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: 8060866
• 负责人: Meng-Meng Fu
• 金额: $ 4.18万
• 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/8060866
• 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; 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传递到突触，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。但是，对轴突运输的调节术语识别和运输行为（速度，方向，暂停）的理解很少。这项研究提出了这样一个想法，即细胞器信号传导的变化可以调节细胞器的运输，例如JNK的激活（C-Jun N末端激酶），这与压力反应和神经元死亡有关。我来自神经元培养物的初步数据表明，对JNK的药理抑制作用抑制了囊泡转运的方向，而JNK的激活会增加逆行转运的速度。介导这些转运反应的极好候选者是JIP1（JNK相互作用蛋白），该蛋白具有与JNK的能力，同时和逆行运动蛋白和囊泡（通过跨膜蛋白）。此外，关于JNK激活的初步观察还导致JIP1的磷酸化支持以下假设：JNK诱导的JIP1的磷酸化会导致通过改变JIP1与运动蛋白之间的相互作用，从而导致轴突转运的变化。为了验证这一假设，提出的实验将使用活细胞显微镜来表征JNK信号的药理学操纵对特定荧光标记的囊泡群体运输的影响。同样，将使用siRNA敲低实验来确定这些JNK诱导的变化是否由JIP1介导。此外，共免疫沉淀将更清楚地阐明JIP1与逆行运动复合物，Dynein/dynactin之间的相互作用。最后，将使用突变的非磷酸化JIP1构建体来确定JIP1磷酸化是否是负责这些JNK诱导的运输变化的机制。这些实验将进一步的想法，即可以通过翻译后修饰可以在货物水平上调节轴突运输，以响应信号通路激活。更好地了解伤害信号和运输之间的联系将为转运失调在神经退行性疾病中的作用提供新的见解。 公共卫生相关性：神经元是专门的细胞，其中包含较长的轴突，沿着电气信息传播的轴突。为了维持正常的神经元功能，必须沿着这些轴突向往返细胞体的轴突远距离运输蛋白质和细胞器。该项目调查了损伤信号通路在调节轴突运输特性中的作用，例如哪些嘉戈斯的运输以及它们的移动速度，并将提供深入了解轴突转运在神经元损伤和神经变性中的作用。