Molecular determinants of neuronal protein homeostasis through plasma membrane-localized proteasome complexes.

通过质膜定位的蛋白酶体复合物神经元蛋白质稳态的分子决定因素。

• 批准号: 10018941
• 负责人: Kapil Ramachandran
• 金额: $ 42.25万
• 依托单位: HARVARD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-16 至 2021-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10018941
• 关键词: 26S proteasome; Affect; Appointment; Biochemical; Biochemistry; Biological; Biological Process; CRISPR interference; Calcium; Cell membrane; Cell physiology; Cells; Cellular biology; Chemicals; Complement; Complex; Cryoelectron Microscopy; Data; Discipline; Electron Microscopy; Electrophysiology (science); Environment; Etiology; Extracellular Space; Faculty; Foundations; Glycoproteins; Heart; Human; Image; International; Knowledge; Left; Light; Mediating; Membrane; Membrane Proteins; Mentorship; Molecular; Morphology; Mutagenesis; Nature; Nervous system structure; Neurobiology; Neurons; Pathologic; Peptide Fragments; Peptides; Phenotype; Physiological; Physiology; Positioning Attribute; Postdoctoral Fellow; Process; Proteasome Binding; Proteasome Inhibition; Protein Biosynthesis; Protein Dynamics; Proteins; Proteome; Proteomics; Research; Research Personnel; Resource Sharing; Resources; Ribosomes; Role; Signal Transduction; Societies; Specific qualifier value; Specificity; Structure; System; Therapeutic; To specify; Training; Ubiquitin; Uncertainty; Work; Zebrafish; career; fascinate; genome-wide; hydrophilicity; in vivo; inhibitor/antagonist; insight; medical schools; multicatalytic endopeptidase complex; nervous system disorder; neuronal excitability; neuropathology; neuroregulation; novel; particle; post-doctoral training; protein degradation; proteostasis; response; ribosome profiling; student mentoring; tool; volunteer

PROJECT SUMMARY / ABSTRACT Cells continuously respond to physiological signals and potentially pathological perturbations. In response, protein synthesis and protein degradation, the latter of which is predominantly driven by the ubiquitin- proteasome system, reciprocally remodel the intracellular proteome. The dynamics of protein turnover determine the physiological response to a large diversity of signals or perturbations and have major ramifications on human physiology. Indeed, over four decades of work on the ubiquitin conjugating cascade and the 26S proteasome has elucidated essential roles for the ubiquitin-proteasome system in nearly every cellular process. The prevailing principles in protein turnover have been that ubiquitylation is necessary for substrate tagging and that the 26S proteasome is the only proteasome species that degrades ubiquitin-protein conjugates. Though 20S proteasomes form the core of the 26S complex, they remain largely understudied because of a prior lack of clear evidence for functional 20S particles in cells and no insight into 20S-specific substrate targeting. I recently discovered a new mechanism of ubiquitin-independent protein turnover through a highly specialized 20S proteasome that is tightly associated with neuronal plasma membranes. These neuronal membrane proteasomes (NMPs) directly associate with ribosomes to degrade ~250-500 nascent chain substrates independent of ubiquitylation. The NMP degrades substrates across the membrane, releasing resulting peptide fragments into the extracellular space that induce signaling in other neurons, and therefore represents a new mechanism of neuromodulation. Here, I propose studies that will lay the foundation necessary to understand this new paradigm in protein turnover. In my first aim, I will identify how NMPs associate with the plasma membrane and reveal the molecular components of this membrane complex. In the second aim, I will determine how the specificity of NMP-mediated degradation of nascent chains is achieved. In the final aim, I will gain insights into the biological processes that NMP-mediated degradation regulates. The proposed research is significant because it opens a new field of research into non-canonical protein turnover in neurons. This work will generate the tools and mechanistic insight necessary to understanding how NMP-mediated degradation is codified in and relevant to the vertebrate nervous system. This will not only shed light onto the new mechanism of neuromodulation through NMPs, but also provide a framework relevant to abnormalities in protein turnover that underlie multiple human neuropathologies.

项目概要/摘要 细胞不断对生理信号和潜在的病理扰动做出反应。在 反应、蛋白质合成和蛋白质降解，后者主要由泛素驱动 蛋白酶体系统，相互重塑细胞内蛋白质组。蛋白质周转的动态决定 对多种信号或扰动的生理反应，对人类有重大影响 生理。事实上，四十多年来对泛素缀合级联和 26S 蛋白酶体的研究 阐明了泛素-蛋白酶体系统在几乎每个细胞过程中的重要作用。这 蛋白质周转的普遍原则是泛素化对于底物标记是必要的，并且 26S 蛋白酶体是唯一能降解泛素-蛋白质缀合物的蛋白酶体。虽然20S 蛋白酶体构成了 26S 复合体的核心，但由于之前缺乏明确的研究，它们在很大程度上仍未被充分研究。 细胞中具有功能性 20S 颗粒的证据，但没有深入了解 20S 特异性底物靶向。我最近 通过高度专业化的 20S 发现了一种不依赖于泛素的蛋白质周转的新机制 与神经元质膜紧密相关的蛋白酶体。这些神经元膜 蛋白酶体 (NMP) 直接与核糖体结合，降解约 250-500 个新生链底物 独立于泛素化。 NMP 跨膜降解底物，释放所得肽 碎片进入细胞外空间，在其他神经元中诱导信号传导，因此代表了一种新的 神经调节机制。在这里，我提出研究将为理解这一点奠定必要的基础 蛋白质周转的新范例。在我的第一个目标中，我将确定 NMP 如何与质膜结合 并揭示该膜复合物的分子成分。在第二个目标中，我将确定如何 实现了 NMP 介导的新生链降解的特异性。在最终目标中，我将深入了解 NMP 介导的降解调节的生物过程。拟议的研究意义重大 因为它开辟了神经元中非典型蛋白质周转的新研究领域。这项工作将产生 了解 NMP 介导的降解如何编码和编码所需的工具和机制见解 与脊椎动物的神经系统有关。这不仅将揭示新的机制 通过 NMP 进行神经调节，还提供了与蛋白质周转异常相关的框架 是多种人类神经病理学的基础。