Combinatorial proteomics and transcriptomics identify AMPK in the control of the axonal regeneration programme of DRG sensory neurons after spinal injury

Regeneration after injury occurs in axons that lie in the peripheral nervous system but it fails in the central nervous system limiting functional recovery. Despite recent progress, the signalling response to injury of peripheral versus central projecting axons that might underpin this differential regenerative ability is currently largely uncharacterized. To fill this knowledge gap, here we combined axoplasmic proteomics from peripheral sciatic or central projecting dorsal root axons from sciatic DRG neurons. Proteomics was combined with cell body RNAseq to compare axonal and soma responses between a spinal regeneration-incompetent versus sciatic regeneration-competent nerve injury. This allowed the identification of injury-dependent signalling pathways uniquely represented in peripheral versus central projecting sciatic DRG axons. RNAseq and proteomics analysis suggested AMPK as a putative regulator of axonal regenerative signalling pathways. AMPK immunoprecipitation followed by mass spectrometry from DRG suggested that the 26S proteasome and its regulatory subunit PSMC5 preferentially interact with AMPKα for proteasomal degradation following sciatic axotomy. Mechanistically, we found that the proteasome and CaMKIIα-dependent proteasomal subunit PSMC5 regulates AMPKα1 protein expression. Finally, conditional deletion of AMPKα1 promoted multiple regenerative signalling pathways and robust axonal growth across the injured spinal cord, suggesting inhibition of AMPK as a novel regenerative target following spinal injury.