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bioRxiv · 10.64898/2026.01.26.701783

Microtubule occupancy at kinetochores links checkpoint silencing with mitotic memory

Soares-de-Oliveira, J.·Okada, N.·Kletter, T.·Weiler, E. S.·Pereira, A. J.·Maiato, H.
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Abstract

The spindle assembly checkpoint (SAC) promotes faithful chromosome segregation by delaying mitosis until all kinetochores attach to spindle microtubules. Paradoxically, a p53-dependent memory mechanism--the mitotic stopwatch" --blocks daughter cell proliferation after unusually prolonged mitoses. How the SAC coordinates with the mitotic stopwatch remains unknown. Here, we found that microtubule occupancy at kinetochores is a cornerstone linking SAC silencing with mitotic memory. By combining live-cell with super-resolution microscopy, FRAP, laser microsurgery, biochemistry and molecular perturbations in Indian muntjac fibroblasts, we show that SAC silencing at kinetochores is gradual, non-uniform and confined to highly-localized microtubule attachments. Augmin promotes timely SAC silencing with high microtubule occupancy at kinetochores, whereas MPS1/CDK1 inhibition bypasses this requirement. Conversely, low microtubule occupancy delays SAC silencing, increases segregation errors and blocks daughter cell proliferation due to mitotic stopwatch surveillance. Thus, timely SAC silencing with high microtubule occupancy avoids "bad memories" of mitosis to allow daughter cell proliferation.

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Soares-de-Oliveira, J., Okada, N., Kletter, T., Weiler, E. S., Pereira, A. J., Maiato, H.. 2026-01-28. Microtubule occupancy at kinetochores links checkpoint silencing with mitotic memory. https://doi.org/10.64898/2026.01.26.701783

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