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Molecular Cell May 2023Cell cycle and metabolism are intimately intertwined, but how metabolites directly regulate cell-cycle machinery remains elusive. Liu et al. reveal that glycolysis...
Cell cycle and metabolism are intimately intertwined, but how metabolites directly regulate cell-cycle machinery remains elusive. Liu et al. reveal that glycolysis end-product lactate directly binds and inhibits the SUMO protease SENP1 to govern the E3 ligase activity of the anaphase-promoting complex, leading to efficient mitotic exit in proliferative cells.
Topics: Anaphase; Lactic Acid; Mitosis; Anaphase-Promoting Complex-Cyclosome; Cell Cycle Proteins
PubMed: 37207623
DOI: 10.1016/j.molcel.2023.04.013 -
Seminars in Cell & Developmental Biology Sep 2021Mitotic cell divisions ensure stable transmission of genetic information from a mother to daughter cells in a series of generations. To ensure this crucial task is... (Review)
Review
Mitotic cell divisions ensure stable transmission of genetic information from a mother to daughter cells in a series of generations. To ensure this crucial task is accomplished, the cell forms a bipolar structure called the mitotic spindle that divides sister chromatids to the opposite sides of the dividing mother cell. After successful establishment of stable attachments of microtubules to chromosomes and inspection of connections between them, at the heart of mitosis, the cell starts the process of segregation. This spectacular moment in the life of a cell is termed anaphase, and it involves two distinct processes: depolymerization of microtubules bound to chromosomes, which is also known as anaphase A, and elongation of the spindle or anaphase B. Both processes ensure physical separation of disjointed sister chromatids. In this chapter, we review the mechanisms of anaphase B spindle elongation primarily in mammalian systems, combining different pioneering ideas and concepts with more recent findings that shed new light on the force generation and regulation of biochemical modules operating during spindle elongation. Finally, we present a comprehensive model of spindle elongation that includes structural, biophysical, and molecular aspects of anaphase B.
Topics: Anaphase; Chromosome Segregation; Humans; Microtubules
PubMed: 33849764
DOI: 10.1016/j.semcdb.2021.03.023 -
PLoS Biology Jul 2022Precise spatiotemporal control of microtubule nucleation and organization is critical for faithful segregation of cytoplasmic and genetic material during cell division...
Precise spatiotemporal control of microtubule nucleation and organization is critical for faithful segregation of cytoplasmic and genetic material during cell division and signaling via the primary cilium in quiescent cells. Microtubule-associated proteins (MAPs) govern assembly, maintenance, and remodeling of diverse microtubule arrays. While a set of conserved MAPs are only active during cell division, an emerging group of MAPs acts as dual regulators in dividing and nondividing cells. Here, we elucidated the nonciliary functions and molecular mechanism of action of the ciliopathy-linked protein CCDC66, which we previously characterized as a regulator of ciliogenesis in quiescent cells. We showed that CCDC66 dynamically localizes to the centrosomes, the bipolar spindle, the spindle midzone, the central spindle, and the midbody in dividing cells and interacts with the core machinery of centrosome maturation and MAPs involved in cell division. Loss-of-function experiments revealed its functions during mitotic progression and cytokinesis. Specifically, CCDC66 depletion resulted in defective spindle assembly and orientation, kinetochore fiber stability, chromosome alignment in metaphase as well as central spindle and midbody assembly and organization in anaphase and cytokinesis. Notably, CCDC66 regulates mitotic microtubule nucleation via noncentrosomal and centrosomal pathways via recruitment of gamma-tubulin to the centrosomes and the spindle. Additionally, CCDC66 bundles microtubules in vitro and in cells by its C-terminal microtubule-binding domain. Phenotypic rescue experiments showed that the microtubule and centrosome-associated pools of CCDC66 individually or cooperatively mediate its mitotic and cytokinetic functions. Collectively, our findings identify CCDC66 as a multifaceted regulator of the nucleation and organization of the diverse mitotic and cytokinetic microtubule arrays and provide new insight into nonciliary defects that underlie ciliopathies.
Topics: Anaphase; Centrosome; Ciliopathies; Cytokinesis; Eye Proteins; Humans; Microtubule-Associated Proteins; Microtubules; Mitosis; Spindle Apparatus
PubMed: 35849559
DOI: 10.1371/journal.pbio.3001708 -
Cellular and Molecular Life Sciences :... Jul 2010The mitotic spindle uses dynamic microtubules and mitotic motors to generate the pico-Newton scale forces that are needed to drive the mitotic movements that underlie... (Review)
Review
The mitotic spindle uses dynamic microtubules and mitotic motors to generate the pico-Newton scale forces that are needed to drive the mitotic movements that underlie chromosome capture, alignment and segregation. Here, we consider the biophysical and molecular basis of force-generation for chromosome movements in the spindle, and, with reference to the Drosophila embryo mitotic spindle, we briefly discuss how mathematical modeling can complement experimental analysis to illuminate the mechanisms of chromosome-to-pole motility during anaphase A and spindle elongation during anaphase B.
Topics: Anaphase; Animals; Chromosome Segregation; Drosophila; Microtubules; Mitosis; Models, Theoretical; Molecular Motor Proteins; Spindle Apparatus
PubMed: 20221784
DOI: 10.1007/s00018-010-0326-6 -
ELife Apr 2023The trapping of Poly-ADP-ribose polymerase (PARP) on DNA caused by PARP inhibitors (PARPi) triggers acute DNA replication stress and synthetic lethality (SL) in...
The trapping of Poly-ADP-ribose polymerase (PARP) on DNA caused by PARP inhibitors (PARPi) triggers acute DNA replication stress and synthetic lethality (SL) in BRCA2-deficient cells. Hence, DNA damage is accepted as a prerequisite for SL in BRCA2-deficient cells. In contrast, here we show that inhibiting ROCK in BRCA2-deficient cells triggers SL independently from acute replication stress. Such SL is preceded by polyploidy and binucleation resulting from cytokinesis failure. Such initial mitosis abnormalities are followed by other M phase defects, including anaphase bridges and abnormal mitotic figures associated with multipolar spindles, supernumerary centrosomes and multinucleation. SL was also triggered by inhibiting Citron Rho-interacting kinase, another enzyme that, similarly to ROCK, regulates cytokinesis. Together, these observations demonstrate that cytokinesis failure triggers mitotic abnormalities and SL in BRCA2-deficient cells. Furthermore, the prevention of mitotic entry by depletion of Early mitotic inhibitor 1 (EMI1) augmented the survival of BRCA2-deficient cells treated with ROCK inhibitors, thus reinforcing the association between M phase and cell death in BRCA2-deficient cells. This novel SL differs from the one triggered by PARPi and uncovers mitosis as an Achilles heel of BRCA2-deficient cells.
Topics: Anaphase; DNA Damage; Mitosis; Poly(ADP-ribose) Polymerase Inhibitors; Poly(ADP-ribose) Polymerases; Synthetic Lethal Mutations; rho-Associated Kinases; BRCA2 Protein; Humans
PubMed: 37073955
DOI: 10.7554/eLife.80254 -
The Journal of Cell Biology Jun 2022Errors in mitosis that cause chromosome missegregation lead to aneuploidy and micronucleus formation, which are associated with cancer. Accurate segregation requires the...
Errors in mitosis that cause chromosome missegregation lead to aneuploidy and micronucleus formation, which are associated with cancer. Accurate segregation requires the alignment of all chromosomes by the mitotic spindle at the metaphase plate, and any misalignment must be corrected before anaphase is triggered. The spindle is situated in a membrane-free "exclusion zone"; beyond this zone, endomembranes (mainly endoplasmic reticulum) are densely packed. We investigated what happens to misaligned chromosomes localized beyond the exclusion zone. Here we show that such chromosomes become ensheathed in multiple layers of endomembranes. Chromosome ensheathing delays mitosis and increases the frequency of chromosome missegregation and micronucleus formation. We use an induced organelle relocalization strategy in live cells to show that clearance of endomembranes allows for the rescue of chromosomes that were destined for missegregation. Our findings indicate that endomembranes promote the missegregation of misaligned chromosomes that are outside the exclusion zone and therefore constitute a risk factor for aneuploidy.
Topics: Anaphase; Aneuploidy; Cell Membrane; Chromosome Segregation; Chromosomes; Endoplasmic Reticulum; Humans; Metaphase; Mitosis; Spindle Apparatus
PubMed: 35486148
DOI: 10.1083/jcb.202203021 -
ELife Jun 2022The chromokinesin KIF22 generates forces that contribute to mitotic chromosome congression and alignment. Mutations in the α2 helix of the motor domain of KIF22 have...
The chromokinesin KIF22 generates forces that contribute to mitotic chromosome congression and alignment. Mutations in the α2 helix of the motor domain of KIF22 have been identified in patients with abnormal skeletal development, and we report the identification of a patient with a novel mutation in the KIF22 tail. We demonstrate that pathogenic mutations do not result in a loss of KIF22's functions in early mitosis. Instead, mutations disrupt chromosome segregation in anaphase, resulting in reduced proliferation, abnormal daughter cell nuclear morphology, and, in a subset of cells, cytokinesis failure. This phenotype could be explained by a failure of KIF22 to inactivate in anaphase. Consistent with this model, constitutive activation of the motor via a known site of phosphoregulation in the tail phenocopied the effects of pathogenic mutations. These results suggest that the motor domain α2 helix may be an important site for regulation of KIF22 activity at the metaphase to anaphase transition. In support of this conclusion, mimicking phosphorylation of α2 helix residue T158 also prevents inactivation of KIF22 in anaphase. These findings demonstrate the importance of both the head and tail of the motor in regulating the activity of KIF22 and offer insight into the cellular consequences of preventing KIF22 inactivation and disrupting force balance in anaphase.
Topics: Anaphase; Chromosome Segregation; DNA-Binding Proteins; Kinesins; Metaphase; Mitosis; Mutation; Nuclear Proteins; Spindle Apparatus
PubMed: 35730929
DOI: 10.7554/eLife.78653 -
Methods in Molecular Biology (Clifton,... 2021A balanced progression through mitosis and cell division is largely dependent on orderly phosphorylation and ubiquitin-mediated proteolysis of regulatory and structural...
A balanced progression through mitosis and cell division is largely dependent on orderly phosphorylation and ubiquitin-mediated proteolysis of regulatory and structural proteins. These series of events ultimately secure genome stability and time-invariant cellular properties during cell proliferation. Two of the core enzymes regulating mitotic milestones in all eukaryotes are cyclin dependent kinase 1 (CDK1) with its coactivator cyclin B, and the E3 ubiquitin ligase anaphase promoting complex/cyclosome (APC/C). Discovering mechanisms and substrates for these enzymes is vital to understanding how cells move through mitosis and segregate chromosomes with high fidelity. However, the study of these enzymes has significant challenges. Purely in vitro studies discount the contributions of yet to be described regulators and misses the physiological context of cellular environment. In vivo studies are complicated by the fact that each of these enzymes, as well as many of their regulators and downstream targets, are essential. Moreover, long-term in vivo manipulations can result in cascading, indirect effects that can distort data analysis and interpretation. Many of these challenges can be circumvented using cell-free systems, which have historically played a critical role in identifying these enzymes and their contributions under quasicellular environments. Here, we describe the preparation of a newly developed human cell-free system that recapitulates an anaphase-like state of human cells. This new toolkit complements traditional cell-free systems from human cells and frog eggs and can be easily implemented in cell biology labs for direct and quantitative studies of mitotic signaling regulated by phosphorylation, APC/C-mediated proteolysis, and beyond.
Topics: Anaphase; Anaphase-Promoting Complex-Cyclosome; CDC2 Protein Kinase; Cell-Free System; Cyclin B1; HEK293 Cells; Humans; Mitosis; Mutation; Phosphorylation; Proteolysis; Ubiquitination
PubMed: 34085221
DOI: 10.1007/978-1-0716-1538-6_11 -
Cell Cycle (Georgetown, Tex.) Oct 2021To ensure genome stability, chromosomes need to undergo proper condensation into two linked sister chromatids from prophase to prometaphase, followed by equal...
To ensure genome stability, chromosomes need to undergo proper condensation into two linked sister chromatids from prophase to prometaphase, followed by equal segregation at anaphase. Emerging evidence has shown that persistent DNA entanglements connecting the sister chromatids lead to the formation of ultrafine anaphase bridges (UFBs). If UFBs are not resolved soon after anaphase, they can induce chromosome missegregation. PICH (PLK1-interacting checkpoint helicase) is a DNA translocase that localizes on chromosome arms, centromeres and UFBs. It plays multiple essential roles in mitotic chromosome organization and segregation. PICH also recruits other associated proteins to UFBs, and together they mediate UFB resolution. Here, the proposed mechanism behind PICH's functions in chromosome organization and UFB resolution will be discussed. We summarize the regulation of PICH action at chromosome arms and centromeres, how PICH recognizes UFBs and recruits other UFB-associated factors, and finally how PICH promotes UFB resolution together with other DNA processing enzymes.
Topics: Anaphase; Centromere; Chromatids; Chromosome Segregation; Genomic Instability; Humans; Mitosis
PubMed: 34530686
DOI: 10.1080/15384101.2021.1970877 -
Trends in Genetics : TIG Jul 2012Sister chromatids are held together by a protein complex named cohesin. Shugoshin proteins protect cohesin from cleavage by separase during meiosis I in eukaryotes and... (Review)
Review
Sister chromatids are held together by a protein complex named cohesin. Shugoshin proteins protect cohesin from cleavage by separase during meiosis I in eukaryotes and from phosphorylation-mediated removal during mitosis in vertebrates. This protection is crucial for chromosome segregation during mitosis and meiosis. Mechanistically, shugoshins shield cohesin by forming a complex with the phosphatase PP2A, which dephosphorylates cohesin, leading to its retention at centromeres during the onset of meiotic anaphase and vertebrate mitotic prophase I. In addition to this canonical function, shugoshins have evolved novel, species-specific cellular functions, the mechanisms of which remain a subject of intense debate, but are likely to involve spatio-temporally coordinated interactions with the chromosome passenger complex, the spindle checkpoint and the anaphase promoting complex. Here, we compare and contrast these remarkable features of shugoshins in model organisms and humans.
Topics: Anaphase; Animals; Cell Cycle Proteins; Centromere; Chromatids; Chromosomal Proteins, Non-Histone; Humans; Mitosis; Phosphorylation; Cohesins
PubMed: 22542109
DOI: 10.1016/j.tig.2012.03.003