Histone methyltransferase Ezh2 coordinates mammalian axon regeneration via regulation of key regenerative pathways (RNA-Seq I)

Current treatments for neurodegenerative diseases and neural injuries fall short of success. One primary reason is that neurons in the mammalian central nervous system (CNS) lose their regeneration ability as they mature. Here, we investigated the role of Ezh2, a histone methyltransferase, in regulation of mammalian axon regeneration. We found that Ezh2 declined in the mouse nervous system during maturation but was upregulated in adult dorsal root ganglion neurons to support spontaneous axon regeneration following peripheral nerve injury. In addition, overexpression of Ezh2 in retinal ganglion cells in the CNS promoted optic nerve regeneration via both histone methylation-dependent and -independent mechanisms. Further investigation revealed that Ezh2 supported axon regeneration by systematically silencing the transcription of genes regulating synaptic function and inhibiting axon regeneration, while simultaneously activating various axon regeneration promoting factors. In particular, our study demonstrated that the GABA transporter 2 encoded by the gene Slc6a13 acted downstream of Ezh2 to control axon regeneration. Our study suggested that modulating chromatin accessibility was a promising strategy to promote CNS axon regeneration. Overall design: To study how Ezh2 supports axon regeneration of retinal ganglion cells, we profiled the transcriptomic changes in retinal ganglion cells induced by Ezh2 overexpression with RNA-seq. We intravitreally injected AAV2-GFP (control) or AAV2-Ezh2 and crushed the optic nerve after two weeks. Three days after the optic nerve crush, retinal ganglion cells were enriched from dissociated retinal cells.

当前针对神经退行性疾病与神经损伤的治疗手段均难以取得理想疗效。究其核心原因之一，在于哺乳动物中枢神经系统（central nervous system, CNS）内的神经元在成熟后会丧失再生能力。本研究聚焦组蛋白甲基转移酶（histone methyltransferase）Ezh2在哺乳动物轴突再生调控中的作用。我们发现，小鼠神经系统发育成熟过程中Ezh2表达水平会下调，但在成年背根神经节神经元中Ezh2则被上调，以支持外周神经损伤后自发的轴突再生。此外，在中枢神经系统的视网膜神经节细胞中过表达Ezh2，可通过组蛋白甲基化依赖与非依赖双重机制促进视神经再生。进一步研究揭示，Ezh2通过系统性沉默调控突触功能、抑制轴突再生的基因的转录，同时激活多种促轴突再生因子，从而发挥支持轴突再生的作用。尤为重要的是，本研究证实由基因Slc6a13编码的γ-氨基丁酸转运体2（GABA transporter 2）可作为Ezh2的下游靶点参与调控轴突再生。本研究表明，调控染色质开放性是促进中枢神经系统轴突再生的极具潜力的治疗策略。
整体实验设计：为探究Ezh2如何支持视网膜神经节细胞的轴突再生，我们通过RNA测序（RNA-seq）分析了Ezh2过表达诱导的视网膜神经节细胞转录组变化。我们通过玻璃体内注射AAV2-GFP（对照组）或AAV2-Ezh2，于两周后对视神经进行钳夹损伤。视神经钳夹损伤三天后，我们从解离的视网膜细胞中富集得到视网膜神经节细胞。