Growth cone-localized microtubule organizing center establishes microtubule orientation in dendrites

Abstract

A polarized arrangement of neuronal microtubule arrays is the foundation of membrane trafficking and subcellular compartmentalization. Conserved among both invertebrates and vertebrates, axons contain exclusively 'plus-end-out' microtubules while dendrites contain a high percentage of 'minus-end-out' microtubules, the origins of which have been a mystery. Here we show that in Caenorhabditis elegans the dendritic growth cone contains a non-centrosomal microtubule organizing center, which generates minus-end-out microtubules along outgrowing dendrites and plus-end-out microtubules in the growth cone. RAB-11-positive endosomes accumulate in this region and co-migrate with the microtubule nucleation complex γ-TuRC. The MTOC tracks the extending growth cone by kinesin-1/UNC-116-mediated endosome movements on distal plus-end-out microtubules and dynein clusters this advancing MTOC. Critically, perturbation of the function or localization of the MTOC causes reversed microtubule polarity in dendrites. These findings unveil the endosome-localized dendritic MTOC as a critical organelle for establishing axon-dendrite polarity.

Proximity labeling at non-centrosomal microtubule-organizing centers reveals VAB-10B and WDR-62 as distinct microtubule regulators

“…As human and mouse WDR62 are known to play prominent roles in neuronal development (Shohayeb et al, 2018), we investigated the localization of WDR-62 in the developing PVD neuron, a highly branched nocioceptor. Intriguingly, we found that WDR-62 localizes to the tip of the outgrowing PVD dendrite (arrowhead, Figure 2F), a site that was recently shown to contain a migrating ncMTOC enriched for microtubules and ɣ-TuRC ( Figure 2F) (Liang et al, 2020). Our discovery of WDR-62 in many polarized cell types provided a unique opportunity to study its role beyond the centrosome.…”

“…The ncMTOC could be a phase-separated structure, as PTRN-1 and several PTRN-1 proximity interactors contain intrinsically disordered regions. Alternatively, and consistent with the colocalization of displaced microtubules with membrane puncta in VAB-10 gut (-) embryos, membranous structures could serve as a platform for ncMTOC assembly as Golgi, Golgi outposts, and endosomes have all been shown to associate with non-centrosomal microtubules (Ori-McKenney et al, 2012;Efimov et al, 2007;Hehnly & Doxsey 2014;Liang et al, 2020). Thus PAR-3 and other polarity regulators could merely position an otherwise autonomous structure.…”

“…F) Live imaging of WDR-62::ZF::GFP (green) in the developing PVD neuron (magenta). Arrowhead indicates WDR-62 localization to the presumptive ncMTOC(Liang et al 2020), as represented in adjacent cartoon.…”

Microtubules originate asymmetrically at the somatic golgi and are guided via Kinesin2 to maintain polarity within neurons

“…Our observation that endogenously-tagged γ-tubulin localises to branchpoints supports this conclusion. Moreover, it was recently shown that an endosome-based MTOC tracks the growing dendritic growth cone within C. elegans PVD neurons and nucleates microtubules that travel back towards the soma ( Liang et al, 2020 ). The authors also suggest that a similar process may occur in newly developing Drosophila class I neurons.…”

“…Nevertheless, microtubules continue to grow within the soma ( Nguyen et al, 2011 ; Sánchez-Huertas et al, 2016 ), and in mammalian neurons this depends in part on the HAUS complex ( Sánchez-Huertas et al, 2016 ), which is also important for microtubule growth within axons and dendrites ( Cunha-Ferreira et al, 2018 ; Sánchez-Huertas et al, 2016 ). Some MTOCs have been identified within dendrites: the basal body, or its surrounding region, within the distal part of C. elegans ciliated neurons acts as an MTOC, as does a similar region within the URX non-ciliated neuron ( Harterink et al, 2018 ); an MTOC made from endosomes that tracks the dendritic growth cone in C. elegans PVD neurons has recently been identified ( Liang et al, 2020 ); and fragments of Golgi called Golgi outposts within the dendrites of Drosophila dendritic arborisation (da) neurons are thought to recruit γ-TuRCs and act as MTOCs ( Ori-McKenney et al, 2012 ; Yalgin et al, 2015 ; Zhou et al, 2014 ).…”