Nociceptor translational profiling reveals the RagA-mTORC1 network as a critical generator of neuropathic pain

Pain sensing neurons, nociceptors, are key drivers of neuropathic pain. We used translating ribosome affinity purification (trap) to comprehensively characterize up- and down-regulated mRNA translation in Scn10a-positive nociceptors in chemotherapy-induced neuropathic pain. We provide evidence that an underlying mechanism driving these changes in gene expression is a sustained mTORC1 activation driven by MNK1-eIF4E signaling. RagA, a GTPase controlling mTORC1 activity, is identified as a novel target of MNK1-eIF4E signaling, demonstrating a new link between these distinct signaling pathways. Neuropathic pain and RagA translation are strongly attenuated by genetic ablation of eIF4E phosphorylation, MNK1 elimination or treatment with the MNK inhibitor eFT508. We reveal a novel translational circuit for the genesis of neuropathic pain with important implications for next generation neuropathic pain therapeutics. Overall design: Examination of paired unbiased mRNA profile and mRNA profile based on eGFP + L10a (RPL10A) pulldown (TRAP assay) for normal mouse DRG (4 assays / replicates) and DRG from CIPN (Paclitaxel) model mouse (5 assays, 2 pooled to make 4 replicates) Associated NIH grants: R01NS065926 (TJP), R01NS098826 (TJP and GD), R01CA200263 (PMD) and R01NS100788 (ZTC)

伤害感受神经元（nociceptors）是神经病理性疼痛的关键驱动因子。本研究采用翻译核糖体亲和纯化（translating ribosome affinity purification, TRAP）技术，全面解析化疗诱导神经病理性疼痛模型中Scn10a阳性伤害感受神经元内mRNA翻译的上调与下调变化。研究证实，驱动此类基因表达改变的潜在机制为MNK1-eIF4E信号通路介导的持续哺乳动物雷帕霉素靶蛋白复合物1（mammalian target of rapamycin complex 1, mTORC1）激活。作为调控mTORC1活性的GTP酶（guanosine triphosphatase, GTPase），RagA被鉴定为MNK1-eIF4E信号通路的全新靶点，揭示了两条不同信号通路间的未知关联。通过敲除真核翻译起始因子4E（eukaryotic translation initiation factor 4E, eIF4E）的磷酸化位点、敲除丝裂原活化蛋白激酶相互作用激酶1（MAP kinase-interacting kinase 1, MNK1）或使用MNK抑制剂eFT508处理，均可显著减弱神经病理性疼痛及RagA的翻译水平。本研究揭示了一条介导神经病理性疼痛发生的全新翻译调控环路，为新一代神经病理性疼痛治疗药物的研发提供了重要参考。

整体实验设计：对配对的无偏倚mRNA表达谱与基于增强绿色荧光蛋白（enhanced green fluorescent protein, eGFP）+L10a（RPL10A）下拉纯化（TRAP实验）的mRNA表达谱进行检测，样本包括正常小鼠背根神经节（dorsal root ganglion, DRG，共4次重复实验）以及紫杉醇（Paclitaxel）诱导的化疗诱导神经病理性疼痛（chemotherapy-induced neuropathic pain, CIPN）模型小鼠的背根神经节（共5次实验，其中2份样本混合以构建4个重复样本）。

相关NIH资助项目包括：R01NS065926（TJP）、R01NS098826（TJP与GD）、R01CA200263（PMD）以及R01NS100788（ZTC）