bims-micesi 2024-08-18 papers

bims-micesi

Biomed News

on Mitotic cell signalling

Issue of 2024–08–18
eight papers selected by
Valentina Piano, Uniklinik Köln

B-type plexins regulate mitosis via RanGTPase.
Spatial control of the APC/C ensures the rapid degradation of cyclin B1.
SKAP binding to microtubules reduces friction at the kinetochore-microtubule interface and increases attachment stability under force.
Balancing Plk1 activity levels: The secret of synchrony between the cell and the centrosome cycle.
Drosophila ring chromosomes interact with sisters and homologs to produce anaphase bridges in mitosis.
Architecture of native kinetochores revealed by structural studies utilizing a thermophilic yeast.
β-TrCP-Mediated Proteolysis of Mis18β Prevents Mislocalization of CENP-A and Chromosomal Instability.
HIV-1 Vif disrupts phosphatase feedback regulation at the kinetochore, leading to a pronounced pseudo-metaphase arrest.

Mol Cancer Res. 2024 Aug 13.

B-type plexins regulate mitosis via RanGTPase.

Nicholus Mukhwana, Ritu Garg, Alexandria R Mitchell, Abul Azad, Magali Williamson.

Aberrant mitosis can result in aneuploidy and cancer. The small GTPase, Ran, is a key regulator of mitosis. B-type Plexins regulate Ran activity by acting as RanGTPase activating proteins (GAPs) and have been implicated in cancer progression. However, whether B-type plexins have a role in mitosis has not so far been investigated. We show here that PlexinB1 functions in the control of mitosis. Depletion of PlexinB1 affects mitotic spindle assembly, significantly delaying anaphase. This leads to mitotic catastrophe in some cells, and prolonged application of the spindle assembly checkpoint. PlexinB1 depletion also promoted acentrosomal microtubule nucleation and defects in spindle pole refocussing and increased the number of cells with multipolar or aberrant mitotic spindles. An increase in lagging chromosomes or chromosomal bridges at anaphase was also found upon PlexinB1 depletion. PlexinB1 localises to the mitotic spindle in dividing cells. The mitotic defects observed upon PlexinB1 depletion were rescued by an RCC1 inhibitor, indicating that PlexinB1 signals, via Ran, to affect mitosis. These errors in mitosis generated multinucleate cells, and nuclei of altered morphology and abnormal karyotype. Furthermore, Semaphorin4D-treatment increased the percentage of cells with micronuclei, precursors of chromothripsis. Implications: Defects in B-type plexins may contribute to the well-established role of plexins in cancer progression by inducing chromosomal instability.

DOI: https://doi.org/10.1158/1541-7786.MCR-23-0836
EMBO J. 2024 Aug 14.

Spatial control of the APC/C ensures the rapid degradation of cyclin B1.

Luca Cirillo, Rose Young, Sapthaswaran Veerapathiran, Annalisa Roberti, Molly Martin, Azzah Abubacar, Camilla Perosa, Catherine Coates, Reyhan Muhammad, Theodoros I Roumeliotis, Jyoti S Choudhary, Claudio Alfieri, Jonathon Pines.

  The proper control of mitosis depends on the ubiquitin-mediated degradation of the right mitotic regulator at the right time. This is effected by the Anaphase Promoting Complex/Cyclosome (APC/C) ubiquitin ligase that is regulated by the Spindle Assembly Checkpoint (SAC). The SAC prevents the APC/C from recognising Cyclin B1, the essential anaphase and cytokinesis inhibitor, until all chromosomes are attached to the spindle. Once chromosomes are attached, Cyclin B1 is rapidly degraded to enable chromosome segregation and cytokinesis. We have a good understanding of how the SAC inhibits the APC/C, but relatively little is known about how the APC/C recognises Cyclin B1 as soon as the SAC is turned off. Here, by combining live-cell imaging, in vitro reconstitution biochemistry, and structural analysis by cryo-electron microscopy, we provide evidence that the rapid recognition of Cyclin B1 in metaphase requires spatial regulation of the APC/C. Using fluorescence cross-correlation spectroscopy, we find that Cyclin B1 and the APC/C primarily interact at the mitotic apparatus. We show that this is because Cyclin B1, like the APC/C, binds to nucleosomes, and identify an 'arginine-anchor' in the N-terminus as necessary and sufficient for binding to the nucleosome. Mutating the arginine anchor on Cyclin B1 reduces its interaction with the APC/C and delays its degradation: cells with the mutant, non-nucleosome-binding Cyclin B1 become aneuploid, demonstrating the physiological relevance of our findings. Together, our data demonstrate that mitotic chromosomes promote the efficient interaction between Cyclin B1 and the APC/C to ensure the timely degradation of Cyclin B1 and genomic stability.

Keywords:  Cell Cycle; Chromosome; Mitosis; Nucleosome; Ubiquitin

DOI:  https://doi.org/10.1038/s44318-024-00194-2
bioRxiv. 2024 Aug 08. pii: 2024.08.08.607154. [Epub ahead of print]

SKAP binding to microtubules reduces friction at the kinetochore-microtubule interface and increases attachment stability under force.

Miquel Rosas-Salvans, Caleb Rux, Moumita Das, Sophie Dumont.

The kinetochore links chromosomes to spindle microtubules to drive chromosome segregation at cell division. We recently uncovered that the kinetochore complex Astrin-SKAP, which binds microtubules, reduces rather than increases friction at the mammalian kinetochore-microtubule interface. How it does so is not known. Astrin-SKAP could affect how other kinetochore complexes bind microtubules, reducing their friction along microtubules, or it could itself bind microtubules with similar affinity but lower friction than other attachment factors. Using SKAP mutants unable to bind microtubules, live imaging and laser ablation, we show that SKAP's microtubule binding is essential for sister kinetochore coordination, force dissipation at the interface and attachment responsiveness to force changes. Further, we show that SKAP's microtubule binding is essential to prevent chromosome detachment under both spindle forces and microneedle-generated forces. Together, our findings indicate that SKAP's microtubule binding reduces kinetochore friction and increases attachment responsiveness and stability under force. We propose that having complexes with both high and low sliding friction on microtubules, making a mechanically heterogeneous interface, is key to maintaining robust attachments under force and thus accurate segregation.

DOI: https://doi.org/10.1101/2024.08.08.607154
Bioessays. 2024 Aug 11. e2400048

Balancing Plk1 activity levels: The secret of synchrony between the cell and the centrosome cycle.

Devashish Dwivedi, Patrick Meraldi.

  The accuracy of cell division requires precise regulation of the cellular machinery governing DNA/genome duplication, ensuring its equal distribution among the daughter cells. The control of the centrosome cycle is crucial for the formation of a bipolar spindle, ensuring error-free segregation of the genome. The cell and centrosome cycles operate in close synchrony along similar principles. Both require a single duplication round in every cell cycle, and both are controlled by the activity of key protein kinases. Nevertheless, our comprehension of the precise cellular mechanisms and critical regulators synchronizing these two cycles remains poorly defined. Here, we present our hypothesis that the spatiotemporal regulation of a dynamic equilibrium of mitotic kinases activities forms a molecular clock that governs the synchronous progression of both the cell and the centrosome cycles.

Keywords:  Plk1; centriole; centrosme cycle; centrosomes; genomic stability; kinases; mitosis

DOI:  https://doi.org/10.1002/bies.202400048
bioRxiv. 2024 Aug 09. pii: 2024.08.08.607186. [Epub ahead of print]

Drosophila ring chromosomes interact with sisters and homologs to produce anaphase bridges in mitosis.

Ho-Chen Lin, Mary M Golic, Hunter J Hill, Katherine F Lemons, Truc T Vuong, Madison Smith, Forrest Golic, Kent G Golic.

Ring chromosomes are known in many eukaryotic organisms, including humans. They are typically associated with a variety of maladies, including abnormal development and lethality. Underlying these phenotypes are anaphase chromatin bridges that can lead to chromosome loss, nondisjunction and breakage. By cytological examination of ring chromosomes in Drosophila melanogaster we identified five causes for anaphase bridges produced by ring chromosomes. Catenation of sister chromatids is the most common cause and these bridges frequently resolve during anaphase, presumably by the action of topoisomerase II. Sister chromatid exchange and chromosome breakage followed by sister chromatid union also produce anaphase bridges. Mitotic recombination with the homolog was rare, but was another route to generation of anaphase bridges. Most surprising, was the discovery of homolog capture, where the ring chromosome was connected to its linear homolog in anaphase. We hypothesize that this is a remnant of mitotic pairing and that the linear chromosome is connected to the ring by multiple wraps produced through the action of topoisomerase II during establishment of homolog pairing. In support, we showed that in a ring/ring homozygote the two rings are frequently catenated in mitotic metaphase, a configuration that requires breaking and rejoining of at least one chromosome.

DOI: https://doi.org/10.1101/2024.08.08.607186
Curr Biol. 2024 Aug 07. pii: S0960-9822(24)00939-4. [Epub ahead of print]

Architecture of native kinetochores revealed by structural studies utilizing a thermophilic yeast.

Daniel J Barrero, Sithara S Wijeratne, Xiaowei Zhao, Grace F Cunningham, Rui Yan, Christian R Nelson, Yasuhiro Arimura, Hironori Funabiki, Charles L Asbury, Zhiheng Yu, Radhika Subramanian, Sue Biggins.

  Eukaryotic chromosome segregation requires kinetochores, multi-megadalton protein machines that assemble on the centromeres of chromosomes and mediate attachments to dynamic spindle microtubules. Kinetochores are built from numerous complexes, and there has been progress in structural studies on recombinant subassemblies. However, there is limited structural information on native kinetochore architecture. To address this, we purified functional, native kinetochores from the thermophilic yeast Kluyveromyces marxianus and examined them by electron microscopy (EM), cryoelectron tomography (cryo-ET), and atomic force microscopy (AFM). The kinetochores are extremely large, flexible assemblies that exhibit features consistent with prior models. We assigned kinetochore polarity by visualizing their interactions with microtubules and locating the microtubule binder, Ndc80c. This work shows that isolated kinetochores are more dynamic and complex than what might be anticipated based on the known structures of recombinant subassemblies and provides the foundation to study the global architecture and functions of kinetochores at a structural level.

Keywords:  K. marxianus; atomic force microscopy; centromere; cryoelectron tomography; electron microscopy; kinetochore; mitosis

DOI:  https://doi.org/10.1016/j.cub.2024.07.036
Mol Cell Biol. 2024 Aug 13. 1-14

β-TrCP-Mediated Proteolysis of Mis18β Prevents Mislocalization of CENP-A and Chromosomal Instability.

Subhash Chandra Sethi, Roshan Lal Shrestha, Vinutha Balachandra, Geetha Durairaj, Wei-Chun Au, Michael Nirula, Tatiana S Karpova, Peter Kaiser, Munira A Basrai.

  Restricting the localization of evolutionarily conserved histone H3 variant CENP-A to the centromere is essential to prevent chromosomal instability (CIN), an important hallmark of cancers. Overexpressed CENP-A mislocalizes to non-centromeric regions and contributes to CIN in yeast, flies, and human cells. Centromeric localization of CENP-A is facilitated by the interaction of Mis18β with CENP-A specific chaperone HJURP. Cellular levels of Mis18β are regulated by β-transducin repeat containing protein (β-TrCP), an F-box protein of SCF (Skp1, Cullin, F-box) E3-ubiquitin ligase complex. Here, we show that defects in β-TrCP-mediated proteolysis of Mis18β contributes to the mislocalization of endogenous CENP-A and CIN in a triple-negative breast cancer (TNBC) cell line, MDA-MB-231. CENP-A mislocalization in β-TrCP depleted cells is dependent on high levels of Mis18β as depletion of Mis18β suppresses mislocalization of CENP-A in these cells. Consistent with these results, endogenous CENP-A is mislocalized in cells overexpressing Mis18β alone. In summary, our results show that β-TrCP-mediated degradation of Mis18β prevents mislocalization of CENP-A and CIN. We propose that deregulated expression of Mis18β may be one of the key mechanisms that contributes to chromosome segregation defects in cancers.

Keywords:  CENP-A; E3 ubiquitin ligase; Mis18β; centromere; chromosomal instability; β-TrCP

DOI:  https://doi.org/10.1080/10985549.2024.2382445
bioRxiv. 2024 Jul 30. pii: 2024.07.30.605839. [Epub ahead of print]

HIV-1 Vif disrupts phosphatase feedback regulation at the kinetochore, leading to a pronounced pseudo-metaphase arrest.

Dhaval A Ghone, Edward L Evans, Madison Bandini, Kaelyn G Stephenson, Nathan M Sherer, Aussie Suzuki.

The human immunodeficiency virus type 1 (HIV-1) Virion Infectivity Factor (Vif) targets and degrades cellular APOBEC3 proteins, key regulators of intrinsic and innate antiretroviral immune responses, thereby facilitating HIV-1 infection. While Vif's role in degrading APOBEC3G is well-studied, Vif is also known to cause cell cycle arrest but the detailed nature of Vif's effects on the cell cycle has yet to be delineated. In this study, we employed high-temporal single-cell live imaging and super-resolution microscopy to monitor individual cells during Vif-induced cell cycle arrest. Our findings reveal that Vif does not affect the G2/M boundary as previously thought. Instead, Vif triggers a unique and robust pseudo-metaphase arrest, which is markedly distinct from the mild prometaphase arrest induced by the HIV-1 accessory protein, Vpr, known for modulating the cell cycle. During Vif-mediated arrest, chromosomes align properly to form a metaphase plate but later disassemble, resulting in polar chromosomes. Notably, unlike Vpr, Vif significantly reduces the levels of both Phosphatase 1 (PP1) and 2 (PP2) at kinetochores, which are key regulators of chromosome-microtubule interactions. These results reveal a novel function of Vif in kinetochore regulation that governs the spatial organization of chromosomes during mitosis.

DOI: https://doi.org/10.1101/2024.07.30.605839