Dual leucine zipper kinase is necessary for retinal ganglion cell axonal regeneration in Xenopus laevis.

Retinal ganglion cell (RGC) axons of the African clawed frog, Xenopus laevis, unlike those of mammals, are capable of regeneration and functional reinnervation of central brain targets following injury. Here, we describe a tadpole optic nerve crush (ONC) procedure and assessments of brain reinnervation based on live imaging of RGC-specific transgenes which, when paired with CRISPR/Cas9 injections at the one-cell stage, can be used to assess the function of regeneration-associated genes in vivo in F0 animals. Using this assay, we find that map3k12, also known as dual leucine zipper kinase (Dlk), is necessary for RGC axonal regeneration and acts in a dose-dependent manner. Loss of Dlk does not affect RGC innervation of the brain during development or visually driven behavior but does block both axonal regeneration and functional vision restoration after ONC. Dlk loss does not alter the acute changes in mitochondrial movement that occur within RGC axons hours after ONC but does completely block the phosphorylation and nuclear translocation of the transcription factor Jun within RGCs days after ONC; yet, Jun is dispensable for reinnervation. These results demonstrate that in a species fully capable of regenerating its RGC axons, Dlk is essential for the axonal injury signal to reach the nucleus but may affect regeneration through a different pathway than by which it signals in mammalian RGCs.

The basic science of optic nerve regeneration.

Diverse insults to the optic nerve result in partial to total vision loss as the axons of retinal ganglion cells are destroyed. In glaucoma, axons are injured at the optic nerve head; in other optic neuropathies, axons can be damaged along the entire visual pathway. In all cases, as mammals cannot regenerate injured central nervous system cells, once the axons are lost, vision loss is irreversible. However, much has been learned about how retinal ganglion cells respond to axon injuries, and many of these crucial discoveries offer hope for future regenerative therapies. Here we review the current understanding regarding the temporal progression of axonal degeneration. We summarize known survival and regenerative mechanisms in mammals, including specific signaling pathways, key transcription factors, and reprogramming genes. We cover mechanisms intrinsic to retinal ganglion cells as well as their interactions with myeloid and glial cell populations in the retina and optic nerve that affect survival and regeneration. Finally, we highlight some non-mammalian species that are able to regenerate their retinal ganglion cell axons after injury, as understanding these successful regenerative responses may be essential to the rational design of future clinical interventions to regrow the optic nerve. In the end, a combination of many different molecular and cellular interventions will likely be the only way to achieve functional recovery of vision and restore quality of life to millions of patients around the world.