bims-micesi 2023-07-09 papers

bims-micesi

Biomed News

on Mitotic cell signalling

Issue of 2023–07–09
twelve papers selected by
Valentina Piano, Uniklinik Köln

Centrosome linker diversity and its function in centrosome clustering and mitotic spindle formation.
miR-31-mediated local translation at the mitotic spindle is important for early development.
A new layer of regulation of chromosomal passenger complex (CPC) translocation in budding yeast.
Kinetochore-fiber lengths are maintained locally but coordinated globally by poles in the mammalian spindle.
The oncogene cyclin D1 promotes bipolar spindle integrity under compressive force.
SUMOylation at the inner nuclear membrane facilitates nuclear envelope biogenesis during mitosis.
Physcomitrium patens SUN2 mediates MTOC association to the nuclear envelope and facilitates chromosome alignment during spindle assembly.
Myosin 10 uses its MyTH4 and FERM domains differentially to support two aspects of spindle pole biology required for mitotic spindle bipolarity.
Kinetochore component function in C. elegans oocytes revealed by 4D tracking of holocentric chromosomes.
Temporal Changes in Nuclear Envelope Permeability during Semi-Closed Mitosis in Dictyostelium Amoebae.
Length limitation of astral microtubules orients cell divisions in murine intestinal crypts.
Maintaining asymmetry in cell division.

EMBO J. 2023 Jul 04. e109738

Centrosome linker diversity and its function in centrosome clustering and mitotic spindle formation.

Laura Theile, Xue Li, Hairuo Dang, Dorothee Mersch, Simon Anders, Elmar Schiebel.

  The centrosome linker joins the two interphase centrosomes of a cell into one microtubule organizing center. Despite increasing knowledge on linker components, linker diversity in different cell types and their role in cells with supernumerary centrosomes remained unexplored. Here, we identified Ninein as a C-Nap1-anchored centrosome linker component that provides linker function in RPE1 cells while in HCT116 and U2OS cells, Ninein and Rootletin link centrosomes together. In interphase, overamplified centrosomes use the linker for centrosome clustering, where Rootletin gains centrosome linker function in RPE1 cells. Surprisingly, in cells with centrosome overamplification, C-Nap1 loss prolongs metaphase through persistent activation of the spindle assembly checkpoint indicated by BUB1 and MAD1 accumulation at kinetochores. In cells lacking C-Nap1, the reduction of microtubule nucleation at centrosomes and the delay in nuclear envelop rupture in prophase probably cause mitotic defects like multipolar spindle formation and chromosome mis-segregation. These defects are enhanced when the kinesin HSET, which normally clusters multiple centrosomes in mitosis, is partially inhibited indicating a functional interplay between C-Nap1 and centrosome clustering in mitosis.

Keywords:  Ninein; Rootletin; centrosome clustering; centrosome cohesion; centrosome linker

DOI:  https://doi.org/10.15252/embj.2021109738
Res Sq. 2023 Jun 14. pii: rs.3.rs-3044775. [Epub ahead of print]

miR-31-mediated local translation at the mitotic spindle is important for early development.

Carolyn Remsburg, Kalin Konrad, Nadezda Stepicheva, Michael Testa, Kelvin Lee, Leila Choe, Shawn Polson, Jaysheel Bhavsar, Hongzhan Huang, Jia Song.

miR-31 is a highly conserved microRNA that plays critical roles in cell proliferation, migration, and differentiation. We discovered miR-31 and some of its validated targets are enriched on the mitotic spindle of the dividing sea urchin embryo and mammalian cells. Using the sea urchin embryo, we found that miR-31 inhibition led to developmental delay correlated with increased cytoskeleton and chromosomal defects. We identified miR-31 to directly suppress several actin remodeling transcripts, β-actin , Gelsolin , Rab35 and Fascin , which were localized to the mitotic spindle. miR-31 inhibition leads to increased newly translated Fascin at the spindles. Forced ectopic localization of Fascin transcripts to the cell membrane and translation led to significant developmental and chromosomal segregation defects, leading to our hypothesis that miR-31 regulates local translation at the mitotic spindle to ensure proper cell division. Furthermore, miR-31-mediated post-transcriptional regulation at the mitotic spindle may be an evolutionarily conserved regulatory paradigm of mitosis.

DOI: https://doi.org/10.21203/rs.3.rs-3044775/v1
Mol Biol Cell. 2023 Jul 05. mbcE23020063

A new layer of regulation of chromosomal passenger complex (CPC) translocation in budding yeast.

Delaney Sherwin, Emily Gutierrez-Morton, Michael Bokros, Cory Haluska, Yanchang Wang.

The conserved CPC consists of Ipl1Aurora-B, Sli15INCENP, Bir1Survivin, and Nbl1Borealin, and localizes at the kinetochore/centromere to correct kinetochore attachment errors and to prevent checkpoint silencing. After anaphase entry, the CPC moves from the kinetochore/centromere to the spindle. In budding yeast, CPC subunit Sli15 is phosphorylated by both cyclin-dependent kinase (CDK) and Ipl1 kinase. Following anaphase onset, activated Cdc14 phosphatase reverses Sli15 phosphorylation imposed by CDK to promote CPC translocation. Although abolished Sli15 phosphorylation imposed by Ipl1 also causes CPC translocation, the regulation of Ipl1-imposed Sli15 phosphorylation remains unclear. In addition to Sli15, Cdc14 also dephosphorylates Fin1, a regulatory subunit of protein phosphatase 1 (PP1), to enable kinetochore localization of Fin1-PP1. Here we present evidence supporting the notion that kinetochore-localized Fin1-PP1 likely reverses Ipl1-imposed Sli15 phosphorylation to promote CPC translocation from the kinetochore/centromere to the spindle. Importantly, premature Fin1 kinetochore localization or phospho-deficient sli15 mutation causes checkpoint defects in response to tensionless attachments, resulting in chromosome missegregation. In addition, our data indicate that reversion of CDK- and Ipl1-imposed Sli15 phosphorylation shows an additive effect on CPC translocation. Together, these results reveal a previously unidentified pathway to regulate CPC translocation, which is important for accurate chromosome segregation.

DOI: https://doi.org/10.1091/mbc.E23-02-0063
Elife. 2023 Jul 03. pii: e85208. [Epub ahead of print]12

Kinetochore-fiber lengths are maintained locally but coordinated globally by poles in the mammalian spindle.

Manuela Richter, Lila Neahring, Jinghui Tao, Renaldo Sutanto, Nathan H Cho, Sophie Dumont.

  At each cell division, nanometer-scale components self-organize to build a micron-scale spindle. In mammalian spindles, microtubule bundles called kinetochore-fibers attach to chromosomes and focus into spindle poles. Despite evidence suggesting that poles can set spindle length, their role remains poorly understood. In fact, many species do not have spindle poles. Here, we probe the pole's contribution to mammalian spindle length, dynamics, and function by inhibiting dynein to generate spindles whose kinetochore-fibers do not focus into poles, yet maintain a metaphase steady-state length. We find that unfocused kinetochore-fibers have a mean length indistinguishable from control, but a broader length distribution, and reduced length coordination between sisters and neighbors. Further, we show that unfocused kinetochore-fibers, like control, can grow back to their steady-state length if acutely shortened by drug treatment or laser ablation: they recover their length by tuning their end dynamics, albeit slower due to their reduced baseline dynamics. Thus, kinetochore-fiber dynamics are regulated by their length, not just pole-focusing forces. Finally, we show that spindles with unfocused kinetochore-fibers can segregate chromosomes but fail to correctly do so. We propose that mammalian spindle length emerges locally from individual k-fibers while spindle poles globally coordinate k-fibers across space and time.

Keywords:  cell biology; human; physics of living systems

DOI:  https://doi.org/10.7554/eLife.85208
bioRxiv. 2023 May 31. pii: 2023.05.30.542893. [Epub ahead of print]

The oncogene cyclin D1 promotes bipolar spindle integrity under compressive force.

Renaldo Sutanto, Lila Neahring, Andrea Serra Marques, Seda Kilinc, Andrei Goga, Sophie Dumont.

The mitotic spindle is the bipolar, microtubule-based structure that segregates chromosomes at each cell division. Aberrant spindles are frequently observed in cancer cells, but how oncogenic transformation affects spindle mechanics and function, particularly in the mechanical context of solid tumors, remains poorly understood. Here, we constitutively overexpress the oncogene cyclin D1 in human MCF10A cells to probe its effects on spindle architecture and response to compressive force. We find that cyclin D1 overexpression increases the incidence of spindles with extra poles, centrioles, and chromosomes. However, it also protects spindle poles from fracturing under compressive force, a deleterious outcome linked to multipolar cell divisions. Our findings suggest that cyclin D1 overexpression may adapt cells to increased compressive stress, contributing to its prevalence in cancers such as breast cancer by allowing continued proliferation in mechanically challenging environments.

DOI: https://doi.org/10.1101/2023.05.30.542893
J Cell Biol. 2023 08 07. pii: e202208137. [Epub ahead of print]222(8):

SUMOylation at the inner nuclear membrane facilitates nuclear envelope biogenesis during mitosis.

Natasha O Saik, Christopher Ptak, Saif Rehman, John D Aitchison, Ben Montpetit, Richard W Wozniak.

As eukaryotic cells progress through cell division, the nuclear envelope (NE) membrane must expand to accommodate the formation of progeny nuclei. In Saccharomyces cerevisiae, closed mitosis allows visualization of NE biogenesis during mitosis. During this period, the SUMO E3 ligase Siz2 binds the inner nuclear membrane (INM) and initiates a wave of INM protein SUMOylation. Here, we show these events increase INM levels of phosphatidic acid (PA), an intermediate of phospholipid biogenesis, and are necessary for normal mitotic NE membrane expansion. The increase in INM PA is driven by the Siz2-mediated inhibition of the PA phosphatase Pah1. During mitosis, this results from the binding of Siz2 to the INM and dissociation of Spo7 and Nem1, a complex required for the activation of Pah1. As cells enter interphase, the process is then reversed by the deSUMOylase Ulp1. This work further establishes a central role for temporally controlled INM SUMOylation in coordinating processes, including membrane expansion, that regulate NE biogenesis during mitosis.

DOI: https://doi.org/10.1083/jcb.202208137
Plant Cell Physiol. 2023 Jul 08. pii: pcad074. [Epub ahead of print]

Physcomitrium patens SUN2 mediates MTOC association to the nuclear envelope and facilitates chromosome alignment during spindle assembly.

Mari W Yoshida, Noiri Oguri, Gohta Goshima.

  Plant cells lack centrosomes and instead utilise acentrosomal microtubule organising centres (MTOCs) to rapidly increase the number of microtubules at the onset of spindle assembly. Although several proteins required for MTOC formation have been identified, how the MTOC is positioned at the right place is not known. Here, we show that the inner nuclear membrane protein SUN2 is required for MTOC association with the nuclear envelope (NE) during mitotic prophase in the moss Physcomitrium patens. In actively dividing protonemal cells, microtubules accumulate around the NE during prophase. In particular, regional MTOC is formed at the apical surface of the nucleus. However, microtubule accumulation around the NE was impaired and apical MTOCs were mislocalised in sun2 knockout (KO) cells. Upon nuclear envelope breakdown (NEBD), the mitotic spindle was assembled with mislocalised MTOCs. However, completion of chromosome alignment in the spindle was delayed; in severe cases, the chromosome was transiently detached from the spindle body. SUN2 tended to localise to the apical surface of the nucleus during prophase in a microtubule-dependent manner. Based on these results, we propose that SUN2 facilitates the attachment of microtubules to chromosomes during spindle assembly by localising microtubules to the NE. MTOC mispositioning was also observed during the first division of the gametophore tissue. Thus, this study suggests that microtubule-nucleus linking, a well-known function of SUN in animals and yeast, is conserved in plants.

Keywords:  Microtubule organising centre (MTOC)/LINC complex/chromosome congression/nuclear migration/Physcomitrium patens

DOI:  https://doi.org/10.1093/pcp/pcad074
bioRxiv. 2023 Jun 15. pii: 2023.06.15.545002. [Epub ahead of print]

Myosin 10 uses its MyTH4 and FERM domains differentially to support two aspects of spindle pole biology required for mitotic spindle bipolarity.

Yang-In Yim, Antonio Pedrosa, Xufeng Wu, Krishna Chinthalapudi, Richard E Cheney, John A Hammer.

Myosin 10 (Myo10) has the ability to link actin filaments to integrin-based adhesions and to microtubules by virtue of its integrin-binding FERM domain and microtubule-binding MyTH4 domain, respectively. Here we used Myo10 knockout cells to define Myo10's contribution to the maintenance of spindle bipolarity, and complementation to quantitate the relative contributions of its MyTH4 and FERM domains. Myo10 knockout HeLa cells and mouse embryo fibroblasts (MEFs) both exhibit a pronounced increase in the frequency of multipolar spindles. Staining of unsynchronized metaphase cells showed that the primary driver of spindle multipolarity in knockout MEFs and knockout HeLa cells lacking supernumerary centrosomes is pericentriolar material (PCM) fragmentation, which creates y-tubulin-positive acentriolar foci that serve as additional spindle poles. For HeLa cells possessing supernumerary centrosomes, Myo10 depletion further accentuates spindle multipolarity by impairing the clustering of the extra spindle poles. Complementation experiments show that Myo10 must interact with both integrins and microtubules to promote PCM/pole integrity. Conversely, Myo10's ability to promote the clustering of supernumerary centrosomes only requires that it interact with integrins. Importantly, images of Halo-Myo10 knock-in cells show that the myosin localizes exclusively within adhesive retraction fibers during mitosis. Based on these and other results, we conclude that Myo10 promotes PCM/pole integrity at a distance, and that it facilitates supernumerary centrosome clustering by promoting retraction fiber-based cell adhesion, which likely provides an anchor for the microtubule-based forces driving pole focusing.

DOI: https://doi.org/10.1101/2023.06.15.545002
Nat Commun. 2023 Jul 07. 14(1): 4032

Kinetochore component function in C. elegans oocytes revealed by 4D tracking of holocentric chromosomes.

Laras Pitayu-Nugroho, Mélanie Aubry, Kimberley Laband, Hélène Geoffroy, Thadshagine Ganeswaran, Audi Primadhanty, Julie C Canman, Julien Dumont.

During cell division, chromosome congression to the spindle center, their orientation along the spindle long axis and alignment at the metaphase plate depend on interactions between spindle microtubules and kinetochores, and are pre-requisite for chromosome bi-orientation and accurate segregation. How these successive phases are controlled during oocyte meiosis remains elusive. Here we provide 4D live imaging during the first meiotic division in C. elegans oocytes with wild-type or disrupted kinetochore protein function. We show that, unlike in monocentric organisms, holocentric chromosome bi-orientation is not strictly required for accurate chromosome segregation. Instead, we propose a model in which initial kinetochore-localized BHC module (comprised of BUB-1Bub1, HCP-1/2CENP-F and CLS-2CLASP)-dependent pushing acts redundantly with Ndc80 complex-mediated pulling for accurate chromosome segregation in meiosis. In absence of both mechanisms, homologous chromosomes tend to co-segregate in anaphase, especially when initially mis-oriented. Our results highlight how different kinetochore components cooperate to promote accurate holocentric chromosome segregation in oocytes of C. elegans.

DOI: https://doi.org/10.1038/s41467-023-39702-z
Cells. 2023 May 13. pii: 1380. [Epub ahead of print]12(10):

Temporal Changes in Nuclear Envelope Permeability during Semi-Closed Mitosis in Dictyostelium Amoebae.

Kristina Mitic, Irene Meyer, Ralph Gräf, Marianne Grafe.

  The Amoebozoan Dictyostelium discoideum exhibits a semi-closed mitosis in which the nuclear membranes remain intact but become permeabilized to allow tubulin and spindle assembly factors to access the nuclear interior. Previous work indicated that this is accomplished at least by partial disassembly of nuclear pore complexes (NPCs). Further contributions by the insertion process of the duplicating, formerly cytosolic, centrosome into the nuclear envelope and nuclear envelope fenestrations forming around the central spindle during karyokinesis were discussed. We studied the behavior of several Dictyostelium nuclear envelope, centrosomal, and nuclear pore complex (NPC) components tagged with fluorescence markers together with a nuclear permeabilization marker (NLS-TdTomato) by live-cell imaging. We could show that permeabilization of the nuclear envelope during mitosis occurs in synchrony with centrosome insertion into the nuclear envelope and partial disassembly of nuclear pore complexes. Furthermore, centrosome duplication takes place after its insertion into the nuclear envelope and after initiation of permeabilization. Restoration of nuclear envelope integrity usually occurs long after re-assembly of NPCs and cytokinesis has taken place and is accompanied by a concentration of endosomal sorting complex required for transport (ESCRT) components at both sites of nuclear envelope fenestration (centrosome and central spindle).

Keywords:  Dictyostelium; centrosome; mitosis; nuclear envelope; nuclear pore complex

DOI:  https://doi.org/10.3390/cells12101380
Dev Cell. 2023 Jul 04. pii: S1534-5807(23)00302-7. [Epub ahead of print]

Length limitation of astral microtubules orients cell divisions in murine intestinal crypts.

Jad Saleh, Marc-Antoine Fardin, Amlan Barai, Matis Soleilhac, Olivia Frenoy, Cécile Gaston, Hongyue Cui, Tien Dang, Noémie Gaudin, Audrey Vincent, Nicolas Minc, Delphine Delacour.

  Planar spindle orientation is critical for epithelial tissue organization and is generally instructed by the long cell-shape axis or cortical polarity domains. We introduced mouse intestinal organoids in order to study spindle orientation in a monolayered mammalian epithelium. Although spindles were planar, mitotic cells remained elongated along the apico-basal (A-B) axis, and polarity complexes were segregated to basal poles, so that spindles oriented in an unconventional manner, orthogonal to both polarity and geometric cues. Using high-resolution 3D imaging, simulations, and cell-shape and cytoskeleton manipulations, we show that planar divisions resulted from a length limitation in astral microtubules (MTs) which precludes them from interacting with basal polarity, and orient spindles from the local geometry of apical domains. Accordingly, lengthening MTs affected spindle planarity, cell positioning, and crypt arrangement. We conclude that MT length regulation may serve as a key mechanism for spindles to sense local cell shapes and tissue forces to preserve mammalian epithelial architecture.

Keywords:  actomyosin; astral microtubules; cell division; organoids; polarity; spindle positioning

DOI:  https://doi.org/10.1016/j.devcel.2023.06.004
Science. 2023 Jul 07. 381(6653): 27-28

Maintaining asymmetry in cell division.

Camila Goldy, Marie-Cécile Caillaud.

Promoting asymmetric division through microtubule dynamics establishes cell fate.

DOI: https://doi.org/10.1126/science.adi6664