Axonally Synthesized β-Actin and GAP-43 Proteins Support Distinct Modes of Axonal Growth

Increasing evidence points to the importance of local protein synthesis for axonal growth and responses to axotomy, yet there is little insight into the functions of individual locally synthesized proteins. We recently showed that expression of a reporter mRNA with the axonally localizing beta-actin mRNA 3'UTR competes with endogenous beta-actin and GAP-43 mRNAs for binding to ZBP1 and axonal localization in adult sensory neurons (Donnelly et al., 2011). Here, we show that the 3'UTR of GAP-43 mRNA can deplete axons of endogenous beta-actin mRNA. We took advantage of this 3'UTR competition to address the functions of axonally synthesized beta-actin and GAP-43 proteins. In cultured rat neurons, increasing axonal synthesis of beta-actin protein while decreasing axonal synthesis of GAP-43 protein resulted in short highly branched axons. Decreasing axonal synthesis of beta-actin protein while increasing axonal synthesis of GAP-43 protein resulted in long axons with few branches. siRNA-mediated depletion of overall GAP-43 mRNA from dorsal root ganglia (DRGs) decreased the length of axons, while overall depletion of beta-actin mRNA from DRGs decreased the number of axon branches. These deficits in axon growth could be rescued by transfecting with siRNA-resistant constructs encoding beta-actin or GAP-43 proteins, but only if them RNAs were targeted for axonal transport. Finally, in ovo electroporation of axonally targeted GAP-43 mRNA increased length and axonally targeted beta-actin mRNA increased branching of sensory axons growing into the chick spinal cord. These studies indicate that axonal translation of beta-actin mRNA supports axon branching and axonal translation of GAP-43 mRNA supports elongating growth.