bims-micesi Biomed News
on Mitotic cell signalling
Issue of 2023–04–02
thirteen papers selected by
Valentina Piano, Uniklinik Köln
  1. Aurora kinases: Generators of spatial control during mitosis.
  2. Kinetochore dynein is sufficient to biorient chromosomes and remodel the outer kinetochore.
  3. p97/VCP drives turnover of SUMOylated centromeric CCAN proteins and CENP-A.
  4. The Multiple Mitotic Roles of the ASPM Orthologous Proteins: Insight into the Etiology of ASPM-Dependent Microcephaly.
  5. Src inhibition induces mitotic arrest associated with chromosomal passenger complex.
  6. Mechanisms underlying spindle assembly and robustness.
  7. A Dynamic population of prophase CENP-C is required for meiotic chromosome segregation.
  8. HPV16 E6 induces chromosomal instability due to polar chromosomes caused by E6AP-dependent degradation of the mitotic kinesin CENP-E.
  9. Mitotic DNA damage promotes chromokinesin-mediated missegregation of polar chromosomes in cancer cells.
  10. RHINO restricts MMEJ activity to mitosis.
  11. Misaligned Chromosomes are a Major Source of Chromosomal Instability in Breast Cancer.
  12. Actin-Microtubule Crosstalk Imparts Stiffness to the Contractile Ring in Fission Yeast.
  13. Mitotic phosphorylation of Tau/MAPT modulates cell cycle progression in prostate cancer cells.
  1. Front Cell Dev Biol. 2023 ;11 1139367
    Aurora kinases: Generators of spatial control during mitosis.
    Aamir Ali, P Todd Stukenberg.
      Cell division events require regulatory systems to ensure that events happen in a distinct order. The classic view of temporal control of the cell cycle posits that cells order events by linking them to changes in Cyclin Dependent Kinase (CDK) activities. However, a new paradigm is emerging from studies of anaphase where chromatids separate at the central metaphase plate and then move to opposite poles of the cell. These studies suggest that distinct events are ordered depending upon the location of each chromosome along its journey from the central metaphase plate to the elongated spindle poles. This system is dependent upon a gradient of Aurora B kinase activity that emerges during anaphase and acts as a spatial beacon to control numerous anaphase/telophase events and cytokinesis. Recent studies also suggest that Aurora A kinase activity specifies proximity of chromosomes or proteins to spindle poles during prometaphase. Together these studies argue that a key role for Aurora kinases is to provide spatial information that controls events depending upon the location of chromosomes or proteins along the mitotic spindle.
    Keywords:  Aurora kinases; chromosome biorientation; kinase gradients; kinetochore-microtubule interactions; micronuclei; mitosis; mitotic spindle; nuclear envelope reformation
    DOI:  https://doi.org/10.3389/fcell.2023.1139367
  2. bioRxiv. 2023 Mar 24. pii: 2023.03.23.534015. [Epub ahead of print]
    Kinetochore dynein is sufficient to biorient chromosomes and remodel the outer kinetochore.
    Bram Prevo, Dhanya K Cheerambathur, William C Earnshaw, Arshad Desai.
      Multiple microtubule-directed activities concentrate on chromosomes during mitosis to ensure their accurate distribution to daughter cells. These activities include couplers and dynamics regulators localized at the kinetochore, the specialized microtubule interface built on centromeric chromatin, as well as motor proteins recruited to kinetochores and to mitotic chromatin. Here, we describe an in vivo reconstruction approach in which the effect of removing the major microtubule-directed activities on mitotic chromosomes is compared to the selective presence of individual activities. This approach revealed that the kinetochore dynein module, comprised of the minus end-directed motor cytoplasmic dynein and its kinetochore-specific adapters, is sufficient to biorient chromosomes and to remodel outer kinetochore composition following microtubule attachment; by contrast, the kinetochore dynein module is unable to support chromosome congression. The chromosome-autonomous action of kinetochore dynein, in the absence of the other major microtubule-directed factors on chromosomes, rotates and orients a substantial proportion of chromosomes such that their sister chromatids attach to opposite spindle poles. In tight coupling with orientation, the kinetochore dynein module drives removal of outermost kinetochore components, including the dynein motor itself and spindle checkpoint activators. The removal is independent of the other major microtubule-directed activities and kinetochore-localized protein phosphatase 1, suggesting that it is intrinsic to the kinetochore dynein module. These observations indicate that the kinetochore dynein module has the ability coordinate chromosome biorientation with attachment state-sensitive remodeling of the outer kinetochore that facilitates cell cycle progression.
    DOI:  https://doi.org/10.1101/2023.03.23.534015
  3. Mol Biol Cell. 2023 Mar 29. mbcE23010035
    p97/VCP drives turnover of SUMOylated centromeric CCAN proteins and CENP-A.
    Sebastiaan J W van den Berg, Samuel East, Sreyoshi Mitra, Lars E T Jansen.
      The centromere is a unique chromatin domain that links sister chromatids and forms the attachment site for spindle microtubules in mitosis. Centromere inheritance is largely DNA sequence-independent but strongly reliant on a self-propagating chromatin domain featuring nucleosomes containing the H3 variant CENP-A. Unlike other histones, CENP-A is maintained with unusually high stability in chromatin. Previously, we have shown that mitotic maintenance of CENP-A and other CCAN proteins is controlled by a dynamic SUMO cycle and that the deSUMOylase SENP6 is necessary for stable maintenance of CENP-A at the centromere. Here, we discover that the removal of SENP6 leads to a rapid loss of the constitutive centromere-associated network (CCAN), followed by a delayed loss of centromeric CENP-A, indicating that the CCAN is the primary SUMO target. We found the ATP-dependent segregase p97/VCP removes centromeric CENP-A in a SUMO-dependent manner and physically interacts with the CCAN and CENP-A chromatin. Our data suggest a direct role of p97 in removing centromeric CENP-A via SUMOylated CCAN proteins thereby ensuring centromere homeostasis and potentially preventing ectopic CENP-A accumulation.
    DOI:  https://doi.org/10.1091/mbc.E23-01-0035
  4. Cells. 2023 Mar 16. pii: 922. [Epub ahead of print]12(6):
    The Multiple Mitotic Roles of the ASPM Orthologous Proteins: Insight into the Etiology of ASPM-Dependent Microcephaly.
    Alyona V Razuvaeva, Lucia Graziadio, Valeria Palumbo, Gera A Pavlova, Julia V Popova, Alexey V Pindyurin, Silvia Bonaccorsi, Maria Patrizia Somma, Maurizio Gatti.
      The Drosophila abnormal spindle (asp) gene was discovered about 40 years ago and shown to be required for both mitotic and meiotic cell division. Subsequent studies showed that asp is highly conserved and that mutations in its human ortholog ASPM (Abnormal Spindle-like Microcephaly-associated; or MCPH5) are the most common cause of autosomal recessive primary microcephaly. This finding greatly stimulated research on ASPM and its fly and mouse (Aspm) orthologs. The three Asp orthologous proteins bind the microtubules (MTs) minus ends during cell division and also function in interphase nuclei. Investigations on different cell types showed that Asp/Aspm/ASPM depletion disrupts one or more of the following mitotic processes: aster formation, spindle pole focusing, centrosome-spindle coupling, spindle orientation, metaphase-to-anaphase progression, chromosome segregation, and cytokinesis. In addition, ASPM physically interacts with components of the DNA repair and replication machineries and is required for the maintenance of chromosomal DNA stability. We propose the working hypothesis that the asp/Aspm/ASPM genes play the same conserved functions in Drosophila, mouse, and human cells. Human microcephaly is a genetically heterogeneous disorder caused by mutations in 30 different genes that play a variety of functions required for cell division and chromosomal DNA integrity. Our hypothesis postulates that ASPM recapitulates the functions of most human microcephaly genes and provides a justification for why ASPM is the most frequently mutated gene in autosomal recessive primary microcephaly.
    Keywords:  DNA repair; DNA replication; Drosophila Asp; asters; cell cycle progression; central spindle; human ASPM; microcephaly; mouse Aspm; spindle poles
    DOI:  https://doi.org/10.3390/cells12060922
  5. Cell Tissue Res. 2023 Mar 29.
    Src inhibition induces mitotic arrest associated with chromosomal passenger complex.
    Song Yang, Youguang Luo, Mulin Yang, Hua Ni, Hanxiao Yin, Ming Hu, Min Liu, Jun Zhou, Yunfan Yang, Dengwen Li.
      The non-receptor tyrosine kinase Src plays a key role in cell division, migration, adhesion, and survival. Src is overactivated in several cancers, where it transmits signals that promote cell survival, mitosis, and other important cancer hallmarks. Src is therefore a promising target in cancer therapy, but the underlying mechanisms are still uncertain. Here we show that Src is highly conserved across different species. Src expression increases during mitosis and is localized to the chromosomal passenger complex. Knockdown or inhibition of Src induces multipolar spindle formation, resulting in abnormal expression of the Aurora B and INCENP components of the chromosomal passenger complex. Molecular mechanism studies have found that Src interacts with and phosphorylates INCENP. This then leads to incorrect chromosome arrangement and segregation, resulting in cell division failure. Herein, Src and chromosomal passenger complex co-localize and Src inhibition impedes mitotic progression by inducing multipolar spindle formation. These findings provide novel insights into the molecular basis for using Src inhibitors to treat cancer.
    Keywords:  Chromosomal passenger complex; Mitosis; Multipolar spindle; Src
    DOI:  https://doi.org/10.1007/s00441-023-03765-7
  6. Nat Rev Mol Cell Biol. 2023 Mar 28.
    Mechanisms underlying spindle assembly and robustness.
    Venecia A Valdez, Lila Neahring, Sabine Petry, Sophie Dumont.
      The microtubule-based spindle orchestrates chromosome segregation during cell division. Following more than a century of study, many components and pathways contributing to spindle assembly have been described, but how the spindle robustly assembles remains incompletely understood. This process involves the self-organization of a large number of molecular parts - up to hundreds of thousands in vertebrate cells - whose local interactions give rise to a cellular-scale structure with emergent architecture, mechanics and function. In this Review, we discuss key concepts in our understanding of spindle assembly, focusing on recent advances and the new approaches that enabled them. We describe the pathways that generate the microtubule framework of the spindle by driving microtubule nucleation in a spatially controlled fashion and present recent insights regarding the organization of individual microtubules into structural modules. Finally, we discuss the emergent properties of the spindle that enable robust chromosome segregation.
    DOI:  https://doi.org/10.1038/s41580-023-00584-0
  7. bioRxiv. 2023 Mar 23. pii: 2023.03.13.532437. [Epub ahead of print]
    A Dynamic population of prophase CENP-C is required for meiotic chromosome segregation.
    Jessica E Fellmeth, Hannah Sturm, Janet Jang, Neha Changela, Aashka Parikh, Manisha Persaud, Kim S McKim.
      The centromere is an epigenetic mark that is a loading site for the kinetochore during meiosis and mitosis. This mark is characterized by the H3 variant CENP-A, known as CID in Drosophila , which replaces canonical H3 at the centromeres. In Drosophila , CENP-C is critical for maintaining CID at the centromeres and directly recruits outer kinetochore proteins after nuclear envelope break down. It is not clear, however, if these two functions require the same population of CENP-C. In Drosophila and many other metazoan oocytes, centromere maintenance and kinetochore assembly are separated by an extended prophase. We used RNAi knockdown, mutants, and transgenes to study the dynamics and function of CENP-C in meiosis. CENP-C that is incorporated into cells prior to the onset of meiosis is involved in centromere maintenance and CID recruitment. We found this is not sufficient for the other functions of CENP-C. Indeed, CENP-C is loaded during meiotic prophase, while CID and the chaperone CAL1 are not. CENP-C prophase loading is required for meiotic functions at two different times. In early meiotic prophase, CENP-C loading is required for sister centromere cohesion and centromere clustering. In late meiotic prophase, CENP-C loading is required to recruit kinetochore proteins. Thus, CENP-C is one of the few proteins that links the function of the centromeres and kinetochores through the long prophase pause in oocytes.
    DOI:  https://doi.org/10.1101/2023.03.13.532437
  8. Proc Natl Acad Sci U S A. 2023 Apr 04. 120(14): e2216700120
    HPV16 E6 induces chromosomal instability due to polar chromosomes caused by E6AP-dependent degradation of the mitotic kinesin CENP-E.
    Pippa F Cosper, Laura C F Hrycyniak, Maha Paracha, Denis L Lee, Jun Wan, Kathryn Jones, Sophie A Bice, Kwangok Nickel, Samyukta Mallick, Alison M Taylor, Randall J Kimple, Paul F Lambert, Beth A Weaver.
      Chromosome segregation during mitosis is highly regulated to ensure production of genetically identical progeny. Recurrent mitotic errors cause chromosomal instability (CIN), a hallmark of tumors. The E6 and E7 oncoproteins of high-risk human papillomavirus (HPV), which causes cervical, anal, and head and neck cancers (HNC), cause mitotic defects consistent with CIN in models of anogenital cancers, but this has not been studied in the context of HNC. Here, we show that HPV16 induces a specific type of CIN in patient HNC tumors, patient-derived xenografts, and cell lines, which is due to defects in chromosome congression. These defects are specifically induced by the HPV16 oncogene E6 rather than E7. We show that HPV16 E6 expression causes degradation of the mitotic kinesin CENP-E, whose depletion produces chromosomes that are chronically misaligned near spindle poles (polar chromosomes) and fail to congress. Though the canonical oncogenic role of E6 is the degradation of the tumor suppressor p53, CENP-E degradation and polar chromosomes occur independently of p53. Instead, E6 directs CENP-E degradation in a proteasome-dependent manner via the E6-associated ubiquitin protein ligase E6AP/UBE3A. This study reveals a mechanism by which HPV induces CIN, which may impact HPV-mediated tumor initiation, progression, and therapeutic response.
    Keywords:  CIN; mitosis; papillomavirus
    DOI:  https://doi.org/10.1073/pnas.2216700120
  9. Mol Biol Cell. 2023 Mar 29. mbcE22110518
    Mitotic DNA damage promotes chromokinesin-mediated missegregation of polar chromosomes in cancer cells.
    Marco Novais-Cruz, António Pombinho, Mafalda Sousa, André F Maia, Helder Maiato, Cristina Ferrás.
      DNA damage response (DDR) during interphase involves active signaling and repair to ensure genomic stability. However, how mitotic cells respond to DNA damage remains poorly understood. Supported by correlative live-/fixed-cell microscopy, it was found that mitotic cells exposed to several cancer chemotherapy compounds acquire and signal DNA damage, regardless of how they interact with DNA. In-depth analysis upon DNA damage during mitosis revealed a spindle assembly checkpoint (SAC)-dependent, but ataxia telangiectasia mutated-independent, mitotic delay. This delay was due to the presence of misaligned chromosomes that ultimately satisfy the SAC and missegregate, leading to micronuclei formation. Mechanistically, it is shown that mitotic DNA damage causes missegregation of polar chromosomes due to the action of arm-ejection forces by chromokinesins. Importantly, with the exception of DNA damage induced by etoposide-a topoisomerase II inhibitor-this outcome was independent of a general effect on kinetochore microtubule stability. Colony formation assays in pan-cancer cell line models revealed that mitotic DNA damage causes distinct cytotoxic effects, depending on the nature and extent of the damage. Overall, these findings unveil and raise awareness that therapeutic DNA damage regimens may contribute to genomic instability through a surprising link with chromokinesin-mediated missegregation of polar chromosomes in cancer cells.
    DOI:  https://doi.org/10.1091/mbc.E22-11-0518
  10. bioRxiv. 2023 Mar 16. pii: 2023.03.16.532763. [Epub ahead of print]
    RHINO restricts MMEJ activity to mitosis.
    Alessandra Brambati, Olivia Sacco, Sarina Porcella, Joshua Heyza, Mike Kareh, Jens C Schmidt, Agnel Sfeir.
      DNA double-strand breaks (DSBs) are toxic lesions that can lead to genome instability if not properly repaired. Breaks incurred in G1 phase of the cell cycle are predominantly fixed by non-homologous end-joining (NHEJ), while homologous recombination (HR) is the primary repair pathway in S and G2. Microhomology-mediated end-joining (MMEJ) is intrinsically error-prone and considered a backup DSB repair pathway that becomes essential when HR and NHEJ are compromised. In this study, we uncover MMEJ as the major DSB repair pathway in M phase. Using CRISPR/Cas9-based synthetic lethal screens, we identify subunits of the 9-1-1 complex (RAD9A-HUS1-RAD1) and its interacting partner, RHINO, as critical MMEJ factors. Mechanistically, we show that the function of 9-1-1 and RHINO in MMEJ is inconsistent with their well-established role in ATR signaling. Instead, RHINO plays an unexpected and essential role in directing mutagenic repair to M phase by directly binding to Polymerase theta (Polθ) and promoting its recruitment to DSBs in mitosis. In addition, we provide evidence that mitotic MMEJ repairs persistent DNA damage that originates in S phase but is not repaired by HR. The latter findings could explain the synthetic lethal relationship between POLQ and BRCA1/2 and the synergistic effect of Polθ and PARP inhibitors. In summary, our study identifies MMEJ as the primary pathway for repairing DSBs during mitosis and highlights an unanticipated role for RHINO in directing mutagenic repair to M phase.
    DOI:  https://doi.org/10.1101/2023.03.16.532763
  11. Cancer Res Commun. 2023 Jan;3(1): 54-65
    Misaligned Chromosomes are a Major Source of Chromosomal Instability in Breast Cancer.
    John B Tucker, Sarah C Bonema, Rebeca García-Varela, Ryan A Denu, Yang Hu, Stephanie M McGregor, Mark E Burkard, Beth A Weaver.
      Chromosomal instability (CIN), the persistent reshuffling of chromosomes during mitosis, is a hallmark of human cancers that contributes to tumor heterogeneity and has been implicated in driving metastasis and altering responses to therapy. Though multiple mechanisms can produce CIN, lagging chromosomes generated from abnormal merotelic attachments are the major cause of CIN in a variety of cell lines, and are expected to predominate in cancer. Here, we quantify CIN in breast cancer using a tumor microarray, matched primary and metastatic samples, and patient-derived organoids from primary breast cancer. Surprisingly, misaligned chromosomes are more common than lagging chromosomes and represent a major source of CIN in primary and metastatic tumors. This feature of breast cancers is conserved in a majority of breast cancer cell lines. Importantly, though a portion of misaligned chromosomes align before anaphase onset, the fraction that remain represents the largest source of CIN in these cells. Metastatic breast cancers exhibit higher rates of CIN than matched primary cancers, primarily due to increases in misaligned chromosomes. Whether CIN causes immune activation or evasion is controversial. We find that misaligned chromosomes result in immune-activating micronuclei substantially less frequently than lagging and bridge chromosomes and that breast cancers with greater frequencies of lagging chromosomes and chromosome bridges recruit more stromal tumor-infiltrating lymphocytes. These data indicate misaligned chromosomes represent a major mechanism of CIN in breast cancer and provide support for differential immunostimulatory effects of specific types of CIN.
    Significance: We surveyed the single-cell landscape of mitotic defects that generate CIN in primary and metastatic breast cancer and relevant models. Misaligned chromosomes predominate, and are less immunostimulatory than other chromosome segregation errors.
    DOI:  https://doi.org/10.1158/2767-9764.CRC-22-0302
  12. Cells. 2023 Mar 16. pii: 917. [Epub ahead of print]12(6):
    Actin-Microtubule Crosstalk Imparts Stiffness to the Contractile Ring in Fission Yeast.
    Kimberly Bellingham-Johnstun, Zoe L Tyree, Jessica Martinez-Baird, Annelise Thorn, Caroline Laplante.
      Actin-microtubule interactions are critical for cell division, yet how these networks of polymers mutually influence their mechanical properties and functions in live cells remains unknown. In fission yeast, the post-anaphase array (PAA) of microtubules assembles in the plane of the contractile ring, and its assembly relies on the Myp2p-dependent recruitment of Mto1p, a component of equatorial microtubule organizing centers (eMTOCs). The general organization of this array of microtubules and the impact on their physical attachment to the contractile ring remain unclear. We found that Myp2p facilitates the recruitment of Mto1p to the inner face of the contractile ring, where the eMTOCs polymerize microtubules without their direct interaction. The PAA microtubules form a dynamic polygon of Ase1p crosslinked microtubules inside the contractile ring. The specific loss of PAA microtubules affects the mechanical properties of the contractile ring of actin by lowering its stiffness. This change in the mechanical properties of the ring has no measurable impact on cytokinesis or on the anchoring of the ring. Our work proposes that the PAA microtubules exploit the contractile ring for their assembly and function during cell division, while the contractile ring may receive no benefit from these interactions.
    Keywords:  contractile ring; cytokinesis; mechanical properties; microtubules; mitosis; quantitative microscopy
    DOI:  https://doi.org/10.3390/cells12060917
  13. J Cancer Res Clin Oncol. 2023 Mar 31.
    Mitotic phosphorylation of Tau/MAPT modulates cell cycle progression in prostate cancer cells.
    Letizia Clementi, Samantha Sabetta, Veronica Zelli, Chiara Compagnoni, Alessandra Tessitore, Vincenzo Mattei, Adriano Angelucci.
       PURPOSE: Tau/MAPT (microtubule associated protein tau) protein is actively studied for the pathologic consequences of its aberrant proteostasis in central nervous system leading to neurodegenerative diseases. Besides its ability to generate insoluble toxic oligomers, Tau homeostasis has attracted attention for its involvement in the formation of the mitotic spindle. This evidence, in association with the description of Tau expression in extra-neuronal tissues, and mainly in cancer tissues, constitutes the rationale for a more in-depth investigation of Tau role also in neoplastic diseases.
    METHODS: In our study, we investigated the expression of phosphorylated Tau in prostate cancer cell lines with particular focus on the residue Thr231 present in microtubule binding domain.
    RESULTS: The analysis of prostate cancer cells synchronized with nocodazole demonstrated that the expression of Tau protein phosphorylated at residue Thr231 is restricted to G2/M cell cycle phase. The phosphorylated form was unable to bind tubulin and it does not localize on mitotic spindle. As demonstrated by the use of specific inhibitors, the phosphorylation status of Tau is under the direct control of cdk5 and PP2A, while cdk1 activation was able to exert an indirect control. These mechanisms were also active in cells treated with docetaxel, where counteracting the expression of the dephosphorylated form, by kinase inhibition or protein silencing, determined resistance to drug toxicity.
    CONCLUSIONS: We hypothesize that phosphorylation status of Tau is a key marker for G2/M phase in prostate cancer cells and that the forced modulation of Tau phosphorylation can interfere with the capacity of cell to efficiently progress through G2/M phase.
    Keywords:  Cancer therapy; Cell cycle; Docetaxel; Mitosis; Prostate cancer
    DOI:  https://doi.org/10.1007/s00432-023-04721-2
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