Direct observation of microtubule pushing by cortical dynein in living cells

ABSTRACT Microtubules are under the influence of forces mediated by cytoplasmic dynein motors associated with the cell cortex. If such microtubules are free to move, they are rap-idly transported inside cells. Here we directly observe fluorescent protein–labeled cortical dynein speckles and motile microtubules. We find that several dynein complex subunits, in-cluding the heavy chain, the intermediate chain, and the associated dynactin subunit Dctn1 (also known as p150glued) form spatially resolved, dynamic speckles at the cell cortex, which are preferentially associated with microtubules. Measurements of bleaching and dissociation kinetics at the cell cortex reveal that these speckles often contain multiple labeled dynein heavy-chain molecules and turn over rapidly within seconds. The dynamic behavior of micro-tubules, such as directional movement, bending, or rotation, is influenced by association with dynein speckles, suggesting a direct physical and functional interaction. Our results support a model in which rapid turnover of cell cortex–associated dynein complexes facilitates their search to efficiently capture and push microtubules directionally with leading plus ends