bims-micesi Biomed News
on Mitotic cell signalling
Issue of 2024–04–28
eleven papers selected by
Valentina Piano, Uniklinik Köln
  1. Molecular interplay between HURP and Kif18A in mitotic spindle regulation.
  2. Confinement in fibrous environments positions and orients mitotic spindles.
  3. Rules of engagement for condensins and cohesins guide mitotic chromosome formation.
  4. Live Cell Monitoring of Separase Activity, a Key Enzymatic Reaction for Chromosome Segregation, with Chimeric FRET-Based Molecular Sensor upon Cell Cycle Progression.
  5. Mitotic Functions and Characters of KIF11 in Cancers.
  6. Non-canonical role for the BAF complex subunit DPF3 in mitosis and ciliogenesis.
  7. The Kinetochore Proteins KNL-1 and Ndc80 Complex are Required for the Proper Positioning of Axons and Neuronal Cell Body in the C. elegans Nervous System.
  8. Changes in spindle morphology driven by TPX2 overexpression in MYC-driven breast cancer cells.
  9. A conserved germline-specific Dsn1 alternative splice isoform supports oocyte and embryo development.
  10. The centrosome: a critical hub for cell cycle regulation.
  11. The Rac1 homolog CED-10 is a component of the MES-1/SRC-1 pathway for asymmetric division of the C. elegans EMS blastomere.
  1. bioRxiv. 2024 Apr 11. pii: 2024.04.11.589088. [Epub ahead of print]
    Molecular interplay between HURP and Kif18A in mitotic spindle regulation.
    Juan M Perez-Bertoldi, Yuanchang Zhao, Akanksha Thawani, Ahmet Yildiz, Eva Nogales.
      During mitosis, microtubule dynamics are regulated to ensure proper alignment and segregation of chromosomes. The dynamics of kinetochore-attached microtubules are regulated by hepatoma-upregulated protein (HURP) and the mitotic kinesin-8 Kif18A, but the underlying mechanism remains elusive. Using single-molecule imaging in vitro , we demonstrate that Kif18A motility is regulated by HURP. While sparse decoration of HURP activates the motor, higher concentrations hinder processive motility. To shed light on this behavior, we determined the binding mode of HURP to microtubules using Cryo-EM. The structure reveals that one HURP motif spans laterally across β-tubulin, while a second motif binds between adjacent protofilaments. HURP partially overlaps with the microtubule-binding site of the Kif18A motor domain, indicating that excess HURP inhibits Kif18A motility by steric exclusion. We also observed that HURP and Kif18A function together to suppress dynamics of the microtubule plus-end, providing a mechanistic basis for how they collectively serve in spindle length control.
    DOI:  https://doi.org/10.1101/2024.04.11.589088
  2. bioRxiv. 2024 Apr 15. pii: 2024.04.12.589246. [Epub ahead of print]
    Confinement in fibrous environments positions and orients mitotic spindles.
    Apurba Sarkar, Aniket Jana, Atharva Agashe, Ji Wang, Rakesh Kapania, Nir S Gov, Jennifer G DeLuca, Raja Paul, Amrinder S Nain.
      Accurate positioning of the mitotic spindle within the rounded cell body is critical to physiological maintenance. Adherent mitotic cells encounter confinement from neighboring cells or the extracellular matrix (ECM), which can cause rotation of mitotic spindles and, consequently, titling of the metaphase plate (MP). To understand the positioning and orientation of mitotic spindles under confinement by fibers (ECM-confinement), we use flexible ECM-mimicking nanofibers that allow natural rounding of the cell body while confining it to differing levels. Rounded mitotic bodies are anchored in place by actin retraction fibers (RFs) originating from adhesion clusters on the ECM-mimicking fibers. We discover the extent of ECM-confinement patterns RFs in 3D: triangular and band-like at low and high confinement, respectively. A stochastic Monte-Carlo simulation of the centrosome (CS), chromosome (CH), membrane interactions, and 3D arrangement of RFs on the mitotic body recovers MP tilting trends observed experimentally. Our mechanistic analysis reveals that the 3D shape of RFs is the primary driver of the MP rotation. Under high ECM-confinement, the fibers can mechanically pinch the cortex, causing the MP to have localized deformations at contact sites with fibers. Interestingly, high ECM-confinement leads to low and high MP tilts, which mechanistically depend upon the extent of cortical deformation, RF patterning, and MP position. We identify that cortical deformation and RFs work in tandem to limit MP tilt, while asymmetric positioning of MP leads to high tilts. Overall, we provide fundamental insights into how mitosis may proceed in fibrous ECM-confining microenvironments in vivo.
    DOI:  https://doi.org/10.1101/2024.04.12.589246
  3. bioRxiv. 2024 Apr 20. pii: 2024.04.18.590027. [Epub ahead of print]
    Rules of engagement for condensins and cohesins guide mitotic chromosome formation.
    Kumiko Samejima, Johan H Gibcus, Sameer Abraham, Fernanda Cisneros-Soberanis, Itaru Samejima, Alison J Beckett, Nina Pučeková, Maria Alba Abad, Bethan Medina-Pritchard, James R Paulson, Linfeng Xie, A Arockia Jeyaprakash, Ian A Prior, Leonid A Mirny, Job Dekker, Anton Goloborodko, William C Earnshaw.
      During mitosis, interphase chromatin is rapidly converted into rod-shaped mitotic chromosomes. Using Hi-C, imaging, proteomics and polymer modeling, we determine how the activity and interplay between loop-extruding SMC motors accomplishes this dramatic transition. Our work reveals rules of engagement for SMC complexes that are critical for allowing cells to refold interphase chromatin into mitotic chromosomes. We find that condensin disassembles interphase chromatin loop organization by evicting or displacing extrusive cohesin. In contrast, condensin bypasses cohesive cohesins, thereby maintaining sister chromatid cohesion while separating the sisters. Studies of mitotic chromosomes formed by cohesin, condensin II and condensin I alone or in combination allow us to develop new models of mitotic chromosome conformation. In these models, loops are consecutive and not overlapping, implying that condensins do not freely pass one another but stall upon encountering each other. The dynamics of Hi-C interactions and chromosome morphology reveal that during prophase loops are extruded in vivo at ∼1-3 kb/sec by condensins as they form a disordered discontinuous helical scaffold within individual chromatids.
    DOI:  https://doi.org/10.1101/2024.04.18.590027
  4. Biosensors (Basel). 2024 Apr 15. pii: 192. [Epub ahead of print]14(4):
    Live Cell Monitoring of Separase Activity, a Key Enzymatic Reaction for Chromosome Segregation, with Chimeric FRET-Based Molecular Sensor upon Cell Cycle Progression.
    Md Shazadur Rahman, Yutaka Shindo, Kotaro Oka, Wataru Ikeda, Miho Suzuki.
      Separase is a key cysteine protease in the separation of sister chromatids through the digestion of the cohesin ring that inhibits chromosome segregation as a trigger of the metaphase-anaphase transition in eukaryotes. Its activity is highly regulated by binding with securin and cyclinB-CDK1 complex. These bindings prevent the proteolytic activity of separase until the onset of anaphase. Chromosome missegregation and aneuploidy are frequently observed in malignancies. However, there are some difficulties in biochemical examinations due to the instability of separase in vitro and the fact that few spatiotemporal resolution approaches exist for monitoring live separase activity throughout mitotic processes. Here, we have developed FRET-based molecular sensors, including GFP variants, with separase-cleavable sequences as donors and covalently attached fluorescent dyes as acceptor molecules. These are applicable to conventional live cell imaging and flow cytometric analysis because of efficient live cell uptake. We investigated the performance of equivalent molecular sensors, either localized or not localized inside the nucleus under cell cycle control, using flow cytometry. Synchronized cell cycle progression rendered significant separase activity detections in both molecular sensors. We obtained consistent outcomes with localized molecular sensor introduction and cell cycle control by fluorescent microscopic observations. We thus established live cell separase activity monitoring systems that can be used specifically or statistically, which could lead to the elucidation of separase properties in detail.
    Keywords:  CDK-1; FRET; cell cycle; chromosome segregation; cohesion; cyclin B1; live cell sensing; securin; separase
    DOI:  https://doi.org/10.3390/bios14040192
  5. Biomolecules. 2024 Mar 22. pii: 386. [Epub ahead of print]14(4):
    Mitotic Functions and Characters of KIF11 in Cancers.
    Wanting Gao, Junjie Lu, Zitao Yang, Enmin Li, Yufei Cao, Lei Xie.
      Mitosis mediates the accurate separation of daughter cells, and abnormalities are closely related to cancer progression. KIF11, a member of the kinesin family, plays a vital role in the formation and maintenance of the mitotic spindle. Recently, an increasing quantity of data have demonstrated the upregulated expression of KIF11 in various cancers, promoting the emergence and progression of cancers. This suggests the great potential of KIF11 as a prognostic biomarker and therapeutic target. However, the molecular mechanisms of KIF11 in cancers have not been systematically summarized. Therefore, we first discuss the functions of the protein encoded by KIF11 during mitosis and connect the abnormal expression of KIF11 with its clinical significance. Then, we elucidate the mechanism of KIF11 to promote various hallmarks of cancers. Finally, we provide an overview of KIF11 inhibitors and outline areas for future work.
    Keywords:  KIF11; angiogenesis; cancer; epithelial–mesenchymal transition; mitosis
    DOI:  https://doi.org/10.3390/biom14040386
  6. J Cell Sci. 2024 Apr 25. pii: jcs.261744. [Epub ahead of print]
    Non-canonical role for the BAF complex subunit DPF3 in mitosis and ciliogenesis.
    Giulia Verrillo, Anna Maria Obeid, Alexia Genco, Jacopo Scrofani, François Orange, Sarah Hanache, Julien Mignon, Tanguy Leyder, Catherine Michaux, Céline Kempeneers, Noëmie Bricmont, Stephanie Herkenne, Isabelle Vernos, Maud Martin, Denis Mottet.
      DPF3, along with other subunits, is a well-known component of the BAF chromatin remodeling complex that plays a key role in regulating chromatin remodeling activity and gene expression. Here, we elucidated a non-canonical localization and role for DPF3. We showed that DPF3 dynamically localizes to the centriolar satellites in interphase and in centrosome, spindle midzone/bridging fiber area and midbodies during mitosis. Loss of DPF3 causes K-fiber instability, unstable kinetochore-microtubules attachment and defects in chromosome alignment, thus resulting in altered mitotic progression, cell death and genomic instability. In addition, we also demonstrated that DPF3 localizes in centriolar satellites at the basis of primary cilia and is required for ciliogenesis by regulating axoneme extension. Together, these findings uncover a moonlighting dual function for DPF3 during mitosis and ciliogenesis.
    Keywords:  Bridging Fiber; Centriolar satellites; Chromosome alignment; Ciliogenesis; DPF3; Genomic instability; Mitosis
    DOI:  https://doi.org/10.1242/jcs.261744
  7. Mol Biol Cell. 2024 Apr 24. mbcE23080325
    The Kinetochore Proteins KNL-1 and Ndc80 Complex are Required for the Proper Positioning of Axons and Neuronal Cell Body in the C. elegans Nervous System.
    Vasileios R Ouzounidis, Mattie Green, Charlotte de Ceuninck van Capelle, Clara Gebhardt, Helena Crellin, Cameron Finlayson, Bram Prevo, Dhanya K Cheerambathur.
      The KMN (Knl1/Mis12/Ndc80) network at the kinetochore, primarily known for its role in chromosome segregation, has been shown to be repurposed during neurodevelopment. Here, we investigate the underlying neuronal mechanism and show that the KMN network promotes the proper axonal organization within the C. elegans head nervous system. Post-mitotic degradation of KNL-1, which acts as a scaffold for signaling and has microtubule-binding activities at the kinetochore, led to disorganized ganglia and aberrant placement and organization of axons in the nerve ring - an interconnected axonal network. Through gene-replacement approaches, we demonstrate that the signaling motifs within KNL-1, responsible for recruiting the protein phosphatase 1, and activating the spindle assembly checkpoint are required for neurodevelopment. Interestingly, while the microtubule-binding activity is crucial to KMN's neuronal function, microtubule dynamics and organization were unaffected in the absence of KNL-1. Instead, the NDC-80 microtubule-binding mutant displayed notable defects in axon bundling during nerve ring formation, indicating its role in facilitating axon-axon contacts. Overall, these findings provide evidence for a non-canonical role for the KMN network in shaping the structure and connectivity of the nervous system in C. elegans during brain development.
    DOI:  https://doi.org/10.1091/mbc.E23-08-0325
  8. MicroPubl Biol. 2024 ;2024
    Changes in spindle morphology driven by TPX2 overexpression in MYC-driven breast cancer cells.
    Guadalupe Pena, Julia Rohrberg, Andrei Goga, Rebecca Heald.
      The MYC oncogene was previously shown to induce mitotic spindle defects, chromosome instability, and reliance on the microtubule-associated protein TPX2 to survive, but how TPX2 levels affect spindle morphology in cancer cells has not previously been examined in detail. We show that breast cancer cell lines expressing high levels of MYC and TPX2 possess shorter spindles with increased TPX2 localization at spindle poles. A similar effect was observed in non-transformed human RPE-1 cells compared to a tumor cell line (HeLa) that overexpresses MYC . These results demonstrate that TPX2 alters spindle length and morphology in cancer cells, which may contribute their ability to divide despite MYC-induced mitotic stress.
    DOI:  https://doi.org/10.17912/micropub.biology.001182
  9. bioRxiv. 2024 Apr 18. pii: 2024.04.17.589883. [Epub ahead of print]
    A conserved germline-specific Dsn1 alternative splice isoform supports oocyte and embryo development.
    Jimmy Ly, Cecilia S Blengini, Sarah L Cady, Karen Schindler, Iain M Cheeseman.
      Alternative mRNA splicing can generate distinct protein isoforms to allow for the differential control of cell processes across cell types. However, alternative splice isoforms that differentially modulate distinct cell division programs have remained elusive. Here, we demonstrate that mammalian germ cells express an alternate mRNA splice isoform for the kinetochore component, DSN1, a subunit of the MIS12 complex that links the centromeres to spindle microtubules during chromosome segregation. This germline DSN1 isoform bypasses the requirement for Aurora kinase phosphorylation for its centromere localization due to the absence of a key regulatory region allowing DSN1 to display persistent centromere localization. Expression of the germline DSN1 isoform in somatic cells results in constitutive kinetochore localization, chromosome segregation errors, and growth defects, providing an explanation for its tight cell type-specific expression. Reciprocally, precisely eliminating expression of the germline DSN1 splice isoform in mouse models disrupts oocyte maturation and early embryonic divisions coupled with a reduction in fertility. Together, this work identifies a germline-specific splice isoform for a chromosome segregation component and implicates its role in mammalian fertility.
    DOI:  https://doi.org/10.1101/2024.04.17.589883
  10. Trends Cell Biol. 2024 Apr 25. pii: S0962-8924(24)00072-2. [Epub ahead of print]
    The centrosome: a critical hub for cell cycle regulation.
    Yuki Yoshino, Zhenzhou Fang, Natsuko Chiba.
      Cyclins and cyclin-dependent kinases (CDKs) localize to the centrosome, but their significance in the cell cycle is unclear. Recently, Roberts et al. revealed that centrosomal cyclin B-CDK is required for mitotic entry and phosphorylation of substrates. This suggests that the centrosome acts as a signaling hub controlling the cell cycle.
    Keywords:  cell cycle; centrosome; cyclin; cyclin-dependent kinase; spindle pole body
    DOI:  https://doi.org/10.1016/j.tcb.2024.04.003
  11. bioRxiv. 2024 Apr 08. pii: 2024.04.04.588162. [Epub ahead of print]
    The Rac1 homolog CED-10 is a component of the MES-1/SRC-1 pathway for asymmetric division of the C. elegans EMS blastomere.
    Helen Lamb, Małgorzata Liro, Krista Myles, McKenzi Fernholz, Holly Anderson, Lesilee S Rose.
      Asymmetric cell division is essential for the creation of cell types with different identities and functions. The EMS blastomere of the four-cell Caenorhabditis elegans embryo undergoes an asymmetric division in response to partially redundant signaling pathways. One pathway involves a Wnt signal emanating from the neighboring P2 cell, while the other pathway is defined by the receptor-like MES-1 protein localized at the EMS/P2 cell contact, and the cytoplasmic kinase SRC-1. In response to these pathways, the EMS nuclear-centrosome complex rotates so that the spindle forms on the anterior-posterior axis; after division, the daughter cell contacting P2 becomes the endodermal precursor cell. Here we identify the Rac1 homolog,CED-10, as a new component of the MES-1/SRC-1 pathway. Loss of CED-10 affects both spindle positioning and endoderm specification. Although MES-1 is still present at the EMS/P2 contact in ced-10 embryos, SRC-1 dependent phosphorylation is reduced. These and other results suggest that CED-10 acts downstream of MES-1 and upstream or at the level of SRC-1 activity. In addition, we find that the branched actin regulator ARX-2 is enriched at the EMS/P2 cell contact site, in a CED-10 dependent manner. Loss of ARX-2 results in spindle positioning defects, suggesting that CED-10 acts through branched actin to promote the asymmetric division of the EMS cell.
    DOI:  https://doi.org/10.1101/2024.04.04.588162
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