Mechanism by Which the Bicaudal D2-Nuclear Pore Protein 358 Interaction Activates Microtubule-based Cargo Transport

双尾 D2-核孔蛋白 358 相互作用激活基于微管的货物运输的机制

• 批准号: 10809832
• 负责人: M Yusuf Ali
• 金额: $ 15.6万
• 依托单位: UNIVERSITY OF VERMONT & ST AGRIC COLLEGE
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-01 至 2025-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10809832
• 关键词: Adaptor Signaling Protein; Alzheimer' s Disease; Amyotrophic Lateral Sclerosis; Autophagosome; Axon; Binding; Binding Proteins; Biological; Biological Assay; Cell physiology; Collaborations; Complex; Coupling; Cytoplasm; Data; Dynein ATPase; Exhibits; Fluorescence Microscopy; Frequencies; Huntington Disease; Impairment; In Vitro; Intracellular Transport; Kinesin; LIS1 protein; Length; Link; Liquid substance; Membrane; Messenger RNA; Microtubule-Associated Proteins; Microtubules; Minus End of the Microtubule; Mitochondria; Molecular Conformation; Monitor; Motion; Motor; Movement; Neurodegenerative Disorders; Neurons; Nuclear Pore; Organelles; Parkinson Disease; Play; Plus End of the Microtubule; Pore Proteins; Positioning Attribute; Proteins; Registries; Regulation; Role; Running; Secretory Vesicles; Signaling Protein; Spastic Paraplegia; Speed; Tail; Testing; Time; Vesicle; Work; cell motility; cofactor; dynactin; experimental study; in vivo; insight; intracellular protein transport; knock-down; mutant; novel; protein complex; reconstitution; recruit; scaffold; sensor; single molecule

Project Summary/Abstract There are many cargo such as secretory vesicles, autophagosomes, and liquid droplets that are bi-directionally transported along microtubules by the opposing motors cytoplasmic dynein and kinesin. However, it is still unclear how these two biological motors collaborate to position the cargo and achieve cellular functions. Impaired bidirectional transport leads to a variety of neurodegenerative diseases, including Alzheimer's disease. Microtubule-associated protein MAP7 and dynein cofactor Lis1 are two important proteins that play critical roles in cargo transport. While MAP7 enhances the processive motion of kinesin, Lis1 relieves the auto- inhibition of dynein. In this proposal, we aim to reconstitute the DDBE-K (dynein, dynactin, BicD, Egl, kinesin) complex and observe its motion on microtubule tracks in the presence of both MAP7 and Lis1 using total internal reflection fluorescence (TIRF) microscopy. Given that MAP7 has the ability to prolong the attachment of kinesin to microtubules and Lis1 prefers binding two dyneins, we predict that MAP7 and Lis1 will play key roles in determining the direction of motion of the complexes on microtubules. Using single-molecule assays, we will investigate whether the adapter protein BicD not only functions as a linker between cargo and motor but also plays an essential role in sensing cargo and motor binding. These proposed studies will provide novel insights into the roles of dynein and cargo adapter proteins and contribute to our understanding of how cytoplasmic dynein and kinesin work together to transport cargo bidirectionally on microtubules.

项目摘要/摘要 有许多货物，例如分泌囊泡，自噬体和液滴 由对方电动机细胞质动力蛋白和 动力学。但是，目前尚不清楚这两个生物电机如何合作以定位 货物并实现细胞功能。双向运输受损会导致多种 神经退行性疾病，包括阿尔茨海默氏病。微管相关蛋白 Map7和Dynein cofactor Lis1是两个重要的蛋白质，在货物中起关键作用 运输。虽然MAP7增强了驱动蛋白的进程，但Lis1可以缓解自动 抑制动力蛋白。在此提案中，我们旨在重建DDBE-K（Dynein，dynactin，bicd， EGL，动力素）复合物，并在两个MAP7的存在下观察其在微管轨道上的运动 使用总内反射荧光（TIRF）显微镜使用LIS1。鉴于Map7具有 能够延长微管上的驱动蛋白的附着，而Lis1更喜欢结合两种动力蛋白， 我们预测MAP7和LIS1将在确定运动方向方面发挥关键作用 微管上的复合物。使用单分子测定法，我们将研究是否 适配器蛋白BICD不仅充当货物和电机之间的接头，而且还可以发挥作用 在传感货物和电机结合中的重要作用。这些提出的研究将提供新颖 深入了解动力蛋白和货物适配器蛋白的作用，并有助于我们的理解 关于细胞质动力蛋白和驱动蛋白如何共同运输货物双向运输的 微管。