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

Trupinić, Monika; Kokanović, Barbara; Ponjavić, Ivana; Barišić, Ivan; Šegvić, Siniša; Ivec, Arian; Tolić, Iva M.

doi:10.1016/j.cub.2022.04.035

Cited by 14 publications

(24 citation statements)

References 76 publications

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“…Opposing the effects of KIF4A and MKLP1, dynein and its targeting factors NuMA and LGN are each required to restrain the anaphase spindle's left-handed twist. Although another study found that dynein inhibition did not increase twist in metaphase HeLa or RPE1 spindles (Trupinic et al, 2022), we observe strong phenotypes after dynein depletion in MCF10A cells or NuMA knockout in RPE1 cells (Fig. 4).…”

Section: Dynein Counteracts Left-handed Twist In the Anaphase Spindlecontrasting

confidence: 63%

“…MCF10A spindles exhibit high baseline twist that peaks in late metaphase and anaphase To study torque regulation in the spindle, we sought to identify a cell line in which spindles exhibited higher baseline twist than that observed in previously characterized cell lines. We reasoned that because twist differs between the human cell lines RPE1, HeLa, and U2OS (Neahring et al, 2021;Novak et al, 2018;Trupinic et al, 2022), other human cell lines may exhibit stronger twist, and that stronger twist would allow us greater dynamic range to study factors that both increase and decrease twist. We quantified twist using the optical flow method (Trupinic et al, 2022) in which we liveimaged full spindle volumes, computationally rotated the images to view the spindle along the pole-to-pole axis, and calculated the displacement fields of pixel intensities between successive frames from 30% and 70% of the pole-to-pole axis (Fig.…”

Section: Resultsmentioning

confidence: 99%

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Torques within and outside the human spindle balance twist at anaphase

Neahring,

He,

Cho

et al. 2023

Preprint

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At each cell division, nanometer-scale motors and microtubules give rise to the micron-scale spindle. Many mitotic motors step helically around microtubules in vitro, and most are predicted to twist the spindle in a left-handed direction. However, the human spindle exhibits only slight global twist, raising the question of how these molecular torques are balanced. Here, using lattice light sheet microscopy, we find that anaphase spindles in the epithelial cell line MCF10A have a high baseline twist, and we identify factors that both increase and decrease this twist. The midzone motors KIF4A and MKLP1 are redundantly required for left-handed twist at anaphase, and we show that KIF4A generates left-handed torque in vitro. The actin cytoskeleton also contributes to left-handed twist, but dynein and its cortical recruitment factor LGN counteract it. Together, our work demonstrates that force generators regulate twist in opposite directions from both within and outside the spindle, preventing strong spindle twist during chromosome segregation.

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Section: Dynein Counteracts Left-handed Twist In the Anaphase Spindlecontrasting

confidence: 63%

Section: Resultsmentioning

confidence: 99%

Torques within and outside the human spindle balance twist at anaphase

Neahring,

He,

Cho

et al. 2023

Preprint

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“…The observation that the more stable set of midzone microtubules became more stable is consistent with time dependent modulation of microtubule dynamics as cells exit mitosis (Mollinari et al, 2002; Neef et al, 2007). Previous work showed that midzone microtubules are generated by augmin-mediated nucleation in the half-spindle and midzone regions (Goshima et al, 2008; Štimac et al, 2022; Trupinić et al, 2022; Uehara et al, 2016).…”

Section: Discussionmentioning

confidence: 99%

Lateral and longitudinal compaction of PRC1 overlap zones drive stabilization of interzonal microtubules

Rosário

Zhang

Stadnicki

et al. 2023

Preprint

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During anaphase, antiparallel overlapping midzone microtubules elongate and form bundles, contributing to chromosome segregation and the location of contractile ring formation. Midzone microtubules are dynamic in early but not late anaphase; however, the kinetics and mechanisms of stabilization are incompletely understood. Using photoactivation of cells expressing PA-EGF-α-tubulin we find that immediately after anaphase onset, a single highly dynamic population of midzone microtubules is present; as anaphase progresses, both dynamic and stable populations of midzone microtubules coexist. By midcytokinesis, only static, non-dynamic microtubules are detected. The velocity of microtubule sliding also decreases as anaphase progresses, becoming undetectable by late anaphase. Following depletion of PRC1, midzone microtubules remain highly dynamic in anaphase and fail to form static arrays in telophase despite furrowing. Cells depleted of Kif4a contain elongated zones of PRC1 and fail to form static arrays in telophase. Cells blocked in cytokinesis form short PRC1 overlap zones that do not coalesce laterally; these cells also fail to form static arrays in telophase. Together, our results demonstrate that dynamic turnover and sliding of midzone microtubules is gradually reduced during anaphase and that the final transition to a static array in telophase requires both lateral and longitudinal compaction of PRC1 containing overlap zones.

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“…The k-fiber’s connection (anchorage) to the spindle is mediated by a dense mesh-like network of non-kinetochore microtubules (non-kMTs) which connect to k-fibers along their length ( Mastronarde et al, 1993 ; O’Toole et al, 2020 ) via both motor and non-motor proteins. Although this network cannot be easily visualized with light microscopy, physical perturbations such as laser ablation ( Kajtez et al, 2016 ; Milas and Tolić, 2016 ; Elting et al, 2017 ) and cell compression ( Trupinić et al, 2022 ; Neahring et al, 2021 ) have been instrumental in uncovering how this network anchors k-fibers. The non-kMT network bears mechanical load locally ( Milas and Tolić, 2016 ; Elting et al, 2017 ), links sister k-fibers together ( Kajtez et al, 2016 ), and contributes to k-fiber and spindle chirality ( Trupinić et al, 2022 ; Neahring et al, 2021 ).…”

Section: Introductionmentioning

confidence: 99%

“…Although this network cannot be easily visualized with light microscopy, physical perturbations such as laser ablation ( Kajtez et al, 2016 ; Milas and Tolić, 2016 ; Elting et al, 2017 ) and cell compression ( Trupinić et al, 2022 ; Neahring et al, 2021 ) have been instrumental in uncovering how this network anchors k-fibers. The non-kMT network bears mechanical load locally ( Milas and Tolić, 2016 ; Elting et al, 2017 ), links sister k-fibers together ( Kajtez et al, 2016 ), and contributes to k-fiber and spindle chirality ( Trupinić et al, 2022 ; Neahring et al, 2021 ). Recent advances in microneedle manipulation enabled us to mechanically challenge k-fiber anchorage with unprecedented spatiotemporal control ( Long et al, 2020 ; Suresh et al, 2020 ).…”

Section: Introductionmentioning

confidence: 99%

Modeling and mechanical perturbations reveal how spatially regulated anchorage gives rise to spatially distinct mechanics across the mammalian spindle

Suresh

Galstyan

Phillips

et al. 2022

eLife

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During cell division, the spindle generates force to move chromosomes. In mammals, microtubule bundles called kinetochore-fibers (k-fibers) attach to and segregate chromosomes. To do so, k-fibers must be robustly anchored to the dynamic spindle. We previously developed microneedle manipulation to mechanically challenge k-fiber anchorage, and observed spatially distinct response features revealing the presence of heterogeneous anchorage (Suresh et al., 2020). How anchorage is precisely spatially regulated, and what forces are necessary and sufficient to recapitulate the k-fiber’s response to force remain unclear. Here, we develop a coarse-grained k-fiber model and combine with manipulation experiments to infer underlying anchorage using shape analysis. By systematically testing different anchorage schemes, we find that forces solely at k-fiber ends are sufficient to recapitulate unmanipulated k-fiber shapes, but not manipulated ones for which lateral anchorage over a 3 μm length scale near chromosomes is also essential. Such anchorage robustly preserves k-fiber orientation near chromosomes while allowing pivoting around poles. Anchorage over a shorter length scale cannot robustly restrict pivoting near chromosomes, while anchorage throughout the spindle obstructs pivoting at poles. Together, this work reveals how spatially regulated anchorage gives rise to spatially distinct mechanics in the mammalian spindle, which we propose are key for function.

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

Cited by 14 publications

References 76 publications

Torques within and outside the human spindle balance twist at anaphase

Torques within and outside the human spindle balance twist at anaphase

Lateral and longitudinal compaction of PRC1 overlap zones drive stabilization of interzonal microtubules

Modeling and mechanical perturbations reveal how spatially regulated anchorage gives rise to spatially distinct mechanics across the mammalian spindle

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