Microtubules are required for the maintenance of planar cell polarity in monociliated floorplate cells.

The asymmetric localization of planar cell polarity (PCP) proteins is essential for the establishment of many planar polarized cellular processes, but the mechanisms that maintain these asymmetric distributions remain poorly understood. A body of evidence has tied oriented subapical microtubules (MTs) to the establishment of PCP protein polarity, yet recent studies have suggested that the MT cytoskeleton is later dispensable for the maintenance of this asymmetry. As MTs underlie the vesicular trafficking of membrane-bound proteins within cells, the requirement for MTs in the maintenance of PCP merited further investigation. We investigated the complex interactions between PCP proteins and the MT cytoskeleton in the polarized context of the floorplate of the zebrafish neural tube. We demonstrated that the progressive posterior polarization of the primary cilia of floorplate cells requires not only Vangl2 but also Fzd3a. We determined that GFP-Vangl2 asymmetrically localizes to anterior membranes whereas Fzd3a-GFP does not polarize on anterior or posterior membranes but maintains a cytosolic enrichment at the base of the primary cilium. Vesicular Fzd3a-GFP is rapidly trafficked along MTs primarily toward the apical membrane during a period of PCP maintenance, whereas vesicular GFP-Vangl2 is less frequently observed. Nocodazole-induced loss of MT polymerization disrupts basal body positioning as well as GFP-Vangl2 localization and reduces cytosolic Fzd3a-GFP movements. Removal of nocodazole after MT disruption restores MT polymerization but does not restore basal body polarity. Interestingly, GFP-Vangl2 repolarizes to anterior membranes and vesicular Fzd3a-GFP dynamics recover after multiple hours of recovery, even in the context of unpolarized basal bodies. Together our findings challenge previous work by revealing an ongoing role for MT-dependent transport of PCP proteins in maintaining both cellular and PCP protein asymmetry during development.

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