3'UTR regulation of axon translation and optic nerve regeneration

In the adult mammalian central nervous system (CNS), failure of axon regeneration limits recovery after traumatic injury or in neurodegenerative disease. Local protein translation has been implicated in the regulation of axon growth in highly compartmentalized neurons. 3' untranslated regions (3'UTRs) of mRNAs play critical roles in RNA localization and modification. Here we studied the regulation of 3'UTRs in growth cone and axon regeneration.

Using fluorescence recovery after photobleaching (FRAP), we examined dynamic changes of mRNA 3'UTRs-related local translation in distal growth cones of primary neurons, initially comparing 3'UTRs from Gap43, normally localized to axons, and gamma-actin (Actg1), normally distributed to soma and axon. Local translation patterns in response to trophic factors and depolarization stimuli were analyzed, with or without translation inhibitor anisomycin. Adeno-associated viral vectors were used to express constitutively active Src with specific 3'UTRs after optic nerve injury in vivo. Axon growth and Src signaling were detected to identify function of 3'UTRs in axon regeneration. Statistical analysis was performed using one-way ANOVA or Kruskal-Wallis test, two-tailed unpaired t-test or Mann-Whitney test for data sets with different distributions.

We discovered different 3'UTRs led to differences in local translational regulation in growth cones in vitro, including in response to relevant signals such as brain-derived neurotrophic factor (BDNF), forskolin, ciliary neurotrophic factor (CNTF), depolarization and repolarization. In vivo, we found that addition of 3'UTRs from Gap43 or Actg1 to a construct expressing constitutively active Src, which normally regulates growth cone and axon elongation, increased Src mRNA and protein localization, and Src activity measured by phospho-FAK in the optic nerve and optic tract, and the 3'UTR of Gap43 promoted more long-distance axon regeneration after optic nerve injury.

Together these data enhance our understanding of the complexity of 3'UTR-mediated regulation of local axon translation, and point to potential therapeutic avenues for growth cone-related protein expression and local translation via 3'UTR manipulation.