Localization and active transport of mRNA in axons of sympathetic neurons in culture

In neurons, the establishment and maintenance of distinct somatic, dendritic, and axonal domains has long been known to rely on regulated traffic of organelles and proteins. More recently, the local targeting of specific mRNAs has also been demonstrated, at least in dendrites, to provide a local supply of specific proteins. Here we set out to test directly for the presence of mRNA in axons of cultured chick sympathetic neurons, to examine their distribution during axonal outgrowth, to determine the reliance of this distribution on specific cytoskeletal elements, and to assess whether the axonal and somatic mRNA complements differ. Using fluorescent in situ hybridization, we found that sympathetic axons do contain poly(A+) mRNA along their length in a pattern that changes gradually as axons elongate, from an evenly dispersed punctate distribution with strong growth cone staining to a distribution focused at branch points, varicosities, and some growth cones. Selective perturbations of the cytoskeleton revealed that the presence of axonal mRNA was dependent on microtubules (MTs), but not actin filaments, indicating that mRNA transport and/or anchoring within the axon are active processes involving MTs. Finally, reverse transcription-PCR amplification of RNAs from the axonal and somatic compartments showed that beta-actin mRNA was present in both compartments, whereas mRNA encoding alpha-tubulin was restricted to the somatic compartment and entirely absent from the axons. Thus, the mRNA populations in the soma versus the axon are both quantitatively and qualitatively different, and these neurons are able to direct specific mRNAs to the axon.