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(申请人提供)：神经元中的轴突运输对于将功能性蛋白质运送到突触、清除受损或错误折叠的蛋白质以及从远端轴突到细胞体的长距离信号传输是必不可少的。然而，在货物识别和运输行为(速度、方向、停顿)方面对轴突运输的调节知之甚少。这项研究提出细胞器的运输可以通过细胞内信号的变化来调节，例如激活与应激反应和神经元死亡有关的JNK(c-jun N-末端激酶)。我的神经元培养的初步数据显示，JNK的药理抑制抑制了囊泡的双向运输，而JNK的激活则增加了逆行运输的速度。JIP1(JNK相互作用蛋白)是介导这些运输反应的一个很好的候选者，它能够与JNK结合，包括顺行和逆行运动蛋白，以及囊泡(通过跨膜蛋白)。此外，JNK的激活也导致JIP1的磷酸化，这一初步观察支持了JNK诱导的JIP1的磷酸化通过改变JIP1与运动蛋白之间的相互作用而导致轴突运输改变的假设。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药物操纵JNK信号对特定荧光标记的囊泡群体运输的影响。此外，还将使用siRNA敲除实验来确定这些由JNK诱导的变化是否由JIP1介导。此外，免疫共沉淀将更清楚地阐明JIP1与逆行运动复合体dynein/dynactin之间的相互作用。最后，突变的非磷酸化JIP1结构将被用来确定JIP1磷酸化是否是导致JNK诱导的这些运输变化的机制。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。更好地理解损伤信号和运输之间的联系将为运输失调在神经退行性疾病中的作用提供新的见解。 与公共健康相关：神经元是一种特殊的细胞，它包含被称为轴突的长过程，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在很长的距离内往返于细胞体。该项目研究了损伤信号通路在调节轴突运输特性中的作用，如运输哪些货物以及它们移动的速度有多快，并将为理解轴突运输在神经元损伤和神经变性中的作用提供洞察力。