genome Glycolysis Heat shock proteins Homeostasis Human Individual Knowledge Measures Mediating Metabolic Metabolic stress Metabolism Mitochondria Nerve Degeneration Neurodegenerative Disorders Neurons Parkinson Disease Pathology Pathway interactions Population Post-Translational Protein Processing Process Production Protein Biosynthesis Proteins Reactive Oxygen Species Respiration Role Stress Technology Ubiquitin Ubiquitin-Activating Enzymes Western Blotting base biological adaptation to stress cell growth consumption measures induced pluripotent stem cell inhibitor/antagonist insight knock-down neuronal survival novel preservation protein degradation protein metabolism proteostasis response screening sensor tool transcriptome sequencing whole genome

项目摘要

PROJECT SUMMARY The aging process and many aging-associated diseases like Parkinson's disease (PD) are hypothesized to be caused by a decline in cellular energy and mitochondrial function. Cellular energy is presumably critical to many cellular processes including protein degradation, a process that is disrupted in neurodegenerative disease. Identifying genes that modulate levels of ATP, the main energy-carrying molecule in all cells, could then be critical to slowing or reversing aging-associated pathology, preserving neuron function in response to stress, and providing insight into how energy failure contributes to disease. However, that analysis has been limited by a lack of tools to screen the genome at high throughput for modifiers of ATP levels, causing a critical gap in knowledge of genetic contributors to energy failure. We have developed a unique screening paradigm to address this knowledge gap by combining genetically encoded ATP sensors with CRISPR-based whole-genome screening technology within cells exposed to acute metabolic stress. With this approach, we have identified three poorly-understood gene pathways that have a prominent impact on ATP levels specifically when cells are metabolically restricted to using only respiration. These pathways are triggered in response to cellular stress, but have also been observed in the pathophysiology of neurodegenerative diseases. While these pathways all regulate key facets of protein metabolism and stress response under normal metabolic conditions, it is unclear how compromised metabolism and low ATP affect these pathways' contributions to the elimination of protein stresses commonly associated with neurodegenerative diseases. We hypothesize that in metabolically-stressed neurons, these stress-responsive pathways exacerbate energy failure and protein accumulation. We will address this hypothesis by investigating if and how these processes affect ATP levels in neurons, as well as the functional consequences of these pathways on protein degradation and survival of metabolically stressed neurons. Successful completion of these aims will provide new insight into the relation between energy homeostasis and proteostasis, as well as the progression of neurodegeneration under metabolic stress.

项目概要衰老过程和许多与衰老相关的疾病，如帕金森病 (PD) 被假设为由细胞能量和线粒体功能下降引起。细胞能量可能对许多人来说至关重要细胞过程，包括蛋白质降解，这一过程在神经退行性疾病中被破坏。然后，识别调节 ATP（所有细胞中主要能量携带分子）水平的基因对于减缓或逆转与衰老相关的病理、保持神经元应对压力的功能至关重要，以及深入了解能量衰竭如何导致疾病。然而，该分析受到以下因素的限制：缺乏高通量筛选基因组中 ATP 水平修饰因子的工具，导致在了解导致能量衰竭的遗传因素。我们开发了一种独特的筛选模式来解决通过将基因编码的 ATP 传感器与基于 CRISPR 的全基因组相结合来弥补这一知识差距暴露于急性代谢应激的细胞内的筛选技术。通过这种方法，我们已经确定三个鲜为人知的基因途径对 ATP 水平有显着影响，特别是当细胞处于代谢仅限于仅使用呼吸作用。这些途径是响应细胞压力而触发的，但也在神经退行性疾病的病理生理学中观察到。虽然这些途径都在正常代谢条件下调节蛋白质代谢和应激反应的关键方面，目前尚不清楚代谢受损和 ATP 低如何影响这些途径对蛋白质消除的贡献通常与神经退行性疾病相关的压力。我们假设在代谢压力下神经元中，这些应激反应通路会加剧能量衰竭和蛋白质积累。我们将通过研究这些过程是否以及如何影响神经元中的 ATP 水平以及这些途径对蛋白质降解和代谢应激生存的功能影响神经元。这些目标的成功完成将为能源与能源之间的关系提供新的见解。稳态和蛋白质稳态，以及代谢应激下神经变性的进展。