The polycomb group protein Ezh2 promotes optic nerve regeneration

Abstract

Purpose : In mammals, neurons in the central nervous system (CNS) lose their axon growth abilities as they mature. On the contrary, mature neurons in the peripheral nervous system (PNS) still possess such an ability and can spontaneously regenerate axons upon axonal injury by initiating a regenerative response. Emerging evidence suggests that such a response largely involves epigenetic regulation of chromatin accessibility. Here we first characterized the role of Ezh2, a H3K27 methyltransferase, in PNS axon regeneration, and then tested if its modulation could induce CNS axon regeneration.

Methods : Western blot and immunostaining were used to compare protein levels. Dorsal root ganglion (DRG) neuronal culture and sciatic nerve regeneration were used as in vitro and in vivo PNS regeneration models, respectively. Optic nerve regeneration was used as an in vivo CNS axon regeneration model. Fluorescence-activated cell sorting was used to enrich RGCs from dissociated retinal cells. RNA-seq of RGCs was conducted to explore molecular mechanisms by which Ezh2 supports axon regeneration.

Results : Ezh2 was significantly upregulated in mouse lumbar 4 and 5 (L4/5) DRGs 3 days after sciatic nerve transection. Functionally, Ezh2 knockdown (KD) or knockout (KO) in cultured DRG neurons impaired axon growth in vitro. In consistence, KD or KO of Ezh2 in L4/5 DRGs or conditional KO of Ezh2 in sensory neurons impaired sensory axon regeneration after sciatic nerve crush. More importantly, we found that Ezh2 overexpression in RGCs significantly promoted optic nerve regeneration and RGC survival after optic nerve crush. Interestingly, the axon regeneration promoting effect of Ezh2 was not solely dependent on its methyltransferase function, as overexpression of a mutant form of Ezh2 lacking the methytransferase function also produced RGC axon regeneration, although to a lesser extent. RNA-seq revealed that mRNA levels of a large number of genes involved in synaptic transmission were downregulated by Ezh2 overexpression. Overexpression of Slc6a13 (encoding Gat2), a gene suppressed by Ezh2, partially blocked RGC axon regeneration induced by Ezh2 overexpression.

Conclusions : Our study demonstrated that Ezh2 was necessary for the successful spontaneous axon regeneration in the PNS, and that Ezh2 gain-of-function in RGCs could promote optic nerve regeneration by silencing synaptic transmission-associated genes.

This is a 2021 ARVO Annual Meeting abstract.