bims-micesi Biomed News
on Mitotic cell signalling
Issue of 2024–03–17
eleven papers selected by
Valentina Piano, Uniklinik Köln
  1. The Arabidopsis BUB1/MAD3 family protein BMF3 requires BUB3.3 to recruit CDC20 to kinetochores in spindle assembly checkpoint signaling.
  2. The Role of Aurora B Kinase in Normal and Cancer Cells.
  3. The Golgi checkpoint: Golgi unlinking during G2 is necessary for spindle formation and cytokinesis.
  4. Survivin Mediates Mitotic Onset in HeLa Cells Through Activation of the Cdk1-Cdc25B Axis.
  5. The oncogene cyclin D1 promotes bipolar spindle integrity under compressive force.
  6. Molecular density-accelerated binding-site maturation underlies CENP-T-dependent kinetochore assembly.
  7. Reduction of cortical pulling at mitotic entry facilitates aster centration.
  8. Broken chromosomes heading into mitosis: More than one way to patch a flat tire.
  9. Architecture and flexibility of native kinetochores revealed by structural studies utilizing a thermophilic yeast.
  10. Phosphate-binding pocket on cyclin B governs CDK substrate phosphorylation and mitotic timing.
  11. Automatic detection of cell-cycle stages using recurrent neural networks.
  1. Proc Natl Acad Sci U S A. 2024 Mar 19. 121(12): e2322677121
    The Arabidopsis BUB1/MAD3 family protein BMF3 requires BUB3.3 to recruit CDC20 to kinetochores in spindle assembly checkpoint signaling.
    Xingguang Deng, Felicia Lei Peng, Xiaoya Tang, Yuh-Ru Julie Lee, Hong-Hui Lin, Bo Liu.
      The spindle assembly checkpoint (SAC) ensures faithful chromosome segregation during cell division by monitoring kinetochore-microtubule attachment. Plants produce both sequence-conserved and diverged SAC components, and it has been largely unknown how SAC activation leads to the assembly of these proteins at unattached kinetochores to prevent cells from entering anaphase. In Arabidopsis thaliana, the noncanonical BUB3.3 protein was detected at kinetochores throughout mitosis, unlike MAD1 and the plant-specific BUB1/MAD3 family protein BMF3 that associated with unattached chromosomes only. When BUB3.3 was lost by a genetic mutation, mitotic cells often entered anaphase with misaligned chromosomes and presented lagging chromosomes after they were challenged by low doses of the microtubule depolymerizing agent oryzalin, resulting in the formation of micronuclei. Surprisingly, BUB3.3 was not required for the kinetochore localization of other SAC proteins or vice versa. Instead, BUB3.3 specifically bound to BMF3 through two internal repeat motifs that were not required for BMF3 kinetochore localization. This interaction enabled BMF3 to recruit CDC20, a downstream SAC target, to unattached kinetochores. Taken together, our findings demonstrate that plant SAC utilizes unconventional protein interactions for arresting mitosis, with BUB3.3 directing BMF3's role in CDC20 recruitment, rather than the recruitment of BUB1/MAD3 proteins observed in fungi and animals. This distinct mechanism highlights how plants adapted divergent versions of conserved cell cycle machinery to achieve specialized SAC control.
    Keywords:  Arabidopsis; BUB3; CDC20; kinetochore; spindle assembly checkpoint
    DOI:  https://doi.org/10.1073/pnas.2322677121
  2. Biochemistry (Mosc). 2023 Dec;88(12): 2054-2062
    The Role of Aurora B Kinase in Normal and Cancer Cells.
    Ekaterina Titova, Galina Shagieva, Vera Dugina, Pavel Kopnin.
      Aurora kinases are essential players in mammalian cell division. These kinases are involved in the regulation of spindle dynamics, microtubule-kinetochore interactions, and chromosome condensation and orientation during mitosis. At least three members of the Aurora family - Aurora kinases A, B, and C - have been identified in mammals. Aurora B is essential for maintaining genomic stability and normal cell division. Mutations and dysregulation of this kinase are implicated in tumor initiation and progression. In this review, we discuss the functions of Aurora B, the relationship between increased Aurora B activity and carcinogenesis, and the prospects for the use of Aurora B kinase inhibitors in antitumor therapy.
    Keywords:  Aurora B; Aurora B inhibitors; cancer; cell cycle; mitosis
    DOI:  https://doi.org/10.1134/S0006297923120088
  3. Life Sci Alliance. 2024 May;pii: e202302469. [Epub ahead of print]7(5):
    The Golgi checkpoint: Golgi unlinking during G2 is necessary for spindle formation and cytokinesis.
    Fabiola Mascanzoni, Inmaculada Ayala, Roberta Iannitti, Alberto Luini, Antonino Colanzi.
      Entry into mitosis requires not only correct DNA replication but also extensive cell reorganization, including the separation of the Golgi ribbon into isolated stacks. To understand the significance of pre-mitotic Golgi reorganization, we devised a strategy to first block Golgi segregation, with the consequent G2-arrest, and then force entry into mitosis. We found that the cells forced to enter mitosis with an intact Golgi ribbon showed remarkable cell division defects, including spindle multipolarity and binucleation. The spindle defects were caused by reduced levels at the centrosome of the kinase Aurora-A, a pivotal spindle formation regulator controlled by Golgi segregation. Overexpression of Aurora-A rescued spindle formation, indicating a crucial role of the Golgi-dependent recruitment of Aurora-A at the centrosome. Thus, our results reveal that alterations of the pre-mitotic Golgi segregation in G2 have profound consequences on the fidelity of later mitotic processes and represent potential risk factors for cell transformation and cancer development.
    DOI:  https://doi.org/10.26508/lsa.202302469
  4. Res Sq. 2024 Feb 28. pii: rs.3.rs-3949429. [Epub ahead of print]
    Survivin Mediates Mitotic Onset in HeLa Cells Through Activation of the Cdk1-Cdc25B Axis.
    Pedro M Cánovas.
      The Survivin protein has roles in repairing incorrect microtubule-kinetochore attachments at prometaphase, and the faithful execution of cytokinesis, both as part of the chromosomal passenger complex (CPC) (1). In this context, errors frequently lead to aneuploidy, polyploidy and cancer (1). Adding to these well-known roles of this protein, this paper now shows for the first time that Survivin is required for cancer cells to enter mitosis, and that, in its absence, HeLa cells accumulate at early prophase, or prior to reported before (2, 3). This early prophase blockage is demonstrated by the presence of an intact nuclear lamina and low Cdk1 activity (4). Importantly, escaping the arrest induced by Survivin abrogation leads to multiple mitotic defects, or mitotic catastrophe, and eventually cell death. Mechanistically, Cdk1 does not localize at the centrosome in the absence of Survivin pointing at an impairment in signaling through the Cdc25B-Cdk1 axis. In agreement, even though Survivin directly interacts with Cdc25B, both in vitro and in vivo, in its absence, an inactive cytosolic Cdc25B-Cdk1-Cyclin B1 complex accumulates. This flaw in Cdc25B activation can however be reversed in Survivin-depleted HeLa cell extracts to which the recombinant Survivin protein is added back. Finally, a role for Survivin in the Cdc25B-mediated activation of Cdk1 is confirmed by overriding the early prophase blockage induced in cells lacking Survivin through the expression of a gain-of-function Cdc25B mutant.
    Keywords:  Cdc25B; Cdk1; G2/M-phase checkpoint; Survivin; cancer; kinase; mitosis; mitotic catastrophe; phosphatase
    DOI:  https://doi.org/10.21203/rs.3.rs-3949429/v1
  5. PLoS One. 2024 ;19(3): e0296779
    The oncogene cyclin D1 promotes bipolar spindle integrity under compressive force.
    Renaldo Sutanto, Lila Neahring, Andrea Serra Marques, Mauricio Jacobo Jacobo, Seda Kilinc, Andrei Goga, Sophie Dumont.
      The mitotic spindle is the bipolar, microtubule-based structure that segregates chromosomes at each cell division. Aberrant spindles are frequently observed in cancer cells, but how oncogenic transformation affects spindle mechanics and function, particularly in the mechanical context of solid tumors, remains poorly understood. Here, we constitutively overexpress the oncogene cyclin D1 in human MCF10A cells to probe its effects on spindle architecture and response to compressive force. We find that cyclin D1 overexpression increases the incidence of spindles with extra poles, centrioles, and chromosomes. However, it also protects spindle poles from fracturing under compressive force, a deleterious outcome linked to multipolar cell divisions. Our findings suggest that cyclin D1 overexpression may adapt cells to increased compressive stress, possibly contributing to its prevalence in cancers such as breast cancer by allowing continued proliferation in mechanically challenging environments.
    DOI:  https://doi.org/10.1371/journal.pone.0296779
  6. bioRxiv. 2024 Feb 26. pii: 2024.02.25.581584. [Epub ahead of print]
    Molecular density-accelerated binding-site maturation underlies CENP-T-dependent kinetochore assembly.
    Ekaterina V Tarasovetc, Gunter B Sissoko, Anna S Mukhina, Aleksandr Maiorov, Fazoil I Ataullakhanov, Iain M Cheeseman, Ekaterina L Grishchuk.
      Formation of macromolecular cellular structures relies on recruitment of multiple proteins, requiring the precisely controlled pairwise binding interactions. At human kinetochores, our recent work found that the high molecular density environment enables strong bonding between the Ndc80 complex and its two binding sites at the CENP-T receptor. However, the mechanistic basis for this unusual density-dependent facilitation remains unknown. Here, using quantitative single-molecule approaches, we reveal two distinct mechanisms that drive preferential recruitment of the Ndc80 complex to higher-order structures of CENP-T, as opposed to CENP-T monomers. First, the Ndc80 binding sites within the disordered tail of the CENP-T mature over time, leading to a stronger grip on the Spc24/25 heads of the Ndc80 complexes. Second, the maturation of Ndc80 binding sites is accelerated when CENP-T molecules are clustered in close proximity. The rates of the clustering-induced maturation are remarkably different for two binding sites within CENP-T, correlating with different interfaces formed by the corresponding CENP-T sequences as they wrap around the Spc24/25 heads. The differential clustering-dependent regulation of these sites is preserved in dividing human cells, suggesting a distinct regulatory entry point to control kinetochore-microtubule interactions. The tunable acceleration of slowly maturing binding sites by a high molecular-density environment may represent a fundamental physicochemical mechanism to assist the assembly of mitotic kinetochores and other macromolecular structures.
    DOI:  https://doi.org/10.1101/2024.02.25.581584
  7. J Cell Sci. 2024 Mar 12. pii: jcs.262037. [Epub ahead of print]
    Reduction of cortical pulling at mitotic entry facilitates aster centration.
    Anne Rosfelter, Ghislain de Labbey, Janet Chenevert, Rémi Dumollard, Sebastien Schaub, Zoltan Machaty, Lydia Besnardeau, Daniel Gonzalez Suarez, Céline Hebras, Hervé Turlier, David Burgess, Alex McDougall.
      Equal cell division relies upon astral microtubule-based centering mechanisms, yet how the interplay between mitotic entry, cortical force generation, and long astral microtubules leads to symmetric cell division is not resolved. We report that a cortically-located sperm aster displaying long astral microtubules that penetrate the whole zygote does not undergo centration until mitotic entry. At mitotic entry we find that microtubule-based cortical pulling is lost. Quantitative measurements of cortical pulling and cytoplasmic pulling together with physical simulations suggested a wavelike loss of cortical pulling at mitotic entry led to aster centration based on cytoplasmic pulling. Cortical actin is lost from the cortex at mitotic entry coincident with a fall in cortical tension from around 300pN/µm to 100pN/µm. Following the loss of cortical force generators at mitotic entry long microtubule-based cytoplasmic pulling is sufficient to displace the aster towards the cell center. These data reveal how mitotic aster centration is coordinated with mitotic entry in chordate zygotes.
    Keywords:  Cell cycle; Mitotic apparatus; Nuclear migration; Sperm aster
    DOI:  https://doi.org/10.1242/jcs.262037
  8. J Cell Biol. 2024 Apr 01. pii: e202401085. [Epub ahead of print]223(4):
    Broken chromosomes heading into mitosis: More than one way to patch a flat tire.
    C Luke Messer, Donald T Fox.
      A cell dealing with a broken chromosome in mitosis is like a driver dealing with a flat tire on the highway: damage repair must occur under non-ideal circumstances. Mitotic chromosome breaks encounter problems related to structures called micronuclei. These aberrant nuclei are linked to cell death, mutagenesis, and cancer. In the last few years, a flurry of studies illuminated two mechanisms that prevent mitotic problems related to micronuclei. One mechanism prevents micronuclei from forming during mitosis and involves DNA Polymerase Theta, a DNA repair regulator that patches up broken mitotic chromosomes. A second mechanism is activated after micronuclei form and then rupture, and involves CIP2A and TOPBP1 proteins, which patch micronuclear fragments to promote their subsequent mitotic segregation. Here, we review recent progress in this field of mitotic DNA damage and discuss why multiple mechanisms exist. Future studies in this exciting area will reveal new DNA break responses and inform therapeutic strategies.
    DOI:  https://doi.org/10.1083/jcb.202401085
  9. bioRxiv. 2024 Feb 29. pii: 2024.02.28.582571. [Epub ahead of print]
    Architecture and flexibility of native kinetochores revealed by structural studies utilizing a thermophilic yeast.
    Daniel J Barrero, Sithara S Wijeratne, Xiaowei Zhao, Grace F Cunningham, Rui Yan, Christian R Nelson, Yasuhiro Arimura, Hironori Funabiki, Charles L Asbury, Zhiheng Yu, Radhika Subramanian, Sue Biggins.
      Eukaryotic chromosome segregation requires kinetochores, multi-megadalton protein machines that assemble on the centromeres of chromosomes and mediate attachments to dynamic spindle microtubules. Kinetochores are built from numerous complexes, and understanding how they are arranged is key to understanding how kinetochores perform their multiple functions. However, an integrated understanding of kinetochore architecture has not yet been established. To address this, we purified functional, native kinetochores from Kluyveromyces marxianus and examined them by electron microscopy, cryo-electron tomography and atomic force microscopy. The kinetochores are extremely large, flexible assemblies that exhibit features consistent with prior models. We assigned kinetochore polarity by visualizing their interactions with microtubules and locating the microtubule binder Ndc80c. This work shows that isolated kinetochores are more dynamic and complex than what might be anticipated based on the known structures of recombinant subassemblies, and provides the foundation to study the global architecture and functions of kinetochores at a structural level.
    DOI:  https://doi.org/10.1101/2024.02.28.582571
  10. bioRxiv. 2024 Feb 29. pii: 2024.02.28.582599. [Epub ahead of print]
    Phosphate-binding pocket on cyclin B governs CDK substrate phosphorylation and mitotic timing.
    Henry Y Ng, Armin N Adly, Devon H Whelpley, Raymond T Suhandynata, Huilin Zhou, David O Morgan.
      Cell cycle progression is governed by complexes of the cyclin-dependent kinases (CDKs) and their regulatory subunits cyclin and Cks1. CDKs phosphorylate hundreds of substrates, often at multiple sites. Multisite phosphorylation depends on Cks1, which binds initial priming phosphorylation sites to promote secondary phosphorylation at other sites. Here, we describe a similar role for a recently discovered phosphate-binding pocket (PP) on B-type cyclins. Mutation of the PP in Clb2, the major mitotic cyclin of budding yeast, alters bud morphology and delays the onset of anaphase. Using phosphoproteomics in vivo and kinase reactions in vitro , we find that mutation of the PP reduces phosphorylation of several CDK substrates, including the Bud6 subunit of the polarisome and the Cdc16 and Cdc27 subunits of the anaphase-promoting complex/cyclosome. We conclude that the cyclin PP, like Cks1, controls the timing of multisite phosphorylation on CDK substrates, thereby helping to establish the robust timing of cell-cycle events.
    DOI:  https://doi.org/10.1101/2024.02.28.582599
  11. PLoS One. 2024 ;19(3): e0297356
    Automatic detection of cell-cycle stages using recurrent neural networks.
    Abin Jose, Rijo Roy, Daniel Moreno-Andrés, Johannes Stegmaier.
      Mitosis is the process by which eukaryotic cells divide to produce two similar daughter cells with identical genetic material. Research into the process of mitosis is therefore of critical importance both for the basic understanding of cell biology and for the clinical approach to manifold pathologies resulting from its malfunctioning, including cancer. In this paper, we propose an approach to study mitotic progression automatically using deep learning. We used neural networks to predict different mitosis stages. We extracted video sequences of cells undergoing division and trained a Recurrent Neural Network (RNN) to extract image features. The use of RNN enabled better extraction of features. The RNN-based approach gave better performance compared to classifier based feature extraction methods which do not use time information. Evaluation of precision, recall, and F-score indicates the superiority of the proposed model compared to the baseline. To study the loss in performance due to confusion between adjacent classes, we plotted the confusion matrix as well. In addition, we visualized the feature space to understand why RNNs are better at classifying the mitosis stages than other classifier models, which indicated the formation of strong clusters for the different classes, clearly confirming the advantage of the proposed RNN-based approach.
    DOI:  https://doi.org/10.1371/journal.pone.0297356
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