REST-mediated epigenomic and transcriptomic signatures in neuropathic pain

项目总结 慢性神经性疼痛很难治疗，而且仍然是一个主要的临床问题，尽管许多临床和 临床前研究。我们的长期目标是确定治疗/预防慢性神经病的新目标 疼痛。越来越多的证据表明，表观遗传机制和通过microRNAs(MiRs)的调控是 与从急性疼痛到慢性疼痛的过渡有关。该项目的目标是确定REST-和G9a- 背根神经节相关转录调控网络和表观基因组调控网络及其在中枢神经系统中的作用 神经病理性疼痛的发展。转录抑制因子REST是主要的表观基因组调控因子。在……里面 神经损伤诱导的神经病理性疼痛，DRG神经元静息过度表达的表观遗传沉默 与OPRM1、Kcnd3、KCNQ2和scn10a基因等靶基因的抑制有关。组蛋白 甲基转移酶G9a也是包括REST在内的许多转录因子的表观基因组辅阻遏子。我们 最近发现，G9a在DRG神经元中几乎所有K通道基因的表观遗传沉默中起关键作用 在急慢性疼痛转变过程中，基因启动子上的组蛋白H3K9me2增加。相应地，老鼠 通过去除背根节中的G9a，神经元在神经损伤后不会出现慢性疼痛。因此，REST和G9a 提出合适的分子探针，以破译慢性疼痛的遗传和表观遗传基础。然而， 关于与REST和G9a相关的转录和表观基因组网络的知识有限 神经病理性疼痛的发展。此多PI资助是两个实验室之间的合作，具有互补性 专业知识。除了上述G9a鼠标模型外，这两个实验室还开发了一种创新的 由互补休息条件敲除(CKO)和新条件(New Conditions)组成的实验系统 人类静息过度表达(COE)小鼠模型。初步结果表明，尽管DRG特异性 REST CKO小鼠表现出神经损伤后减轻的疼痛超敏反应，DRG特异性REST COE小鼠表现出 疼痛过敏。因此，两种不同的小鼠模型概括了慢性疼痛的转变，如 预期基于现场工作，并提供一个体内系统来研究转录机制 慢性疼痛。在这里，我们提出了中心假设，即REST和G9a参与了 DRG神经损伤后慢性疼痛的发生通过独特的转录和表观基因组特征。如果 如果成功，我们提议的研究将识别DRG神经元中的所有REST和G9a靶点，包括mRNAs 和MIR，以及REST和G9a介导的表观基因组变化在慢性疼痛的发生发展中的作用 神经损伤。我们还将从功能上确定那些受REST和G9a监管的MIR，这些MIR可用于 阻断休息和G9a介导的慢性疼痛。我们将把我们的结果与化疗的结果进行比较- 引起慢性疼痛。总而言之，这项工作的结果可能会产生新的签名和 可能用于预测患者对慢性疼痛的易感性或抵抗力的生物标志物 发展。这一结果还可以用来确定治疗慢性疼痛的新的可操作药物靶点。

PROJECT SUMMARY Chronic neuropathic pain is difficult to treat and remains a major clinical problem despite numerous clinical and preclinical studies. Our long-term goal is to identify new targets for treatment/prevention of chronic neuropathic pain. Mounting evidence suggests that epigenetic mechanisms and regulation via microRNAs (miRs) are associated with the transition from acute to chronic pain. The goal of this project is to identify REST- and G9a- related transcriptomic and epigenomic regulatory networks in the dorsal root ganglion (DRG) and their roles in the development of neuropathic pain. The transcriptional repressor REST is a major epigenomic regulator. In nerve injury-induced neuropathic pain, epigenetic silencing by REST overexpression in DRG neurons has been implicated in the repression of targets such as Oprm1, Kcnd3, Kcnq2, and scn10a genes. Histone methyl-transferase G9a is also an epigenomic corepressor of many transcription factors including REST. We recently found that G9a is critical in the epigenetic silencing of almost all K+ channel genes in DRG neurons via increased histone H3K9me2 on the genes' promoters during acute-to-chronic pain transition. Accordingly, mice with G9a ablated from DRG neurons do not develop chronic pain after nerve injury. Thus, both REST and G9a present suitable molecular probes to decipher the genetic and epigenetic bases of chronic pain. However, knowledge is limited about the transcriptomic and epigenomic networks associated with REST and G9a in neuropathic pain development. This multiple-PI grant is a collaboration between two labs with complementary expertise. In addition to the G9a mouse model described above, the two labs have developed an innovative experimental system consisting of complementary Rest conditional knock-out (cKO) and new conditional human REST overexpression (cOE) mouse models. Preliminary results indicate that whereas DRG-specific Rest cKO mice show attenuated pain hypersensitivity after nerve injury, DRG-specific REST cOE mice exhibit pain hypersensitivity. Thus, the two contrasting mouse models recapitulate the chronic pain transition, as anticipated based on work in the field, and provide an in vivo system to study transcriptional mechanisms of chronic pain. Here we propose to the test the central hypothesis that REST and G9a are involved in the development of chronic pain after DRG nerve injury via unique transcriptomic and epigenomic signatures. If successful, our proposed research will identify all REST and G9a targets in DRG neurons including mRNAs and miRs, as well as REST- and G9a-mediated epigenomic changes in the development of chronic pain after nerve injury. We will also functionally identify those REST- and G9a-regulated miRs that could be utilized to block REST- and G9a-mediated chronic pain. We will compare our results to those obtained in chemotherapy- induced chronic pain. In summary, the results from this work are likely to generate new signatures and biomarkers that could potentially be utilized to predict patient susceptibility or resistance to chronic pain development. The results could also be used to identify new actionable drug targets in chronic pain.