Abstract
Microtubules establish the directionality of intracellular transport by kinesins and dynein through polarized assembly, but it remains unclear how directed transport occurs along microtubules organized with mixed polarity. We investigated the ability of the plus end-directed kinesin-4 motor KIF21B to navigate mixed polarity microtubules in mammalian dendrites. Reconstitution assays with recombinant KIF21B and engineered microtubule bundles or extracted neuronal cytoskeletons indicate that nucleotide-independent microtubule-binding regions of KIF21B modulate microtubule dynamics and promote directional switching on antiparallel microtubules. Optogenetic recruitment of KIF21B to organelles in live neurons induces unidirectional transport in axons but bidirectional transport with a net retrograde bias in dendrites. Removal of the secondary microtubule-binding regions of KIF21B or dampening of microtubule dynamics with low concentrations of nocodazole eliminates retrograde bias in live dendrites. Further exploration of the contribution of microtubule dynamics in dendrites to directionality revealed plus end-out microtubules to be more dynamic than plus end-in microtubules, with nocodazole preferentially stabilizing the plus end-out population. We propose a model in which both nucleotide-sensitive and -insensitive microtubule-binding sites of KIF21B motors contribute to the search and selection of stable plus end-in microtubules within the mixed polarity microtubule arrays characteristic of mammalian dendrites to achieve net retrograde movement of KIF21B-bound cargoes.