Visualizing Vesicle-Bound Kinesins in Cultured Hippocampal Neurons.

Abstract

Eukaryotic cells use microtubule-based vesicle transport to exchange molecules between compartments. Kinesin family members mediate all microtubule plus end-directed vesicle transport. Of the 45 kinesins expressed in humans, some 20 mediate microtubule plus-end directed vesicle transport. Here we describe a technique to visualize vesicle-bound kinesins in cultured hippocampal neurons. The method involves the expression of the vesicle-binding tail domain while minimizing the cytoplasmic pool. Using this approach drastically improves vesicle labeling compared to full-length kinesins. This tool is useful for systematically comparing the localization of different kinesins in the same cell type and for identifying cargo proteins that reside in vesicles moved by a specific kinesin family member. While we describe the assay in cultured hippocampal neurons, we expect it to be easily transferable to other eukaryotic cell types.