Destabilization of axonal mRNAs by KHSRP complexes during axon regeneration

轴突再生过程中 KHSRP 复合物导致轴突 mRNA 不稳定

• 批准号: 10306001
• 负责人: JEFFERY L TWISS
• 金额: $ 40.74万
• 依托单位: UNIVERSITY OF SOUTH CAROLINA AT COLUMBIA
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-09-15 至 2026-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10306001
• 关键词: Address; Adult; Affect; Affinity; Alleles; Appearance; Axon; Behavior; Binding; Binding Proteins; Bioinformatics; Biotinylation; Clinical; Collaborations; Complex; Data; Dendrites; Development; Elements; Environment; FMRP; Family; Funding; Genetic Transcription; Genetic Translation; Growth; Heterogeneous-Nuclear Ribonucleoproteins; Hippocampus (Brain); HuD antigen; Impairment; In Vitro; Injury; Knockout Mice; Knowledge; Link; Mediating; Memory; Memory impairment; Messenger RNA; MicroRNAs; Molecular; Mus; Natural regeneration; Nerve; Nerve Regeneration; Nervous system structure; Neuraxis; Neurons; Nuclear; Peripheral; Peripheral Nerves; Peripheral nerve injury; Physiological; Prefrontal Cortex; Protein Binding Domain; Protein Biosynthesis; Proteins; Proteome; RNA; RNA Binding; RNA analysis; RNA-Binding Proteins; RNA-Protein Interaction; Recovery of Function; Regenerative capacity; Regulation; Regulon; Ribonucleoproteins; Rodent; Role; Seizures; Stimulus; Testing; Time; Translating; Translational Regulation; Translations; Up-Regulation; Work; axon growth; axon injury; axon regeneration; base; cohort; exosome; extracellular; in vivo; link protein; mRNA Stability; mRNA Transcript Degradation; memory consolidation; neuron loss; neuronal cell body; neurotransmission; particle; peripheral nerve regeneration; presynaptic; protein complex; response; response to injury; stoichiometry; transcriptome

SUMMARY This application asks how localized mRNA stability modifies axonal regeneration capacity, focusing on contributions of the RNA binding protein [RBP] KHSRP. The nervous system makes extensive use of post- transcriptional mechanisms to regulate cellular proteomes in response to extracellular stimuli and physiologic environments during development, function, & in response to axonal injury. Since one mRNA can be translated into protein many times over, how long a given mRNA is available for translation impacts the amount of protein generated from that mRNA. Stability of mRNAs is indeed regulated, with interactions with RBPs stabilizing & destabilizing different mRNAs, as well as interactions with microRNAs targeting some targets for degradation. Translation of mRNAs clearly supports axon regeneration, but we have little knowledge for how stability of axonal mRNAs is locally regulated. We have shown that the RBPs HuD (also called ELAVL4) and KHSRP (also called FUBP2, MARTA1, & ZBP2) compete for binding to neuronal mRNAs with AU-rich elements, where HuD interaction stabilizes and KHSRP interaction destabilizes target mRNAs. At the molecular level, this interaction is impacted by an mRNA’s affinity for binding to HuD or KHSRP. Our work over years 01-05 show that loss of KHSRP increases KHSRP target mRNA levels, causes excessive axonal and dendritic growth, impairs memory consolidation in hippocampus & prefrontal cortex, and increases presynaptic activity in prefrontal cortex and hippocampus. KHSRP is expressed into adulthood, and we surprisingly find that axonal KHSRP levels rapidly increase in peripheral nerves after injury. This increase in axonal KHSRP occurs through intra-axonal translation of its encoding mRNA. Our preliminary data indicate that KHSRP knockout mice show accelerated nerve regeneration pointing to axon-intrinsic functions for KHSRP in regeneration. Based on these observations, we hypothesize that axonal KHSRP controls the rates of axon regeneration through regulation of localized mRNA stability. We will test this hypothesis with the following specific aims: 1) Does KHSRP regulate PNS axon regeneration through a neuron intrinsic mechanism? 2) Does increased axonal KHSRP limit axon regeneration by destabilizing axonal mRNAs encoding regeneration-associated proteins? and 3) Does KHSRP’s protein interactome influence its intra-axonal functions? Completion of the studies here will begin to fill this knowledge gap by focusing on RNA-protein interactions initiated in axons that can affect mRNA survival. This will provide the first subcellular analyses of RBP domain-specific RNA regulons and will bring the first systematic assessment for contributions of RNA survival to peripheral nerve regeneration.

总结 该应用程序询问局部mRNA稳定性如何改变轴突再生能力，重点是 RNA结合蛋白[RBP] KHSRP的贡献。神经系统广泛使用后- 调节细胞蛋白质组响应细胞外刺激的转录机制， 生理环境中的发展，功能和轴突损伤的反应。由于一个mRNA 可以多次翻译成蛋白质，给定的mRNA可以翻译多久 从mRNA产生蛋白质的量。mRNA的稳定性确实受到调节， 与RBP的相互作用稳定和不稳定不同的mRNA，以及与microRNA的相互作用 瞄准一些目标进行降解。mRNA的翻译明显支持轴突再生，但我们 对轴突mRNA的稳定性如何局部调节知之甚少。我们已经证明， HuD（也称为ELAVL 4）和KHSRP（也称为FUBP 2、MARTA 1和ZBP 2）竞争结合神经元。 具有富含AU元件的mRNA，其中HuD相互作用稳定，KHSRP相互作用使靶不稳定 mRNA。在分子水平上，这种相互作用受到mRNA结合HuD或Hu的亲和力的影响。 KHSRP。我们在01-05年的研究表明，KHSRP的缺失增加了KHSRP靶mRNA的水平， 过度的轴突和树突生长，损害海马和前额皮质中的记忆巩固， 并增加前额叶皮层和海马体的突触前活动。KHSRP表达为 成年后，我们惊讶地发现，轴突KHSRP水平迅速增加后，周围神经 损伤轴突KHSRP的这种增加通过其编码mRNA的轴突内翻译发生。我们 初步数据表明，KHSRP基因敲除小鼠显示出加速的神经再生， 再生中KHSRP的轴突内在功能。基于这些观察，我们假设， 轴突KHSRP通过调节局部mRNA的稳定性来控制轴突再生的速率。我们 本研究将从以下几个方面来验证这一假说：1）KHSRP是否调控PNS轴突再生 通过神经元的内在机制？2)轴突KHSRP增加是否通过 破坏轴突基因编码再生相关蛋白的稳定性？和3）KHSRP的蛋白质 相互作用体影响其轴突内功能？完成这里的研究将开始填补这个 通过专注于轴突中启动的RNA-蛋白质相互作用，可以影响mRNA的存活。 这将提供RBP结构域特异性RNA调节子的第一个亚细胞分析，并将带来 系统评估RNA存活对周围神经再生的贡献。