Ran GTPase regulates non-centrosomal microtubule nucleation and is transported by actin waves towards the neurite tip

Microtubule (MT) is the most abundant cytoskeleton in neurons and controls multiple facets of their development. While the organizing center of MTs in mitotic cells is typically located at the centrosome, MT nucleation in post-mitotic neurons switches to non-centrosomal sites. A handful of proteins and organelle have been shown to promote non-centrosomal MT formation in neurons, yet the regulation mechanism remains unknown. Here we demonstrate that the small GTPase Ran is a key regulator of non-centrosomal MT nucleation in neurons. The GTP-bound Ran (RanGTP) localizes to the neurite tips and around the soma. Using the RanGTP- and RanGDP-mimic mutants, we show that RanGTP promotes MT nucleation at the tip of the neurite. To demonstrate that RanGTP can promote MT nucleation in regions other than the neurite tip, an optogenetic tool called RanTRAP was constructed to enable light-induced local production of RanGTP in the neuronal cytoplasm. An increase of non-centrosomal MT nucleation can be observed by elevating the RanGTP level along the neurite using RanTRAP, establishing a new role for Ran in regulating neuronal MTs. Additionally, the mechanism of RanGTP enrichment at the neurite tip was examined. We discovered that actin waves drive the anterograde transport of RanGTP towards the neurite tip. Pharmacological disruption of actin waves abolishes the enrichment of RanGTP and reduces the non-centrosomal MT nucleation at the neurite tip. These observations provide a novel regulation mechanism of MTs and an indirect connection between the actin and MT cytoskeletons in neurons.