Role of Stress Granule Protein Aggregation in Axon Regeneration

周围神经可以自发再生，但轴突生长速度极其缓慢，以至于 靶点的完全功能再支配在人类身上很少实现。轴突再生在 中枢神经系统甚至更糟糕，以至于患有脊髓损伤(SCI)的人 几乎总是在病变平面以下永久丧失感觉和运动功能。 迫切需要加速周围神经系统中的轴突再生，并 增加中枢神经系统中的轴突再生。我们的研究项目集中在轴突上 再生的内在机制。轴突内合成的蛋白质支持轴突生长 发育中的神经元。我们已经证明三叉神经节神经元保持合成蛋白质的能力。 它们的轴突和这些蛋白质支持受损轴突的生长。培养神经元的轴突含有 数以千计的mRNAs--有几条证据表明中枢神经系统中存在复杂的mRNAs群体 体内的轴突和脊髓轴突包含mRNAs和翻译机制，当鼓励 用允许的底物再生。尽管 2000年初以来的显著进展‘S，决定时间和地点的分子机制 一种特定的mRNA在轴突中的翻译在很大程度上仍不清楚。这一级别的监管是 对调节轴突生长能力至关重要。我们已经证明了mRNAs存储在PNS轴突中 包含应激颗粒蛋白G3BP1的RNA-蛋白质聚集体。G3BP1蛋白可以驱动 应激颗粒聚集，G3BP1磷酸化阻止应激颗粒组装。不像 经典定义的应激颗粒，轴突G3PB1蛋白在未损伤/功能状态下显示聚集 三叉神经节轴突。这些轴突G3BP1集合体在轴突切断后迅速增加，但降至以下 在此之后不久，基础水平和相应的磷酸化G3BP1增加。G3BP1结合 到轴突中的mRNAs并减弱它们的翻译。我们已经发现了外源因子和 触发轴突G3BP1聚合体分解的内源性信号。外源因子 特异性增加轴突蛋白质合成，加快轴突生长速度 活着。这些观察结果使我们假设，压力的生理聚集 轴突颗粒蛋白通过阻断翻译抑制损伤的三叉神经节和中枢神经系统轴突生长 轴突信使核糖核酸组。我们将通过以下具体目标来检验这一假设： 目的1-通过抑制G3BP1功能促进轴突生长。 目的2-轴突G3BP1聚集解体的内源性机制。 目的3-轴突G3BP1聚集体解体时驱动轴突生长的机制。 轴突翻译的功能作用现已曝光，我们有确凿的活体证据 这一机制可以靶向促进急性周围神经损伤后轴突的生长。 拟议研究的完成将为时间监管的机制带来新的见解 轴突损伤和再生中轴突mRNA翻译的研究及探索新的治疗策略 神经修复。

Peripheral nerves spontaneously regenerate but the axon growth rate is abysmally slow, such that complete functional reinnervation of targets is rarely achieved in humans. Axon regeneration in the central nervous system is even worse, such that individuals with spinal cord injury (SCI) almost invariably have permanent lose of sensory and motor functions below the level of the lesion. There is a pressing need to accelerate axon regeneration in the peripheral nervous system and increase axon regeneration in the central nervous system. Our research program focuses on axon intrinsic mechanisms of regeneration. Intra-axonally synthesized proteins support axon growth in developing neurons. We have shown that PNS neurons retain the capacity to synthesize proteins in their axons and these proteins support growth of injured axons. Axons of cultured neurons contain thousands of mRNAs – and several lines of evidence point to complex populations of mRNAs in CNS axons in vivo and spinal cord axons contain mRNAs and translational machinery when encouraged to regenerate with permissive substrates. Despite remarkable advances since the early 2000’s, the molecular mechanisms that determine when and where a specific mRNA is translated in axons remain largely unknown. This level of regulation is critical for regulating axon growth capacity. We have shown that mRNAs are stored in PNS axons in RNA-protein aggregates that contain the stress granule protein G3BP1. G3BP1 protein can drive stress granule aggregation, and G3BP1 phosphorylation blocks stress granule assembly. Unlike the classically defined stress granule, axonal G3PB1 protein shows aggregation in uninjured/functioning PNS axons. These axonal G3BP1 aggregates rapidly increase after axotomy, but decrease to below basal levels shortly thereafter with a corresponding increase in phosphorylated G3BP1. G3BP1 binds to mRNAs in axons and attenuates their translation. We have discovered exogenous agents and endogenous signals that trigger disassembly of axonal G3BP1 aggregates. The exogenous agents specifically increase axonal protein synthesis and accelerate axon growth rates in vitro and in vivo. These observations have led us to hypothesize that physiological aggregation of stress granule proteins in axons attenuates axon growth in the injured PNS and CNS by blocking translation of an axonal mRNA cohort. We will test this hypothesis with the following specific aims: Aim 1 – Promotion of axon growth by inhibition of G3BP1 function. Aim 2 – Endogenous mechanisms for axonal G3BP1 aggregate disassembly. Aim 3 – Mechanisms driving axon growth upon disassembly of axonal G3BP1 aggregates. Functional roles for axonal translation have now come to light and we have solid in vivo evidence that this mechanism can be targeted to accelerate axon growth after acute peripheral nerve injury. Completion of the proposed research will bring new insight into mechanisms for temporal regulation of axonal mRNA translation in axon injury & regeneration and uncover new therapeutic strategies for neural repair.