Unrestrained growth of correctly oriented microtubules instructs axonal microtubule orientation

In many eukaryotic cells, directed molecular transport occurs along microtubules. Within neuronal axons, transport over vast distances particularly relies on uniformly oriented microtubules, whose plus-ends point towards the distal axon tip (+end out). However, axonal microtubules initially have mixed orientations, and what breaks this orientation symmetry is poorly understood. Using live imaging of primary Drosophila melanogaster neurons and physical modelling, we found that +end out microtubules surpass a growth transition and undergo persistent long-term growth near the advancing axon tip. In contrast, oppositely oriented microtubules remain short. Using experimental perturbations, we confirmed that the enhanced growth of +end out microtubules is critical for achieving uniform microtubule orientation. Computer simulations of axon development incorporating our data returned +end out microtubules along the entire axonal length, suggesting that the accelerated growth kinetics of correctly oriented microtubules instructs overall axonal microtubule orientation. Our study thus leads to a holistic explanation of how axonal microtubules orient uniformly, a prerequisite for efficient long-range transport essential for neuronal functioning.