Coordination between actin filaments and microtubules is critical to complete important steps during cell division. For instance, cytoplasmic actin filament dynamics play an active role in the off-center positioning of the spindle during metaphase I in mouse oocytes [1-3] or in gathering the chromosomes to ensure proper spindle formation in starfish oocytes [4, 5], whereas cortical actin filaments control spindle rotation and positioning in adherent cells or in mouse oocytes [6-9]. Several molecular effectors have been found to facilitate anchoring between the meiotic spindle and the cortical actin [10-14]. In vitro reconstitutions have provided detailed insights in the biochemical and physical interactions between microtubules and actin filaments [15-20]. Yet how actin meshwork architecture affects microtubule dynamics is still unclear. Here, we reconstituted microtubule aster in the presence of a meshwork of actin filaments using confined actin-intact Xenopus egg extracts. We found that actin filament branching reduces the lengths and growth rates of microtubules and constrains the mobility of microtubule asters. By reconstituting the interaction between dynamic actin filaments and microtubules in a minimal system based on purified proteins, we found that the branching of actin filaments is sufficient to block microtubule growth and trigger microtubule disassembly. In a further exploration of Xenopus egg extracts, we found that dense and static branched actin meshwork perturbs monopolar spindle assembly by constraining the motion of the spindle pole. Interestingly, monopolar spindle assembly was not constrained in conditions supporting dynamic meshwork rearrangements. We propose that branched actin filament meshwork provides physical barriers that limit microtubule growth.
Keywords: Xenopus egg extracts; actin; actin network architecture; actin-microtubule interaction; branched actin network; cell-free systems; microtubule.
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