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bioRxiv · 10.1101/2022.11.07.515476

Kinetochore microtubules flux poleward along fixed centrosome-anchored microtubules during the metaphase of C. elegans one-cell embryo.

Soler, N.·Chesneau, L.·Bouvrais, H.·Pastezeur, S.·Le Marrec, L.·Pecreaux, J.
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Abstract

The mitotic spindle, a key structure to partition chromosomes during cell division, connects its poles to the chromosomes through microtubules. Their plus-ends, oriented towards the chromosomes, exhibit dynamic instability crucial for kinetochore correct attachments. Involved in this process, the poleward flux implicates the displacement of microtubules towards the spindle poles, coordinated with polymerisation at the plus ends. The mechanisms behind this are diverse. It includes treadmilling powered by microtubule depolymerisation at the spindle poles, sliding of spindle microtubules by molecular motors like Kinesin-5, and pushing microtubules away from the chromosomes by chromokinesins. Interestingly, no such flux was reported in the Caenorhabditis elegans zygote, although all proteins contributing to flux in mammals have homologs in the nematode. To explore this, we fluorescently labelled microtubules and conducted photobleaching. We found no global poleward flux; the bleached zones edges moved inward. The centrosome-side front motion was caused by dynamic instability, while the chromosome-side front exhibited faster recovery, suggesting an additional mechanism. This larger slope was detected only near the chromosomes, indicating that only kinetochore microtubules undergo flux. Consistently, this flux depended on proteins ensuring the chromosome attachment and growth of the kinetochore microtubules, notably NDC-80, CLS-2CLASP, and ZYG-9XMAP215. Furthermore, this flux decreased as metaphase progressed and attachments transitioned from side-to end-on; it was reduced by SKA-1 recruitment. Treadmilling was unlikely to account for these observations, as most kinetochore microtubules do not reach spindle poles in the zygote spindle. Conversely, the depletion of kinesin-12 KLP-18KIF15, which cross-links and focuses microtubules at meiosis, reduced the front rate. Ultimately, we propose that the sole kinetochore microtubules slide along spindle microtubules, likely powered by KLP-18, contrasting with solid displacement in other systems. It aligns with observations in human cells of decreasing flux with increasing chromosome distance.

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Soler, N., Chesneau, L., Bouvrais, H., Pastezeur, S., Le Marrec, L., Pecreaux, J.. 2022-11-07. Kinetochore microtubules flux poleward along fixed centrosome-anchored microtubules during the metaphase of C. elegans one-cell embryo.. https://doi.org/10.1101/2022.11.07.515476

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