Microtubule Polarity in Axons is Sorted by a Molecular Gradient of Dynactin

Abstract

In neuronal axons, almost all microtubules (MTs) are oriented with their growing end (+end) away from the cell body (+end out). Molecular motor proteins rely on this orientation to efficiently move cellular cargo to the far distal regions of the axon. Despite 30 years of research, the mechanism that establishes this unique MT configuration remains unknown. We here analysed MT growth behavior in Drosophila melanogaster neurons with a novel machine learning algorithm; KymoButler. With the unmatched precision of this approach we discovered that +end out MTs grow for longer times than -end out MTs. Together with an analytical model of microtubule growth, this observation predicted dramatic differences in average MT length, leaving -end out MTs are short and unstable. Additionally, we found evidence that the microtubule anti catastrophe factor p150 is responsible for the difference in growth times by promoting microtubule growth at the axon tip through a molecular gradient. These findings suggested a simple mechanism that organises axonal MTs. First, +end out MTs are stabilized by distally located p150. Subsequently, the short -end out MTs are transported out of the axon, depolymerize, or reorient, leaving only longer +end out MTs in the axon. Our results pave the way towards a deeper understanding of how the cytoskeleton in neurons orients to support molecular transport along the axon, potentially shedding new light on pathologies that are characterized by axonal transport deficiencies such as Alzheimer's disease.