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      Contractile acto-myosin network on nuclear envelope remnants positions human chromosomes for mitosis

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          Abstract

          To ensure proper segregation during mitosis, chromosomes must be efficiently captured by spindle microtubules and subsequently aligned on the mitotic spindle. The efficacy of chromosome interaction with the spindle can be influenced by how widely chromosomes are scattered in space. Here, we quantify chromosome-scattering volume (CSV) and find that it is reduced soon after nuclear envelope breakdown (NEBD) in human cells. The CSV reduction occurs primarily independently of microtubules and is therefore not an outcome of interactions between chromosomes and the spindle. We find that, prior to NEBD, an acto-myosin network is assembled in a LINC complex-dependent manner on the cytoplasmic surface of the nuclear envelope. This acto-myosin network remains on nuclear envelope remnants soon after NEBD, and its myosin-II-mediated contraction reduces CSV and facilitates timely chromosome congression and correct segregation. Thus, we find a novel mechanism that positions chromosomes in early mitosis to ensure efficient and correct chromosome–spindle interactions.

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          The Actin Cytoskeleton and Actin-Based Motility

          The actin cytoskeleton—a collection of actin filaments with their accessory and regulatory proteins—is the primary force-generating machinery in the cell. It can produce pushing (protrusive) forces through coordinated polymerization of multiple actin filaments or pulling (contractile) forces through sliding actin filaments along bipolar filaments of myosin II. Both force types are particularly important for whole-cell migration, but they also define and change the cell shape and mechanical properties of the cell surface, drive the intracellular motility and morphogenesis of membrane organelles, and allow cells to form adhesions with each other and with the extracellular matrix.
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            Small-molecule inhibition of Wee1 kinase by MK-1775 selectively sensitizes p53-deficient tumor cells to DNA-damaging agents.

            Wee1 is a tyrosine kinase that phosphorylates and inactivates CDC2 and is involved in G(2) checkpoint signaling. Because p53 is a key regulator in the G(1) checkpoint, p53-deficient tumors rely only on the G(2) checkpoint after DNA damage. Hence, such tumors are selectively sensitized to DNA-damaging agents by Wee1 inhibition. Here, we report the discovery of a potent and selective small-molecule inhibitor of Wee1 kinase, MK-1775. This compound inhibits phosphorylation of CDC2 at Tyr15 (CDC2Y15), a direct substrate of Wee1 kinase in cells. MK-1775 abrogates G(2) DNA damage checkpoint, leading to apoptosis in combination with DNA-damaging chemotherapeutic agents such as gemcitabine, carboplatin, and cisplatin selectively in p53-deficient cells. In vivo, MK-1775 potentiates tumor growth inhibition by these agents, and cotreatment does not significantly increase toxicity. The enhancement of antitumor effect by MK-1775 was well correlated with inhibition of CDC2Y15 phosphorylation in tumor tissue and skin hair follicles. Our data indicate that Wee1 inhibition provides a new approach for treatment of multiple human malignancies.
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              Chromosomes can congress to the metaphase plate before biorientation.

              The stable propagation of genetic material during cell division depends on the congression of chromosomes to the spindle equator before the cell initiates anaphase. It is generally assumed that congression requires that chromosomes are connected to the opposite poles of the bipolar spindle ("bioriented"). In mammalian cells, we found that chromosomes can congress before becoming bioriented. By combining the use of reversible chemical inhibitors, live-cell light microscopy, and correlative electron microscopy, we found that monooriented chromosomes could glide toward the spindle equator alongside kinetochore fibers attached to other already bioriented chromosomes. This congression mechanism depended on the kinetochore-associated, plus end-directed microtubule motor CENP-E (kinesin-7).
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                Author and article information

                Contributors
                Role: Reviewing Editor
                Role: Senior Editor
                Journal
                eLife
                Elife
                eLife
                eLife
                eLife Sciences Publications, Ltd
                2050-084X
                03 July 2019
                2019
                : 8
                : e46902
                Affiliations
                [1 ]deptCentre for Gene Regulation and Expression, School of Life Sciences University of Dundee DundeeUnited Kingdom
                [2 ]deptGenome Damage and Stability Centre University of Sussex BrightonUnited Kingdom
                [3 ]deptCollege of Biological Sciences University of Minnesota MinneapolisUnited States
                Virginia Tech United States
                Utrecht University Netherlands
                Virginia Tech United States
                i3S- Instituto de Investigação e Inovação em Saúde, Universidade do Porto Portugal
                Author information
                http://orcid.org/0000-0002-3320-7919
                https://orcid.org/0000-0003-4115-9686
                http://orcid.org/0000-0002-6180-8906
                https://orcid.org/0000-0001-8366-6198
                https://orcid.org/0000-0002-9886-5947
                Article
                46902
                10.7554/eLife.46902
                6634967
                31264963
                1e4245ef-6ecb-4262-9aeb-a058720382bf
                © 2019, Booth et al

                This article is distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use and redistribution provided that the original author and source are credited.

                History
                : 15 March 2019
                : 01 July 2019
                Funding
                Funded by: FundRef http://dx.doi.org/10.13039/100004440, Wellcome;
                Award ID: 096535/Z/11/Z
                Award Recipient :
                Funded by: FundRef http://dx.doi.org/10.13039/100004440, Wellcome;
                Award ID: 097945/Z/11/Z
                Award Recipient :
                Funded by: FundRef http://dx.doi.org/10.13039/100004440, Wellcome;
                Award ID: 208401/Z/17/Z
                Award Recipient :
                Funded by: FundRef http://dx.doi.org/10.13039/501100000289, Cancer Research UK;
                Award ID: C28206/A114499
                Award Recipient :
                Funded by: FundRef http://dx.doi.org/10.13039/501100000265, Medical Research Council;
                Award ID: MR/K015869/1
                Award Recipient :
                The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
                Categories
                Research Article
                Cell Biology
                Chromosomes and Gene Expression
                Custom metadata
                Live-cell imaging shows that the contractile acto-myosin network on the nuclear envelope remnant positions chromosomes in early mitosis to ensure efficient and correct interactions between chromosomes and the mitotic spindle.

                Life sciences
                chromosomes,mitotic spindle,acto-myosin,nuclear envelope,linc complex,human cells,human
                Life sciences
                chromosomes, mitotic spindle, acto-myosin, nuclear envelope, linc complex, human cells, human

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