Contractile acto-myosin network on nuclear envelope remnants positions human chromosomes for mitosis
Preprint posted on 15 March 2019 https://www.biorxiv.org/content/10.1101/459750v3
Why I think this study is interesting
Microtubules are critical to every step of mitosis; without them, chromosomes cannot segregate to the future daughter nuclei. Yet recent studies have hinted at supporting roles for actin and myosin (1-3). This study employs innovative image analysis to show how actin and myosin forces help microtubules capture chromosomes quickly and accurately, highlighting the interdependency of these cytoskeletal systems.
During mitosis, the nuclear envelope disintegrates, allowing mitotic spindle microtubules to capture the chromosomes at their kinetochores (4-6). Spindle microtubules then align the chromosomes at the center of the spindle and separate them to form two identical daughter nuclei.
The spindle microtubules must search for chromosomes in three dimensions in a crowded environment. After the nuclear envelope disintegrates, the chromosomes can access the entire cell, increasing the search volume for spindle microtubules and reducing chromosome capture efficiency. This study aims to understand how cells tackle this problem and ensure timely and accurate chromosome segregation.
To quantify chromosome scattering, the authors measure the chromosome scattering volume (CSV) in live, dividing cells with fluorescently labled DNA. They use 3D images of the fluorescent chromosomes to define the smallest volume that contains all of the chromosomes (the CSV), which allows them to compare chromosome scattering across experimental conditions and timepoints.
Chromosome scatter decreases following nuclear envelope breakdown.
After envelope breakdown, the chromosome scattering volume (CSV) decreased quickly for 8 minutes, then continued to decrease for about 10 more minutes. Overall, the CSV decreased to about 60% of the value before envelope breakdown.
Chromosome condensation does not drive chromosome scatter reduction.
The authors measured total chromosome volume by outlining individual chromosomes instead of the total scatter volume. This value remained constant for 30 minutes after nuclear envelope breakdown, confirming previous findings that chromosome condensation occurs before nuclear envelope breakdown (7).
Microtubules do not drive chromosome scatter reduction.
Treatment with nocodazole to disrupt microtubule polymerization prevented chromosome alignment but did not affect overall CSV reduction. The CSV decreased faster at first in control cells; this could result from fully functional microtubules compressing envelope fragments.
The actin cytoskeleton and the nucleoskeleton coordinate chromosome scatter reduction.
Actin accumulates around the nucleus around 10 minutes prior to nuclear envelope breakdown and surrounds the chromosomes while CSV decreases. High-resolution Airyscan confocal microscopy revealed that the actin network forms just outside the nuclear envelope. The actin-encased volume shrinks to around 65% of its original volume by 6 minutes after envelope breakdown.
The LINC complex links actin in the cytoplasm to the nucleoskeleton. In cells expressing a dominant negative LINC complex, actin failed to accumulate around the nucleus before envelope breakdown. Although chromosome scatter decreased normally for around 8 minutes, reduction then tapered off. The CSV ultimately decreased half as much as in control cells.
Myosin II contractility reduces chromosome scatter.
Myosin II co-localizes with actin at the nuclear envelope prior to its breakdown. Just as with expressing dominant negative LINC, inhibiting myosin II motor activity with blebbistatin stops actin network accumulation/contraction and chromosome scatter reduction. To confirm the role of myosin, the authors used azidoblebbistatin, which covalently links to myosin upon exposure to infrared light. When they exposed one half of the nucleus to infrared light (inhibiting myosin), it did not contract, while the half of the nucleus did.
Actomyosin-driven chromosome scatter reduction ensures proper anaphase onset.
In cells expressing the dominant-negative LINC complex, the mitotic spindle microtubules assembled correctly but there was a considerable delay in chromosome congression (30 vs 64 minutes) and anaphase onset (40 vs 78 minutes). Many cells also had chromosomes behind the spindle poles, which further delayed anaphase.
The authors speculate that nuclear envelope breakdown reduces tension in the nuclear membrane (and the associated actin network), which may induce contraction of the envelope-associated actomyosin network to facilitate chromosome scatter reduction. Previous work has shown connections between cell membrane tension and actomyosin contractility (8, 9). Future studies could investigate how changes in nuclear membrane tension affect chromosome scatter reduction during mitosis (10).
How are the mechanisms described in this paper conserved across different cell types? The U2OS cells used in this study are nearly triploid. The authors found an actin network around the nucleus in dividing diploid RPE cells but found no similar network in highly aneuploid HeLa cells. Future work could investigate the relationship between actomyosin-induced chromosome scatter reduction, aneuploidy, and cancer.
1. P. Lenart et al., A contractile nuclear actin network drives chromosome congression in oocytes. Nature 436, 812-818 (2005).
2. M. Burdyniuk, A. Callegari, M. Mori, F. Nédélec, P. Lénárt, F-Actin nucleated onchromosomes coordinates their capture by microtubules in oocyte meiosis. The Journal of Cell Biology 217, 2661-2674 (2018).
3. B. Mogessie, M. Schuh, Actin protects mammalian eggs against chromosome segregation errors. Science 357, (2017).
4. T. U. Tanaka, Kinetochore-microtubule interactions: steps towards bi-orientation. The EMBO Journal 29, 4070-4082 (2010).
5. K. M. Godek, L. Kabeche, D. A. Compton, Regulation of kinetochore-microtubule attachments through homeostatic control during mitosis. Nature Reviews. Molecular Cell Biology 16, 57-64 (2015).
6. H. Maiato, A. M. Gomes, F. Sousa, M. Barisic, Mechanisms of Chromosome Congression during Mitosis. Biology 6, 122-177 (2017).
7. F. Mora-Bermudez, D. Gerlich, J. Ellenberg, Maximal chromosome compaction occurs by axial shortening in anaphase and depends on Aurora kinase. Nature Cell Biology 9, 822-831 (2007).
8. B. Pontes, P. Monzo, N. C. Gauthier, Membrane tension: A challenging but universal physical parameter in cell biology. Seminars in Cell & Developmental Biology 71, 30-41 (2017).
9. N. C. Gauthier, M. A. Fardin, P. Roca-Cusachs, M. P. Sheetz, Temporary increase in plasma membrane tension coordinates the activation of exocytosis and contraction during cell spreading. Proceedings of the National Academy of Sciences of the United States of America 108, 14467-14472 (2011).
10. B. Enyedi and P. Niethammer, Nuclear membrane stretch and its role in mechanotransduction. Nucleus 8(2), 156-161 (2017).
Posted on: 9 July 2019 , updated on: 10 July 2019Read preprint
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