TDP43 Degradation by the Lysosomal Proteases in Amyotrophic Lateral Sclerosis

肌萎缩侧索硬化症中溶酶体蛋白酶对 TDP43 的降解

• 批准号: 10650830
• 负责人: Paul Joseph Sampognaro
• 金额: $ 22.44万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-07-01 至 2027-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10650830
• 关键词: Alkalinization; Amyotrophic Lateral Sclerosis; Area; Autophagocytosis; Basic Science; Biological Assay; Cathepsins; Cathepsins B; Cell Compartmentation; Cell Survival; Cell physiology; Cells; Clinical; Clinical Trials; Complex; Critical Pathways; Data; Development; Disease; Enzymes; Faculty; Fellowship; Foundations; Functional disorder; Funding; Goals; Half-Life; Homeostasis; Human; Impairment; In Vitro; Incubated; Induced pluripotent stem cell derived neurons; Investigation; K-Series Research Career Programs; Knowledge; Knowledge acquisition; Lysosomes; Mass Spectrum Analysis; Measures; Membrane; Mentors; Mentorship; Methods; Modeling; Morphology; Motor; Motor Neurons; Mutation; Nerve Degeneration; Neurobiology; Neurodegenerative Disorders; Neurology; Neurons; Organelles; Pathogenesis; Pathogenicity; Pattern; Peptide Hydrolases; Phase; Pilot Projects; Process; Proteins; Recycling; Research; Research Personnel; Residencies; Resistance; Running; Scientist; Site; Solid; TDP-43 aggregation; Testing; Therapeutic; Time; Training; Work; age related; biological adaptation to stress; career; design; experience; gain of function; knock-down; loss of function; medical schools; motor deficit; motor neuron degeneration; mutant; neuron loss; neuropathology; novel; novel therapeutics; overexpression; preservation; prevent; protein TDP-43; protein aggregation; purge; skills; sobriety; undergraduate student

PROJECT SUMMARY/ABSTRACT Amyotrophic lateral sclerosis (ALS) is a progressive, uniformly fatal neurodegenerative disorder caused by the loss of upper and lower motor neurons. Despite multiple advanced clinical trials, there are currently no therapies that can stabilize or reverse the motor deficits of ALS. My goal in pursuing a K08 Mentored Clinical Scientist Research Career Development Award is to acquire the knowledge and practical training to make major advances in our understanding of the mechanisms underlying neuronal loss in ALS. With my career, I seek to develop novel therapies that will prevent motor neuron death and preserve function. In 2006 Neumann and colleagues discovered that TAR DNA-binding protein 43 (TDP43) accumulated abnormally within diseased neurons and represented a unifying, end-stage neuropathologic hallmark of ALS. With this observation, the field then converged on the cellular process of autophagy — the method by which cells use lysosomes to purge proteins aggregates and maintain homeostasis — as a critical pathway involved in ALS’ pathogenesis. Lysosomes, however, are tremendously intricate and not well understood compartments within neurons. Lysosomes house many complex, hydrolytic enzymes; they assist in adaptive stress responses; and they promote cell survival broadly. While it has been shown that TDP43 can be directed to the lysosome for clearance, past efforts have stopped their investigations at the lysosomal membrane, presuming that presuming that TDP43 degradation happens efficiently by an array of unspecified enzymes thereafter. My preliminary data demonstrates that only a subset of lysosomal enzymes (cathepsins B, D, E, G, L, K, S, and V) can degrade TDP43 in a pH-dependent manner and that ALS-causing TDP43 mutations have the potential to disrupt the functions of these enzymes. Therefore, I hypothesize that cathepsins B, D, E, G, L, K, S, and V are responsible for TDP43 degradation, pathogenic TDP43 mutations are capable of conferring resistance to these cathepsins, and that impaired cathepsin activity (whether due to TDP43 mutations or age- related changes in pH) contributes to TDP43 buildup, TDP43 aggregation, and neurodegeneration over time. This proposal builds upon my solid foundation in neurobiology and neurology that I have cultivated working as a basic research scientist during my undergraduate, medical school, residency, fellowship, and early faculty years. My current research mentor has an established record of impactful discoveries in the field of neurodegeneration. I have also assembled a team of highly accomplished advisors at UCSF to guide me through this next phase of training on my path to becoming an independent investigator. My training plan is specifically designed to provide me with the mentorship, training in advanced experimental skills, and experience required to run a research group. Completing the research and obtaining the skill sets and mentorship outlined in this proposal will prepare me well to obtain R01 or equivalent funding to begin my career as an independent investigator.

项目概要/摘要 肌萎缩侧索硬化症 (ALS) 是一种进行性、均致命的神经退行性疾病，由 上、下运动神经元丧失。尽管进行了多项先进的临床试验，但目前还没有 可以稳定或逆转 ALS 运动缺陷的疗法。我追求 K08 指导临床的目标 科学家研究职业发展奖是为了获得知识和实践培训 我们对 ALS 神经元丢失机制的理解取得了重大进展。凭借我的事业，我 寻求开发新的疗法来防止运动神经元死亡并保留功能。 2006 年，Neumann 及其同事发现 TAR DNA 结合蛋白 43 (TDP43) 积累 患病神经元内异常，代表 ALS 的统一的终末期神经病理学标志。 通过这一观察，该领域随后集中在自噬的细胞过程上——自噬的方法 细胞利用溶酶体清除蛋白质聚集体并维持体内平衡——作为参与的关键途径 ALS 的发病机制。然而，溶酶体是极其复杂且尚未被充分理解的隔室 神经元内。溶酶体含有许多复杂的水解酶；它们有助于缓解适应性压力 回应；它们广泛促进细胞存活。虽然已经表明 TDP43 可以定向到 溶酶体进行清除，过去的努力已经停止了对溶酶体膜的研究，假设 假设此后一系列未指定的酶有效地发生了 TDP43 降解。 我的初步数据表明，只有溶酶体酶的一个子集（组织蛋白酶 B、D、E、G、L、K、S、 V) 可以以 pH 依赖性方式降解 TDP43，并且导致 ALS 的 TDP43 突变具有 可能破坏这些酶的功能。因此，我假设组织蛋白酶 B、D、E、G、L、K、 S和V负责TDP43降解，致病性TDP43突变能够赋予 对这些组织蛋白酶产生抗性，并损害组织蛋白酶活性（无论是由于 TDP43 突变还是年龄- 随着时间的推移，pH 值的相关变化会导致 TDP43 积累、TDP43 聚集和神经退行性变。 这个建议建立在我在神经生物学和神经病学方面的坚实基础之上，这些基础是我在工作中培养的。 在我的本科、医学院、住院医师实习、研究员和早期教职期间担任基础研究科学家 年。我目前的研究导师在该领域拥有有影响力的发现的既定记录 神经变性。我还在加州大学旧金山分校组建了一支由非常有成就的顾问组成的团队来指导我 通过下一阶段的培训，我将成为一名独立调查员。我的训练计划是 专门为我提供指导、高级实验技能培训，以及 管理一个研究小组所需的经验。完成研究并获得技能组合和 本提案中概述的指导将使我做好充分准备，以获得 R01 或同等资金来开始我的 独立调查员的职业生涯。