bims-micesi Biomed News
on Mitotic cell signalling
Issue of 2024–03–31
thirteen papers selected by
Valentina Piano, Uniklinik Köln
  1. MAPK-dependent control of mitotic progression in S. pombe.
  2. Early onset of APC/C activity renders SAC inefficient in mouse embryos.
  3. Mitotic chromosomes are self-entangled and disentangle through a topoisomerase-II-dependent two-stage exit from mitosis.
  4. Mechanobiology of the nucleus during the G2-M transition.
  5. A mitotic stopwatch determines cell fate.
  6. Analysis of non-degradable cyclins reveals distinct roles of the mitotic cyclins in Drosophila meiosis.
  7. A dynamic role for transcription factors in restoring transcription through mitosis.
  8. Control of cell proliferation by memories of mitosis.
  9. Recombinant cyclin B-Cdk1-Suc1 capable of multi-site mitotic phosphorylation in vitro.
  10. Cell cycle perturbation uncouples mitotic progression and invasive behavior in a post-mitotic cell.
  11. Deficiency of mastl, a mitotic regulator, results in cell detachment from developing tissues of zebrafish embryos.
  12. A stable microtubule bundle formed through an orchestrated multistep process controls quiescence exit.
  13. Spindle checkpoint activation by fungal orthologs of the S. cerevisiae Mps1 kinase.
  1. BMC Biol. 2024 Mar 25. 22(1): 71
    MAPK-dependent control of mitotic progression in S. pombe.
    Ana Belén Iglesias-Romero, Terersa Soto, Ignacio Flor-Parra, Silvia Salas-Pino, Gabriel Ruiz-Romero, Kathleen L Gould, José Cansado, Rafael R Daga.
       BACKGROUND: Mitogen-activated protein kinases (MAPKs) preserve cell homeostasis by transducing physicochemical fluctuations of the environment into multiple adaptive responses. These responses involve transcriptional rewiring and the regulation of cell cycle transitions, among others. However, how stress conditions impinge mitotic progression is largely unknown. The mitotic checkpoint is a surveillance mechanism that inhibits mitotic exit in situations of defective chromosome capture, thus preventing the generation of aneuploidies. In this study, we investigate the role of MAPK Pmk1 in the regulation of mitotic exit upon stress.
    RESULTS: We show that Schizosaccharomyces pombe cells lacking Pmk1, the MAP kinase effector of the cell integrity pathway (CIP), are hypersensitive to microtubule damage and defective in maintaining a metaphase arrest. Epistasis analysis suggests that Pmk1 is involved in maintaining spindle assembly checkpoint (SAC) signaling, and its deletion is additive to the lack of core SAC components such as Mad2 and Mad3. Strikingly, pmk1Δ cells show up to twofold increased levels of the anaphase-promoting complex (APC/C) activator Cdc20Slp1 during unperturbed growth. We demonstrate that Pmk1 physically interacts with Cdc20Slp1 N-terminus through a canonical MAPK docking site. Most important, the Cdc20Slp1 pool is rapidly degraded in stressed cells undergoing mitosis through a mechanism that requires MAPK activity, Mad3, and the proteasome, thus resulting in a delayed mitotic exit.
    CONCLUSIONS: Our data reveal a novel function of MAPK in preventing mitotic exit and activation of cytokinesis in response to stress. The regulation of Cdc20Slp1 turnover by MAPK Pmk1 provides a key mechanism by which the timing of mitotic exit can be adjusted relative to environmental conditions.
    Keywords:  APC/C; CDC20/Slp1; CIP; Cytokinesis; MAPK; MCC; Mitosis; Osmotic stress; Pmk1; SAPK; Spindle assembly checkpoint; Sty1
    DOI:  https://doi.org/10.1186/s12915-024-01865-6
  2. Front Cell Dev Biol. 2024 ;12 1355979
    Early onset of APC/C activity renders SAC inefficient in mouse embryos.
    Adela Horakova, Marketa Konecna, Lenka Radonova, Martin Anger.
      Control mechanisms of spindle assembly and chromosome segregation are vital for preventing aneuploidy during cell division. The mammalian germ cells and embryos are prone to chromosome segregation errors, and the resulting aneuploidy is a major cause of termination of development or severe developmental disorders. Here we focused on early mouse embryos, and using combination of methods involving microinjection, immunodetection and confocal live cell imaging, we concentrated on the Spindle Assembly Checkpoint (SAC) and Anaphase Promoting Complex/Cyclosome (APC/C). These are two important mechanisms cooperating during mitosis to ensure accurate chromosome segregation, and assessed their activity during the first two mitoses after fertilization. Our results showed, that in zygotes and 2-cell embryos, the SAC core protein Mad1 shows very low levels on kinetochores in comparison to oocytes and its interaction with chromosomes is restricted to a short time interval after nuclear membrane disassembly (NEBD). Exposure of 2-cell embryos to low levels of spindle poison does not prevent anaphase, despite the spindle damage induced by the drug. Lastly, the APC/C is activated coincidentally with NEBD before the spindle assembly completion. This early onset of APC/C activity, together with precocious relocalization of Mad1 from chromosomes, prevents proper surveillance of spindle assembly by SAC. The results contribute to the understanding of the origin of aneuploidy in early embryos.
    Keywords:  Mad1; anaphase; anaphase-promoting complex; chromosome segregation; embryo; spindle; spindle assembly checkpoint
    DOI:  https://doi.org/10.3389/fcell.2024.1355979
  3. Mol Cell. 2024 Mar 13. pii: S1097-2765(24)00144-8. [Epub ahead of print]
    Mitotic chromosomes are self-entangled and disentangle through a topoisomerase-II-dependent two-stage exit from mitosis.
    Erica M Hildebrand, Kirill Polovnikov, Bastiaan Dekker, Yu Liu, Denis L Lafontaine, A Nicole Fox, Ying Li, Sergey V Venev, Leonid A Mirny, Job Dekker.
      The topological state of chromosomes determines their mechanical properties, dynamics, and function. Recent work indicated that interphase chromosomes are largely free of entanglements. Here, we use Hi-C, polymer simulations, and multi-contact 3C and find that, by contrast, mitotic chromosomes are self-entangled. We explore how a mitotic self-entangled state is converted into an unentangled interphase state during mitotic exit. Most mitotic entanglements are removed during anaphase/telophase, with remaining ones removed during early G1, in a topoisomerase-II-dependent process. Polymer models suggest a two-stage disentanglement pathway: first, decondensation of mitotic chromosomes with remaining condensin loops produces entropic forces that bias topoisomerase II activity toward decatenation. At the second stage, the loops are released, and the formation of new entanglements is prevented by lower topoisomerase II activity, allowing the establishment of unentangled and territorial G1 chromosomes. When mitotic entanglements are not removed in experiments and models, a normal interphase state cannot be acquired.
    Keywords:  G1; chromosome entanglement; chromosome folding; cohesin; fractal globule; genome architecture; genome topology; mitosis; self-catenation; topoisomerase II
    DOI:  https://doi.org/10.1016/j.molcel.2024.02.025
  4. Nucleus. 2024 Dec;15(1): 2330947
    Mechanobiology of the nucleus during the G2-M transition.
    Joana T Lima, Jorge G Ferreira.
      Cellular behavior is continuously influenced by mechanical forces. These forces span the cytoskeleton and reach the nucleus, where they trigger mechanotransduction pathways that regulate downstream biochemical events. Therefore, the nucleus has emerged as a regulator of cellular response to mechanical stimuli. Cell cycle progression is regulated by cyclin-CDK complexes. Recent studies demonstrated these biochemical pathways are influenced by mechanical signals, highlighting the interdependence of cellular mechanics and cell cycle regulation. In particular, the transition from G2 to mitosis (G2-M) shows significant changes in nuclear structure and organization, ranging from nuclear pore complex (NPC) and nuclear lamina disassembly to chromosome condensation. The remodeling of these mechanically active nuclear components indicates that mitotic entry is particularly sensitive to forces. Here, we address how mechanical forces crosstalk with the nucleus to determine the timing and efficiency of the G2-M transition. Finally, we discuss how the deregulation of nuclear mechanics has consequences for mitosis.
    Keywords:  Chromosome; G2-M transition; LINC complex; mechanotransduction; nuclear envelope; nuclear pore complex; nucleus
    DOI:  https://doi.org/10.1080/19491034.2024.2330947
  5. Science. 2024 Mar 29. 383(6690): 1414-1415
    A mitotic stopwatch determines cell fate.
    Agustina P Bertolin, Vanesa Gottifredi.
      Surveillance of mitotic timing prevents amplification of damaged cells.
    DOI:  https://doi.org/10.1126/science.ado5703
  6. G3 (Bethesda). 2024 Mar 29. pii: jkae066. [Epub ahead of print]
    Analysis of non-degradable cyclins reveals distinct roles of the mitotic cyclins in Drosophila meiosis.
    Mohammed Bourouh, Rajdeep Dhaliwal, Rajni Rai, Hafsah Qureshi, Andrew Swan.
      Meiosis is a complex variant of the mitotic cell cycle, and as such relies on many of the same proteins involved in mitotis, but utilizes these in novel ways. As in mitosis, Cdk1 and its cyclin partners, Cyclin A, B and B3 are required at multiple steps in meiosis. Here we study the effect of stabilized forms of the three mitotic cyclins to study the consequences of failure to degrade the cyclins in meiosis. We find that stabilized Cyclin B3 promotes ectopic microtubule polymerization throughout the egg, dependent on APC/C activity and apparently due to the consequent destruction of Cyclin A and Cyclin B. We present data that suggests CycB, and possibly CycA, can also promote APC/C activity at specific stages of meiosis. We also present evidence that in meiosis APC/CCort and APC/CFzy are able to target Cyclin B via a novel degron. Overall, our findings highlight the distinct functions of the three mitotic Cdk-cyclin complexes in meiosis.
    Keywords:   Drosophila ; APC/C; cyclin; meiosis
    DOI:  https://doi.org/10.1093/g3journal/jkae066
  7. Biochem Soc Trans. 2024 Mar 25. pii: BST20231022. [Epub ahead of print]
    A dynamic role for transcription factors in restoring transcription through mitosis.
    Marek A Budzyński, Alexander K L Wong, Armin Faghihi, Sheila S Teves.
      Mitosis involves intricate steps, such as DNA condensation, nuclear membrane disassembly, and phosphorylation cascades that temporarily halt gene transcription. Despite this disruption, daughter cells remarkably retain the parent cell's gene expression pattern, allowing for efficient transcriptional memory after division. Early studies in mammalian cells suggested that transcription factors (TFs) mark genes for swift reactivation, a phenomenon termed 'mitotic bookmarking', but conflicting data emerged regarding TF presence on mitotic chromosomes. Recent advancements in live-cell imaging and fixation-free genomics challenge the conventional belief in universal formaldehyde fixation, revealing dynamic TF interactions during mitosis. Here, we review recent studies that provide examples of at least four modes of TF-DNA interaction during mitosis and the molecular mechanisms that govern these interactions. Additionally, we explore the impact of these interactions on transcription initiation post-mitosis. Taken together, these recent studies call for a paradigm shift toward a dynamic model of TF behavior during mitosis, underscoring the need for incorporating dynamics in mechanistic models for re-establishing transcription post-mitosis.
    Keywords:  binding dynamics; mitotic bookmarking; transcription factors; transcriptional memory
    DOI:  https://doi.org/10.1042/BST20231022
  8. Science. 2024 Mar 29. 383(6690): 1441-1448
    Control of cell proliferation by memories of mitosis.
    Franz Meitinger, Hazrat Belal, Robert L Davis, Mallory B Martinez, Andrew K Shiau, Karen Oegema, Arshad Desai.
      Mitotic duration is tightly constrained, and extended mitosis is characteristic of problematic cells prone to chromosome missegregation and genomic instability. We show here that mitotic extension leads to the formation of p53-binding protein 1 (53BP1)-ubiquitin-specific protease 28 (USP28)-p53 protein complexes that are transmitted to, and stably retained by, daughter cells. Complexes assembled through a Polo-like kinase 1-dependent mechanism during extended mitosis and elicited a p53 response in G1 that prevented the proliferation of the progeny of cells that experienced an approximately threefold extended mitosis or successive less extended mitoses. The ability to monitor mitotic extension was lost in p53-mutant cancers and some p53-wild-type (p53-WT) cancers, consistent with classification of TP53BP1 and USP28 as tumor suppressors. Cancers retaining the ability to monitor mitotic extension exhibited sensitivity to antimitotic agents.
    DOI:  https://doi.org/10.1126/science.add9528
  9. PLoS One. 2024 ;19(3): e0299003
    Recombinant cyclin B-Cdk1-Suc1 capable of multi-site mitotic phosphorylation in vitro.
    Keishi Shintomi, Yuki Masahara-Negishi, Masami Shima, Shoji Tane, Tatsuya Hirano.
      Cyclin-dependent kinase 1 (Cdk1) complexed with cyclin B phosphorylates multiple sites on hundreds of proteins during mitosis. However, it is not fully understood how multi-site mitotic phosphorylation by cyclin B-Cdk1 controls the structures and functions of individual substrates. Here we develop an easy-to-use protocol to express recombinant vertebrate cyclin B and Cdk1 in insect cells from a single baculovirus vector and to purify their complexes with excellent homogeneity. A series of in-vitro assays demonstrate that the recombinant cyclin B-Cdk1 can efficiently and specifically phosphorylate the SP and TP motifs in substrates. The addition of Suc1 (a Cks1 homolog in fission yeast) accelerates multi-site phosphorylation of an artificial substrate containing TP motifs. Importantly, we show that mitosis-specific multi-subunit and multi-site phosphorylation of the condensin I complex can be recapitulated in vitro using recombinant cyclin B-Cdk1-Suc1. The materials and protocols described here will pave the way for dissecting the biochemical basis of critical mitotic processes that accompany Cdk1-mediated large-scale phosphorylation.
    DOI:  https://doi.org/10.1371/journal.pone.0299003
  10. Differentiation. 2024 Mar 11. pii: S0301-4681(24)00021-5. [Epub ahead of print]137 100765
    Cell cycle perturbation uncouples mitotic progression and invasive behavior in a post-mitotic cell.
    Michael A Q Martinez, Chris Z Zhao, Frances E Q Moore, Callista Yee, Wan Zhang, Kang Shen, Benjamin L Martin, David Q Matus.
      The acquisition of the post-mitotic state is crucial for the execution of many terminally differentiated cell behaviors during organismal development. However, the mechanisms that maintain the post-mitotic state in this context remain poorly understood. To gain insight into these mechanisms, we used the genetically and visually accessible model of C. elegans anchor cell (AC) invasion into the vulval epithelium. The AC is a terminally differentiated uterine cell that normally exits the cell cycle and enters a post-mitotic state before initiating contact between the uterus and vulva through a cell invasion event. Here, we set out to identify the set of negative cell cycle regulators that maintain the AC in this post-mitotic, invasive state. Our findings revealed a critical role for CKI-1 (p21CIP1/p27KIP1) in redundantly maintaining the post-mitotic state of the AC, as loss of CKI-1 in combination with other negative cell cycle regulators-including CKI-2 (p21CIP1/p27KIP1), LIN-35 (pRb/p107/p130), FZR-1 (Cdh1/Hct1), and LIN-23 (β-TrCP)-resulted in proliferating ACs. Remarkably, time-lapse imaging revealed that these ACs retain their ability to invade. Upon examination of a node in the gene regulatory network controlling AC invasion, we determined that proliferating, invasive ACs do so by maintaining aspects of pro-invasive gene expression. We therefore report that the requirement for a post-mitotic state for invasive cell behavior can be bypassed following direct cell cycle perturbation.
    Keywords:  C. elegans; CKI-1; Dichotomy; Invasion; Proliferation
    DOI:  https://doi.org/10.1016/j.diff.2024.100765
  11. Front Cell Dev Biol. 2024 ;12 1375655
    Deficiency of mastl, a mitotic regulator, results in cell detachment from developing tissues of zebrafish embryos.
    Hideko Utsumi, Taijiro Yabe, Sumito Koshida, Akira Yamashita, Shinji Takada.
      To form tissues with unique functions and structures, it is important that the cells that comprise them maintain physical contact. On the other hand, with each mitosis, drastic changes in cell shapes, cell adhesion, and cytoskeletal architecture may cause such contacts to be temporarily weakened, risking improper development and maintenance of tissues. Despite such risks, tissues form properly during normal development. However, it is not well understood whether mitotic abnormalities affect tissue formation. Here, analysis of zebrafish embryos with aberrant mitosis shows that proper progression of mitosis is important to maintain cell contact in developing tissues. By screening mutants with abnormal trunk and tail development, we obtained a mutant with perturbed expression of some tissue-specific genes in embryonic caudal regions. The responsible gene is mastl/gwl, which is involved in progression of mitosis. Analysis focusing on the chordo-neural hinge (CNH), the primordium of axial tissues, shows that cell detachment from the CNH is increased in mastl mutant embryos. Time-lapse imaging reveals that this cell detachment occurs during mitosis. These results suggest that cells are unable to maintain contact due to abnormalities in progression of mitosis in mastl mutants.
    Keywords:  MASTL; axis elongation; cell detachment; chordo-neural hinge; mitosis; zebrafish
    DOI:  https://doi.org/10.3389/fcell.2024.1375655
  12. Elife. 2024 Mar 25. pii: RP89958. [Epub ahead of print]12
    A stable microtubule bundle formed through an orchestrated multistep process controls quiescence exit.
    Damien Laporte, Aurelie Massoni-Laporte, Charles Lefranc, Jim Dompierre, David Mauboules, Emmanuel T Nsamba, Anne Royou, Lihi Gal, Maya Schuldiner, Mohan L Gupta, Isabelle Sagot.
      Cells fine-tune microtubule assembly in both space and time to give rise to distinct edifices with specific cellular functions. In proliferating cells, microtubules are highly dynamics, and proliferation cessation often leads to their stabilization. One of the most stable microtubule structures identified to date is the nuclear bundle assembled in quiescent yeast. In this article, we characterize the original multistep process driving the assembly of this structure. This Aurora B-dependent mechanism follows a precise temporality that relies on the sequential actions of kinesin-14, kinesin-5, and involves both microtubule-kinetochore and kinetochore-kinetochore interactions. Upon quiescence exit, the microtubule bundle is disassembled via a cooperative process involving kinesin-8 and its full disassembly is required prior to cells re-entry into proliferation. Overall, our study provides the first description, at the molecular scale, of the entire life cycle of a stable microtubule structure in vivo and sheds light on its physiological function.
    Keywords:  S. cerevisiae; cell biology; kinesin; microtubules; quiescence
    DOI:  https://doi.org/10.7554/eLife.89958
  13. PLoS One. 2024 ;19(3): e0301084
    Spindle checkpoint activation by fungal orthologs of the S. cerevisiae Mps1 kinase.
    Amy Fabritius, Anabel Alonso, Andrew Wood, Shaheen Sulthana, Mark Winey.
      There is an ongoing need for antifungal agents to treat humans. Identification of new antifungal agents can be based on screening compounds using whole cell assays. Screening compounds that target a particular molecule is possible in budding yeast wherein sophisticated strain engineering allows for controlled expression of endogenous or heterologous genes. We have considered the yeast Mps1 protein kinase as a reasonable target for antifungal agents because mutant or druggable forms of the protein, upon inactivation, cause rapid loss of cell viability. Furthermore, extensive analysis of the Mps1 in budding yeast has offered potential tactics for identifying inhibitors of its enzymatic activity. One such tactic is based on the finding that overexpression of Mps1 leads to cell cycle arrest via activation of the spindle assembly checkpoint. We have endeavored to adapt this assay to be based on the overexpression of Mps1 orthologs from pathogenic yeast in hopes of having a whole-cell assay system to test the activity of these orthologs. Mps1 orthologous genes from seven pathogenic yeast or other pathogenic fungal species were isolated and expressed in budding yeast. Two orthologs clearly produced phenotypes similar to those produced by the overexpression of budding yeast Mps1, indicating that this system for heterologous Mps1 expression has potential as a platform for identifying prospective antifungal agents.
    DOI:  https://doi.org/10.1371/journal.pone.0301084
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