Dynein-Dynactin-NuMA clusters generate cortical spindle-pulling forces as a multi-arm ensemble

ELife
Masako OkumuraTomomi Kiyomitsu

Abstract

To position the mitotic spindle within the cell, dynamic plus ends of astral microtubules are pulled by membrane-associated cortical force-generating machinery. However, in contrast to the chromosome-bound kinetochore structure, how the diffusion-prone cortical machinery is organized to generate large spindle-pulling forces remains poorly understood. Here, we develop a light-induced reconstitution system in human cells. We find that induced cortical targeting of NuMA, but not dynein, is sufficient for spindle pulling. This spindle-pulling activity requires dynein-dynactin recruitment by NuMA's N-terminal long arm, dynein-based astral microtubule gliding, and NuMA's direct microtubule-binding activities. Importantly, we demonstrate that cortical NuMA assembles specialized focal structures that cluster multiple force-generating modules to generate cooperative spindle-pulling forces. This clustering activity of NuMA is required for spindle positioning, but not for spindle-pole focusing. We propose that cortical Dynein-Dynactin-NuMA (DDN) clusters act as the core force-generating machinery that organizes a multi-arm ensemble reminiscent of the kinetochore.

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Citations

Jul 10, 2018·ELife·Andrea Serra-Marques, Sophie Dumont
Nov 9, 2019·The Journal of Biological Chemistry·Brian Kuhlman
Mar 19, 2020·Open Biology·Francesca RizzelliMarina Mapelli
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Jan 25, 2022·Physiological Reviews·Peng TanYubin Zhou

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Methods Mentioned

BETA
PCR
confocal microscopy

Software Mentioned

Metamorph
NuMA
Prism
ClustalWS
Fiji
Photoshop
GraphPad
iLID

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