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: 10666545
• 负责人: JEFFERY L TWISS
• 金额: $ 39.89万
• 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/10666545
• 关键词: Acceleration; Adult; Affect; Affinity; Alleles; Appearance; Axon; Behavior; Binding; Binding Proteins; Bioinformatics; Biotinylation; Central Nervous System; Clinical; Collaborations; Complex; Cytoplasm; Data; Dendrites; Development; Elements; Environment; FMRP; Family; Funding; Growth; Heterogeneous-Nuclear Ribonucleoproteins; Hippocampus; HuD antigen; Human; Impairment; In Vitro; Injury; Knockout Mice; Knowledge; Link; Mediating; Memory impairment; Messenger RNA; MicroRNAs; Molecular; Mus; Natural regeneration; Nerve; Nerve Regeneration; Nervous System; Neurons; Nuclear; Peripheral Nerves; Peripheral Nervous System; Peripheral nerve injury; Physiological; Prefrontal Cortex; Protein Binding Domain; Protein Biosynthesis; Proteins; Proteome; RNA; RNA analysis; RNA-Binding Proteins; RNA-Protein Interaction; Recovery of Function; Regenerative capacity; Regulation; Regulon; Ribonucleoproteins; Rodent; Role; Seizures; Stimulus; Testing; Translating; Translational Regulation; Translations; Up-Regulation; Work; axon growth; axon injury; axon regeneration; cohort; exosome; extracellular; in vivo; link protein; mRNA Stability; mRNA Transcript Degradation; mRNA Translation; memory consolidation; neuron loss; neuronal cell body; neuronal growth; neurotransmission; particle; peripheral nerve regeneration; posttranscriptional; presynaptic; protein complex; response; response to injury; stoichiometry; transcriptome; translational impact

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.

概括 该申请询问局部 mRNA 稳定性如何改变轴突再生能力，重点关注 RNA结合蛋白[RBP] KHSRP的贡献。神经系统广泛利用后 调节细胞蛋白质组以响应细胞外刺激的转录机制 发育、功能和响应轴突损伤期间的生理环境。由于一个 mRNA 可以多次翻译成蛋白质，给定的 mRNA 可用于翻译的时间有多长 从该 mRNA 产生的蛋白质的量。 mRNA 的稳定性确实受到调节， 与稳定和不稳定不同 mRNA 的 RBP 相互作用，以及与 microRNA 的相互作用 针对某些退化目标。 mRNA 的翻译显然支持轴突再生，但我们 对于轴突 mRNA 的稳定性如何局部调节知之甚少。我们已经证明 RBP HuD（也称为 ELAVL4）和 KHSRP（也称为 FUBP2、MARTA1 和 ZBP2）竞争与神经元的结合 具有富含 AU 元件的 mRNA，其中 HuD 相互作用使靶标稳定，KHSRP 相互作用使靶标不稳定 mRNA。在分子水平上，这种相互作用受到 mRNA 与 HuD 或 KHSRP。我们 01-05 年的工作表明，KHSRP 的缺失会增加 KHSRP 靶标 mRNA 水平，导致 轴突和树突过度生长，损害海马体和前额皮质的记忆巩固， 并增加前额皮质和海马体的突触前活动。 KHSRP 表示为 成年后，我们惊讶地发现周围神经中轴突 KHSRP 水平在成年后迅速增加 受伤。轴突 KHSRP 的增加是通过其编码 mRNA 的轴突内翻译而发生的。我们的 初步数据表明 KHSRP 敲除小鼠表现出加速的神经再生 KHSRP 在再生中的轴突固有功能。根据这些观察，我们假设 轴突 KHSRP 通过调节局部 mRNA 稳定性来控制轴突再生速率。我们 将通过以下具体目标来检验这一假设：1）KHSRP 是否调节 PNS 轴突再生 通过神经元内在机制？ 2) 轴突 KHSRP 增加是否会通过以下方式限制轴突再生： 编码再生相关蛋白的轴突 mRNA 不稳定？ 3) KHSRP 的蛋白质 相互作用组影响其轴突内功能？完成这里的学习将开始填补这个 知识差距，重点关注轴突中启动的 RNA-蛋白质相互作用，这些相互作用可能影响 mRNA 的存活。 这将提供第一个 RBP 域特异性 RNA 调节子的亚细胞分析，并将带来第一个 系统评估 RNA 存活对周围神经再生的贡献。

项目成果

An Essential Postdevelopmental Role for Lis1 in Mice.

• DOI: 10.1523/eneuro.0350-17.2018
• 发表时间: 2018-01
• 期刊: eNeuro
• 影响因子: 3.4
• 作者: Hines TJ;Gao X;Sahu S;Lange MM;Turner JR;Twiss JL;Smith DS
• 通讯作者: Smith DS

Intra-axonal protein synthesis - a new target for neural repair?

轴突内蛋白质合成——神经修复的新靶点？

• DOI: 10.4103/1673-5374.191193
• 发表时间: 2016
• 期刊: Neural regeneration research
• 影响因子: 6.1
• 作者: Twiss,JefferyL;Kalinski,AshleyL;Sachdeva,Rahul;Houle,JohnD
• 通讯作者: Houle,JohnD

Neuroproteomics: How Many Angels can be Identified in an Extract from the Head of a Pin?

神经蛋白质组学：从针头提取物中可以鉴定出多少个天使？

• DOI: 10.1074/mcp.e116.057828
• 发表时间: 2016
• 期刊: Molecular & cellular proteomics : MCP
• 作者: Twiss,JefferyL;Fainzilber,Mike
• 通讯作者: Fainzilber,Mike

Intra-axonal mechanisms driving axon regeneration.

• DOI: 10.1016/j.brainres.2020.146864
• 发表时间: 2020-08-01
• 期刊: Brain research
• 影响因子: 2.9
• 作者: Smith TP;Sahoo PK;Kar AN;Twiss JL
• 通讯作者: Twiss JL

Intra-axonal translation of Khsrp mRNA slows axon regeneration by destabilizing localized mRNAs.

• DOI: 10.1093/nar/gkac337
• 发表时间: 2022-06-10
• 期刊: Nucleic acids research
• 影响因子: 14.9
• 作者: Patel P;Buchanan CN;Zdradzinski MD;Sahoo PK;Kar AN;Lee SJ;Vaughn LS;Urisman A;Oses-Prieto J;Dell'Orco M;Cassidy DE;Costa ID;Miller S;Thames E;Smith TP;Burlingame AL;Perrone-Bizzozero N;Twiss JL
• 通讯作者: Twiss JL