bims-micesi Biomed News
on Mitotic cell signalling
Issue of 2022–01–16
twelve papers selected by
Valentina Piano, Max Planck Institute of Molecular Physiology
  1. Microtubule organizing centers regulate spindle positioning in mouse oocytes.
  2. The mitotic spindle protein CKAP2 potently increases formation and stability of microtubules.
  3. A Novel Hyperactive Nud1 Mitotic Exit Network Scaffold Causes Spindle Position Checkpoint Bypass in Budding Yeast.
  4. Mechanical Torque Promotes Bipolarity of the Mitotic Spindle Through Multi-centrosomal Clustering.
  5. Molecular and cellular dynamics of early embryonic cell divisions in Volvox carteri.
  6. Rigosertib and Cholangiocarcinoma: A Cell Cycle Affair.
  7. Association of mitotic checkpoint regulator MAD2L1 with fibrosis progression in chronic hepatitis C patients.
  8. G2/M arrest and mitotic slippage induced by fenbendazole in canine melanoma cells.
  9. Dose-dependent Transmissibility of Chromosome Aberrations at First Mitosis after Exposure to Gamma Rays. I. Modeling and Implications Related to Risk Assessment.
  10. Incorporation of CENP-A/CID into centromeres during early Drosophila embryogenesis does not require RNA polymerase II-mediated transcription.
  11. The cell cycle-related genes RHAMM, AURKA, TPX2, PLK1, and PLK4 are associated with the poor prognosis of breast cancer patients.
  12. Pristine Cu-MOF Induces Mitotic Catastrophe and Alterations of Gene Expression and Cytoskeleton in Ovarian Cancer Cells.
  1. Dev Cell. 2021 Dec 28. pii: S1534-5807(21)01031-5. [Epub ahead of print]
    Microtubule organizing centers regulate spindle positioning in mouse oocytes.
    Daniela Londoño-Vásquez, Katherine Rodriguez-Lukey, Susanta K Behura, Ahmed Z Balboula.
      During female meiosis I (MI), spindle positioning must be tightly regulated to ensure the fidelity of the first asymmetric division and faithful chromosome segregation. Although the role of F-actin in regulating these critical processes has been studied extensively, little is known about whether microtubules (MTs) participate in regulating these processes. Using mouse oocytes as a model system, we characterize a subset of MT organizing centers that do not contribute directly to spindle assembly, termed mcMTOCs. Using laser ablation, STED super-resolution microscopy, and chemical manipulation, we show that mcMTOCs are required to regulate spindle positioning and faithful chromosome segregation during MI. We discuss how forces exerted by F-actin on the spindle are balanced by mcMTOC-nucleated MTs to anchor the spindle centrally and to regulate its timely migration. Our findings provide a model for asymmetric cell division, complementing the current F-actin-based models, and implicate mcMTOCs as a major player in regulating spindle positioning.
    Keywords:  MTOC; cytoplasmic MTOC; meiosis; microtubule; oocyte; spindle migration; spindle positioning
    DOI:  https://doi.org/10.1016/j.devcel.2021.12.011
  2. Elife. 2022 Jan 14. pii: e72202. [Epub ahead of print]11
    The mitotic spindle protein CKAP2 potently increases formation and stability of microtubules.
    Thomas S McAlear, Susanne Bechstedt.
      Cells increase microtubule dynamics to make large rearrangements to their microtubule cytoskeleton during cell division. Changes in microtubule dynamics are essential for the formation and function of the mitotic spindle, and misregulation can lead to aneuploidy and cancer. Using in vitro reconstitution assays we show that the mitotic spindle protein Cytoskeleton-Associated Protein 2 (CKAP2) has a strong effect on nucleation of microtubules by lowering the critical tubulin concentration 100-fold. CKAP2 increases the apparent rate constant ka of microtubule growth by 50-fold and increases microtubule growth rates. In addition, CKAP2 strongly suppresses catastrophes. Our results identify CKAP2 as the most potent microtubule growth factor to date. These finding help explain CKAP2's role as an important spindle protein, proliferation marker, and oncogene.
    Keywords:  biochemistry; cell biology; chemical biology; none
    DOI:  https://doi.org/10.7554/eLife.72202
  3. Cells. 2021 Dec 24. pii: 46. [Epub ahead of print]11(1):
    A Novel Hyperactive Nud1 Mitotic Exit Network Scaffold Causes Spindle Position Checkpoint Bypass in Budding Yeast.
    Michael Vannini, Victoria R Mingione, Ashleigh Meyer, Courtney Sniffen, Jenna Whalen, Anupama Seshan.
      Mitotic exit is a critical cell cycle transition that requires the careful coordination of nuclear positioning and cyclin B destruction in budding yeast for the maintenance of genome integrity. The mitotic exit network (MEN) is a Ras-like signal transduction pathway that promotes this process during anaphase. A crucial step in MEN activation occurs when the Dbf2-Mob1 protein kinase complex associates with the Nud1 scaffold protein at the yeast spindle pole bodies (SPBs; centrosome equivalents) and thereby becomes activated. This requires prior priming phosphorylation of Nud1 by Cdc15 at SPBs. Cdc15 activation, in turn, requires both the Tem1 GTPase and the Polo kinase Cdc5, but how Cdc15 associates with SPBs is not well understood. We have identified a hyperactive allele of NUD1, nud1-A308T, that recruits Cdc15 to SPBs in all stages of the cell cycle in a CDC5-independent manner. This allele leads to early recruitment of Dbf2-Mob1 during metaphase and requires known Cdc15 phospho-sites on Nud1. The presence of nud1-A308T leads to loss of coupling between nuclear position and mitotic exit in cells with mispositioned spindles. Our findings highlight the importance of scaffold regulation in signaling pathways to prevent improper activation.
    Keywords:  Cdc15; Dbf2; MEN; Mob1; Nud1; mitotic exit; spindle position checkpoint
    DOI:  https://doi.org/10.3390/cells11010046
  4. Bull Math Biol. 2022 Jan 10. 84(2): 29
    Mechanical Torque Promotes Bipolarity of the Mitotic Spindle Through Multi-centrosomal Clustering.
    Christopher E Miles, Jie Zhu, Alex Mogilner.
      Intracellular forces shape cellular organization and function. One example is the mitotic spindle, a cellular machine consisting of multiple chromosomes and centrosomes which interact via dynamic microtubule filaments and motor proteins, resulting in complicated spatially dependent forces. For a cell to divide properly, it is important for the spindle to be bipolar, with chromosomes at the center and multiple centrosomes clustered into two 'poles' at opposite sides of the chromosomes. Experimental observations show that in unhealthy cells, the spindle can take on a variety of patterns. What forces drive each of these patterns? It is known that attraction between centrosomes is key to bipolarity, but what prevents the centrosomes from collapsing into a monopolar configuration? Here, we explore the hypothesis that torque rotating chromosome arms into orientations perpendicular to the centrosome-centromere vector promotes spindle bipolarity. To test this hypothesis, we construct a pairwise-interaction model of the spindle. On a continuum version of the model, an integro-PDE system, we perform linear stability analysis and construct numerical solutions which display a variety of spatial patterns. We also simulate a discrete particle model resulting in a phase diagram that confirms that the spindle bipolarity emerges most robustly with torque. Altogether, our results suggest that rotational forces may play an important role in dictating spindle patterning.
    Keywords:  Collective behavior; Intracellular mechanics; Mitotic spindle; Multi-centrosomal clustering
    DOI:  https://doi.org/10.1007/s11538-021-00985-2
  5. Plant Cell. 2022 Jan 09. pii: koac004. [Epub ahead of print]
    Molecular and cellular dynamics of early embryonic cell divisions in Volvox carteri.
    Eva Laura von der Heyde, Armin Hallmann.
      Cell division is fundamental to all organisms, and the green alga used here exhibits both key animal and plant functions. Specifically, we analyzed the molecular and cellular dynamics of early embryonic divisions of the multicellular green alga Volvox carteri (Chlamydomonadales). Relevant proteins related to mitosis and cytokinesis were identified in silico, the corresponding genes were cloned, fused to yfp, and stably expressed in Volvox, and the tagged proteins were studied by live-cell imaging. We reveal rearrangements of the microtubule cytoskeleton during centrosome separation, spindle formation, establishment of the phycoplast and generation of previously unknown structures. The centrosomes participate in initiation of spindle formation and determination of spindle orientation. Although the nuclear envelope does not break down during early mitosis, intermixing of cytoplasm and nucleoplasm results in loss of nuclear identity. Finally, we present a model for mitosis in Volvox. Our study reveals enormous dynamics, clarifies spatio-temporal relationships of subcellular structures and provides insight into the evolution of cell division.
    DOI:  https://doi.org/10.1093/plcell/koac004
  6. Int J Mol Sci. 2021 Dec 25. pii: 213. [Epub ahead of print]23(1):
    Rigosertib and Cholangiocarcinoma: A Cell Cycle Affair.
    Alessio Malacrida, Guido Cavaletti, Mariarosaria Miloso.
      Rigosertib is multi-kinase inhibitor that could represent an interesting therapeutic option for non-resectable patients with cholangiocarcinoma, a very aggressive hepatic cancer with limited effective treatments. The Western blotting technique was used to evaluate alterations in the expression of proteins involved in the regulation of the cell cycle of cholangiocarcinoma EGI-1 cells. Our results show an increase in EMI1 and Cyclin B protein levels after Rigosertib treatment. Moreover, the phosphorylation of CDK1 is significantly reduced by Rigosertib, while PLK1 expression increased after 24 h of treatment and decreased after 48 h. Finally, we evaluated the role of p53. Its levels increase after Rig treatment, and, as shown in the cell viability experiment with the p53 inhibitor Pifithrin, its activity is necessary for the effects of Rigosertib against the cell viability of EGI-1 cells. In conclusion, we hypothesized the mechanism of the action of Rigosertib against cholangiocarcinoma EGI-1 cells, highlighting the importance of proteins involved in the regulation of cell cycles. The CDK1-Cyclin B complex and p53 play an important role, explaining the Block in the G2/M phase of the cell cycle and the effect on cell viability.
    Keywords:  CDK1; Cyclin B; EGI-1 cells; EMI1; PLK1 inhibitor; Rigosertib; cell cycle; cholangiocarcinoma; p53
    DOI:  https://doi.org/10.3390/ijms23010213
  7. Clin Res Hepatol Gastroenterol. 2022 Jan 06. pii: S2210-7401(22)00003-1. [Epub ahead of print] 101860
    Association of mitotic checkpoint regulator MAD2L1 with fibrosis progression in chronic hepatitis C patients.
    Saira S Khalid, Ayman A Abdo, Waleed Al-Hamoudi, Khalid Alswat.
      
    Keywords:  Hepatitis C virus; cell cycle inhibitors; cell cycle proteins; chronic hepatitis C; microRNAs; mitosis regulator
    DOI:  https://doi.org/10.1016/j.clinre.2022.101860
  8. Vet Med Sci. 2022 Jan 12.
    G2/M arrest and mitotic slippage induced by fenbendazole in canine melanoma cells.
    Sehoon Kim, Shashini Kanchanamala Perera, Seo-In Choi, Robert B Rebhun, Kyoung-Won Seo.
       BACKGROUND: The use of fenbendazole (FBZ) in terminal cancer patients has recently increased, as anthelminthic drugs, such as FBZ and benzimidazole, exhibit anti-tubulin effects in tumour cells.
    OBJECTIVES: The present study evaluated the in vitro anti-cancer effects of FBZ in five canine melanoma cell lines originating from the oral cavity (UCDK9M3, UCDK9M4, UCDK9M5, KMeC and LMeC).
    METHODS: Five canine melanoma cell lines were treated with FBZ and analysed with cell viability assay, cell cycle analysis, western blot assay and immunofluorescence staining to identify apoptotic effect, cell cycle arrest, microtubule disruption and mitotic slippage.
    RESULTS: Cell viability was reduced in all melanoma cell lines in a dose-dependent manner after FBZ treatment. Through cell cycle analysis, G2/M arrest and mitotic slippage were identified, which showed a time-dependent change. All treatment concentrations induced increased cleaved PARP signals in western blot analysis compared to the control groups. Immunofluorescence of cells treated for 24 h revealed defects in microtubule structure, multinucleation or macronucleation. With the exception of UCDK9M3, the melanoma cells showed mitotic slippage and post-slippage death, indicative of mitotic catastrophe.
    CONCLUSIONS: These results indicate that FBZ exhibits anti-cancer effects in vitro against canine melanoma cells; however, further in vivo studies regarding the clinical applications of FBZ are required.
    Keywords:  G2/M cell cycle arrest; anti-tumour effect; apoptosis; canine melanoma cell; fenbendazole; mitotic slippage
    DOI:  https://doi.org/10.1002/vms3.733
  9. Radiat Res. 2022 Jan 14.
    Dose-dependent Transmissibility of Chromosome Aberrations at First Mitosis after Exposure to Gamma Rays. I. Modeling and Implications Related to Risk Assessment.
    Bradford D Loucas, Igor Shuryak, Stephen R Kunkel, Michael N Cornforth.
      The relationship between certain chromosomal aberration (CA) types and cell lethality is well established. On that basis we used multi-fluor in situ hybridization (mFISH) to tally the number of mitotic human lymphocytes exposed to graded doses of gamma rays that carried either lethal or nonlethal CA types. Despite the fact that a number of nonlethal complex exchanges were observed, the cells containing them were seldom deemed viable, due to coincident lethal chromosome damage. We considered two model variants for describing the dose responses. The first assumes independent linear-quadratic (LQ) dose response shapes for the yields of both lethal and nonlethal CAs. The second (simplified) variant assumes that the mean number of nonlethal CAs per cell is proportional to the mean number of lethal CAs per cell, meaning that the shapes and magnitudes of both aberration types differ only by a multiplicative proportionality constant. Using these models allowed us to assemble dose response curves for the frequency of aberration-bearing cells that would be expected to survive. This took the form of a joint probability distribution for cells containing ≥1 nonlethal CAs but having zero lethal CAs. The simplified second model variant turned out to be marginally better supported than the first, and the joint probability distribution based on this model yielded a crescent-shaped dose response reminiscent of those observed for mutagenesis and transformation for cells "at risk" (i.e. not corrected for survival). Among the implications of these findings is the suggestion that similarly shaped curves form the basis for deriving metrics associated with radiation risk models.
    DOI:  https://doi.org/10.1667/RADE-21-00180.1
  10. Chromosoma. 2022 Jan 11.
    Incorporation of CENP-A/CID into centromeres during early Drosophila embryogenesis does not require RNA polymerase II-mediated transcription.
    Samadri Ghosh, Christian F Lehner.
      In many species, centromere identity is specified epigenetically by special nucleosomes containing a centromere-specific histone H3 variant, designated as CENP-A in humans and CID in Drosophila melanogaster. After partitioning of centromere-specific nucleosomes onto newly replicated sister centromeres, loading of additional CENP-A/CID into centromeric chromatin is required for centromere maintenance in proliferating cells. Analyses with cultured cells have indicated that transcription of centromeric DNA by RNA polymerase II is required for deposition of new CID into centromere chromatin. However, a dependence of centromeric CID loading on transcription is difficult to reconcile with the notion that the initial embryonic stages appear to proceed in the absence of transcription in Drosophila, as also in many other animal species. To address the role of RNA polymerase II-mediated transcription for CID loading in early Drosophila embryos, we have quantified the effects of alpha-amanitin and triptolide on centromeric CID-EGFP levels. Our analyses demonstrate that microinjection of these two potent inhibitors of RNA polymerase II-mediated transcription has at most a marginal effect on centromeric CID deposition during progression through the early embryonic cleavage cycles. Thus, we conclude that at least during early Drosophila embryogenesis, incorporation of CID into centromeres does not depend on RNA polymerase II-mediated transcription.
    Keywords:  Alpha-amanitin; CENP-A/CID deposition; Centromere chromatin; Centromere transcription; Triptolide
    DOI:  https://doi.org/10.1007/s00412-022-00767-2
  11. J Cell Biochem. 2022 Jan 10.
    The cell cycle-related genes RHAMM, AURKA, TPX2, PLK1, and PLK4 are associated with the poor prognosis of breast cancer patients.
    Iris Kahl, Julian Mense, Christopher Finke, Anna-Lena Boller, Clara Lorber, Balázs Győrffy, Burkhard Greve, Martin Götte, Nancy A Espinoza-Sánchez.
      Breast cancer is the third most common type of cancer diagnosed. Cell cycle is a complex but highly organized and controlled process, in which normal cells sense mitogenic growth signals that instruct them to enter and progress through their cell cycle. This process culminates in cell division generating two daughter cells with identical amounts of genetic material. Uncontrolled proliferation is one of the hallmarks of cancer. In this study, we analyzed the expression of the cell cycle-related genes receptor for hyaluronan (HA)-mediated motility (RHAMM), AURKA, TPX2, PLK1, and PLK4 and correlated them with the prognosis in a collective of 3952 breast cancer patients. A high messenger RNA expression of all studied genes correlated with a poor prognosis. Stratifying the patients according to the expression of hormonal receptors, we found that in patients with estrogen and progesterone receptor-positive and human epithelial growth factor receptor 2-negative tumors, and Luminal A and Luminal B tumors, the expression of the five analyzed genes correlates with worse survival. qPCR analysis of a panel of breast cancer cell lines representative of major molecular subtypes indicated a predominant expression in the luminal subtype. In vitro experiments showed that radiation influences the expression of the five analyzed genes both in luminal and triple-negative model cell lines. Functional analysis of MDA-MB-231 cells showed that small interfering RNA knockdown of PLK4 and TPX2 and pharmacological inhibition of PLK1 had an impact on the cell cycle and colony formation. Looking for a potential upstream regulation by microRNAs, we observed a differential expression of RHAMM, AURKA, TPX2, PLK1, and PLK4 after transfecting the MDA-MB-231 cells with three different microRNAs. Survival analysis of miR-34c-5p, miR-375, and miR-142-3p showed a different impact on the prognosis of breast cancer patients. Our study suggests that RHAMM, AURKA, TPX2, PLK1, and PLK4 can be used as potential targets for treatment or as a prognostic value in breast cancer patients.
    Keywords:  KM-plotter; breast cancer; cell cycle check points; miRNAs; radiation; survival analysis
    DOI:  https://doi.org/10.1002/jcb.30205
  12. ACS Appl Bio Mater. 2020 Jul 20. 3(7): 4081-4094
    Pristine Cu-MOF Induces Mitotic Catastrophe and Alterations of Gene Expression and Cytoskeleton in Ovarian Cancer Cells.
    Daomei Chen, Bin Li, Liang Jiang, Yizhou Li, Yepeng Yang, Zhifang Luo, Jiaqiang Wang.
      Metals-organic frameworks (MOFs) have been widely explored in biomedicine, mostly in drug delivery, biosensing, and bioimaging due to their large surface area, tunable porosity, readily chemical functionalization, and good biocompatibility. However, the underlining cellular mechanisms controlling the process for MOF cytotoxicity remains almost completely unknown. Here, we demonstrate that pristine Cu-MOF without any loaded drug selectively inhibited ovarian cancer mainly through promoting tubulin polymerization and destroying the cell actin cytoskeleton (F-actin) to trigger the mitotic catastrophe, accompanying by conventional programmed cell death. To our knowledge, this is the first report claiming that mitotic catastrophe may be an explaining mechanism of MOF cytotoxicity. Cu-MOF with an intrinsic protease-like activity also hydrolyzed cellular cytoskeleton proteins (F-actin). The RNA sequencing data indicated the differential expressional mRNA of cell proliferation and actin cytoskeleton (ACTA2, ACTN3, FSCN2, and SCIN) and mitotic spindles (PLK1 and TPX2) related genes. We found that Cu-MOF as a promising candidate in the disruption of cellular cytoskeleton and the change of the gene expression could be actin altering and antimitotic agents against cancer cells, allowing for fundamental biological and biophysical studies of MOFs.
    Keywords:  F-actin; MOFs; cells arrest; gene expression; mitotic catastrophe; ovarian cancer cells
    DOI:  https://doi.org/10.1021/acsabm.0c00175
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