Naslov
The chirality of the mitotic spindle provides a mechanical response to forces and depends on microtubule motors and crosslinkers

Autori
Trupinić, Monika ; Kokanović, Barbara ; Ponjavić, Ivana ; Ivec, Arian ; Tolić, Iva M.

Vrsta, podvrsta i kategorija rada
Sažeci sa skupova, neobjavljeni rad, znanstveni

Skup
9th World Congress of Biomechanics

Mjesto i datum
Taipei, Tajvan, 10.07.2022. - 14.07.2022

Vrsta sudjelovanja
Predavanje

Vrsta recenzije
Nije recenziran

Ključne riječi
Mitotic spindle ; Mitosis ; Chirality ; Twist ; Torques ; Rotation ; Motor proteins ; Kinesins ; Augmin ; Spindle compression

Sažetak
Introduction: Forces produced by motor proteins and microtubule dynamics within the mitotic spindle are crucial for proper chromosome segregation1. In addition to linear forces, rotational forces or torques are present in the spindle, reflected in the left- handed twisted shapes of microtubule bundles that make the spindle chiral2. However, the biological role and molecular origins of spindle chirality are unknown. Results: By developing methods for measuring spindle twist, we show that spindles are most chiral near the metaphase-to-anaphase transition. To assess the role of chirality in maintaining spindle robustness under force, we compressed the spindles along their axis. This resulted in stronger left- handed twist with a 2.3-fold increase, suggesting that the twisted shape allows for a mechanical response to forces. Inhibition or depletion of motor proteins that perform chiral stepping and localize to antiparallel microtubule overlaps, Eg5/kinesin-5 or Kif18A/kinesin-8, decreased the left-handed twist to -0.47 ± 0.14 °/μm in HeLa and -0.06 ± 0.19 °/μm in RPE1 cells or led to right- handed twist of 0.11 ± 0.14 °/μm in HeLa and 0.28 ± 0.13 °/μm in RPE1 cells, respectively, implying that these motors regulate the twist by rotating antiparallel microtubules around one another. Right-handed twist was also observed after depletion of the microtubule crosslinker PRC1 in RPE1 cells (0.21 ± 0.13 °/μm) or the nucleator augmin in both HeLa and RPE1 cells (0.18 ± 0.21 °/ μm and 0.49 ± 0.21 °/μm, respectively), indicating that PRC1 contributes to the twist by constraining microtubule rotation, and augmin by nucleating antiparallel bridging microtubules. Discussion: The uncovered switch from left-handed to right- handed twist reveals the existence of competing mechanisms that promote twisting in opposite directions. As round spindles were more twisted than elongated ones, we infer that bending and twisting moments are generated by similar molecular mechanisms and propose a physiological role for spindle chirality in reducing the risk of spindle breakage under load. Methods: Live RPE1 and HeLa cells were imaged using Bruker Opterra Multipoint Scanning Confocal Microscope. To quantify spindle twist, we used 3 complementary approaches: visual assessment, optical flow, and bundle tracing. Acknowledgements: We thank Nenad Pavin, Maja Novak and all members of Tolić and Pavin groups for helpful discussions. This work was funded by the European Research Council (ERC Consolidator Grant, GA Number 647077), the Croatian Science Foundation (HRZZ project IP-2019-04-5967), the Science and Innovation Grant co-financed by the European Structural and Investment Funds (ESIF) within the Operational Programme Competitiveness and Cohesion (OPCC) 2014–2020 (Grant KK.01.1.1.04.0057), and the QuantiXLie Center of Excellence, a project co- financed by the Croatian Government and European Union through the European Regional Development Fund—the Competitiveness and Cohesion Operational Programme (Grant KK.01.1.1.01.0004). We also acknowledge new support by the ERC (Synergy Grant, GA Number 855158). The work of doctoral students M.T. and A.I. have been supported in part by the “Young researchers' career development project – training of doctoral students” of the Croatian Science Foundation. References: 1. Prosser, S.L., and Pelletier, L. (2017). Mitotic spindle assembly in animal cells: a fine balancing act. Nat Rev Mol Cell Biol 18, 187-201. 2. Novak, M., Polak, B., Simunic, J., Boban, Z., Kuzmic, B., Thomae, A.W., Tolic, I.M., and Pavin, N. (2018). The mitotic spindle is chiral due to torques within microtubule bundles. Nat Commun 9, 3571.

Izvorni jezik
Engleski

Znanstvena područja
Biologija, Interdisciplinarne prirodne znanosti

POVEZANOST RADA