Regulation of Axonal Transport by the JNK (c-Jun N-terminal Kinase) Pathway

JNK（c-Jun N 末端激酶）途径对轴突运输的调节

• 批准号: 8219211
• 负责人: Meng-Meng Fu
• 金额: $ 4.22万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-01-01 至 2013-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8219211
• 关键词: Affect; Alzheimer' s Disease; Amyloid beta-Protein Precursor; Axon; Axonal Transport; Behavior; Binding; Cell Death Signaling Process; Cells; Cessation of life; Co-Immunoprecipitations; Complex; Data; Defect; Distal; Dominant-Negative Mutation; Dynein ATPase; Exhibits; Fluorescent Dyes; Injury; Integral Membrane Protein; Kinesin; LRRK2 gene; Label; Life; Link; Measures; Mediating; Microscopy; Molecular; Motor; Mutate; Nerve Degeneration; Neurodegenerative Disorders; Neuronal Injury; Neurons; Organelles; Parkinson Disease; Pathway interactions; Phosphorylation; Population; Post-Translational Protein Processing; Process; Property; Proteins; Regulation; Research; Role; Scaffolding Protein; Signal Pathway; Signal Transduction; Small Interfering RNA; Speed; Staging; Stress; Synapses; Testing; Transfection; Travel; Vesicle; anterograde transport; base; biological adaptation to stress; dynactin; insight; knock-down; mutant; neuronal cell body; phosphatase inhibitor; protein function; protein misfolding; public health relevance; research study; response; retrograde transport; stress-activated protein kinase 1; therapeutic target

DESCRIPTION (provided by applicant): Axonal transport in neurons is essential for the delivery of functional proteins to the synapse, for clearance of damaged or misfolded proteins, and for long-distance signaling from the distal axon to the cell body. However, regulation of axonal transport in terms cargo recognition and transport behavior (speed, direction, pausing) is poorly understood. This study proposes the idea that transport of organelles can be modulated by changes in intracellular signaling, such as activation of JNK (c-Jun N-terminal kinase), which has been implicated in stress response and neuronal death. My preliminary data from neuronal cultures shows that pharmacological inhibition of JNK inhibits vesicular transport in both directions and that activation of JNK increases the speed of retrograde transport. An excellent candidate for mediating these transport responses is JIP1 (JNK-interacting protein), which has the ability to associate with JNK, both anterograde and retrograde motor proteins, and vesicles (via transmembrane proteins). In addition, the preliminary observation that JNK activation also leads to phosphorylation of JIP1 supports the hypothesis that JNK-induced phosphorylation of JIP1 leads to changes in axonal transport by altering the interaction between JIP1 and motor proteins. To verify this hypothesis, the proposed experiments will use live-cell microscopy to characterize the effects of pharmacological manipulation of JNK signaling on the transport of specific fluorescently labeled vesicle populations. Also, siRNA knockdown experiments will be used to determine whether these JNK-induced changes are mediated by JIP1. Further, coimmunoprecipitations will more clearly elucidate the interaction between JIP1 and the retrograde motor complex, dynein/dynactin. Finally, mutant nonphosphorylatable JIP1 constructs will be used to determine whether JIP1 phosphorylation is the mechanism responsible for these JNK-induced changes in transport. These experiments will further the idea that axonal transport can be regulated at the cargo level via post-translational modification in response to signaling pathway activation. A better understanding of the connection between injury signaling and transport will provide new insight into the role of transport dysregulation in neurodegenerative diseases. PUBLIC HEALTH RELEVANCE: Neurons are specialized cells that contain long processes called axons along which electrical information travels. In order to maintain normal neuronal function, proteins and organelles must be transported at great distances along these axons to and from the cell body. This project investigates the role of an injury signaling pathway in regulating axonal transport properties, such as which cargos are transported and how fast they move, and will provide insight into understanding the role of axonal transport in neuronal injury and neurodegeneration.

描述（由申请人提供）：神经元中的轴突运输对于将功能蛋白递送到突触、清除受损或错误折叠的蛋白以及从远端轴突到细胞体的长距离信号传导至关重要。然而，人们对轴突运输在货物识别和运输行为（速度、方向、暂停）方面的调节知之甚少。这项研究提出了这样的想法：细胞器的运输可以通过细胞内信号传导的变化来调节，例如 JNK（c-Jun N 末端激酶）的激活，这与应激反应和神经元死亡有关。我从神经元培养中获得的初步数据表明，JNK 的药理学抑制会抑制双向的囊泡运输，并且 JNK 的激活会增加逆行运输的速度。介导这些转运反应的一个很好的候选者是 JIP1（JNK 相互作用蛋白），它能够与 JNK、顺行和逆行运动蛋白以及囊泡（通过跨膜蛋白）结合。此外，初步观察到JNK激活也会导致JIP1磷酸化，这支持了JNK诱导的JIP1磷酸化通过改变JIP1和运动蛋白之间的相互作用而导致轴突运输变化的假设。为了验证这一假设，拟议的实验将使用活细胞显微镜来表征 JNK 信号传导的药理学操作对特定荧光标记囊泡群体运输的影响。此外，siRNA 敲低实验将用于确定这些 JNK 诱导的变化是否是由 JIP1 介导的。此外，免疫共沉淀将更清楚地阐明 JIP1 和逆行运动复合体动力蛋白/动力蛋白之间的相互作用。最后，突变的不可磷酸化的 JIP1 构建体将用于确定 JIP1 磷酸化是否是导致这些 JNK 诱导的运输变化的机制。这些实验将进一步证明轴突运输可以通过响应信号通路激活的翻译后修饰在货物水平上进行调节。更好地了解损伤信号传导和转运之间的联系将为了解转运失调在神经退行性疾病中的作用提供新的见解。 公共健康相关性：神经元是一种特殊的细胞，包含称为轴突的长过程，电信息沿着轴突传播。为了维持正常的神经元功能，蛋白质和细胞器必须沿着这些轴突远距离运输至细胞体或从细胞体运输出去。该项目研究损伤信号通路在调节轴突运输特性中的作用，例如运输哪些货物以及它们移动的速度，并将深入了解轴突运输在神经元损伤和神经变性中的作用。