JNK Signaling Mediates Glial Proliferation in the Regenerating Zebrafish Spinal Cord

Zebrafish possess the remarkable capacity to regenerate from spinal cord injuries that would leave mammals such as humans permanently paralyzed. Much research into zebrafish spinal cord regeneration has focused on identifying extracellular growth factors and matrix components which create a pro-regenerative environment; however, it is just as important to identify and understand the transcription factors which control pro-regenerative transcriptional responses within the resident stem cell population of the spinal cord, and the signaling cascades which translate the known extracellular ligands into cellular responses. Using CRISPR/Cas9, we generated two novel transcription factor knockout zebrafish lines which we tested for spinal cord regeneration defects and found no difference in regenerative capacity. Using a chemical screen, we identify JNK signaling as a necessary regulator of glial cell cycling and tissue bridging during spinal cord regeneration in larval zebrafish. With a kinase translocation reporter, we visualize and quantify JNK signaling dynamics at single-cell resolution in glial cell populations in developing larvae and during injury-induced regeneration. Glial JNK signaling is patterned in time and space during development and regeneration, decreasing globally as the tissue matures and increasing in the rostral cord stump upon transection injury. Thus, we present a tool to visualize signaling activity in the larval zebrafish spinal cord and demonstrate that dynamic JNK activity after spinal cord injury directs a proliferative response of glial cells during spinal cord regeneration.