bims-micesi 2024-06-02 papers

bims-micesi

Biomed News

on Mitotic cell signalling

Issue of 2024–06–02
ten papers selected by
Valentina Piano, Uniklinik Köln

The Aurora B-controlled PP1/RepoMan complex determines the spatial and temporal distribution of mitotic H2B S6 phosphorylation.
Chronic spindle assembly checkpoint activation causes myelosuppression and gastrointestinal atrophy.
A noncanonical GTPase signaling mechanism controls exit from mitosis in budding yeast.
Acentric chromosome congression and alignment on the metaphase plate via kinetochore-independent forces in Drosophila.
Genome folding principles uncovered in condensin-depleted mitotic chromosomes.
Temporal and anteriorly positioned mitotic zones drive asymmetric microtubule patterns needed for Left-Right Organizer development.
PLK1 inhibition leads to mitotic arrest and triggers apoptosis in cholangiocarcinoma cells.
Mouse KL2 is a unique MTSE involved in chromosome-based spindle organization and regulated by multiple kinases during female meiosis.
An alternative cell cycle coordinates multiciliated cell differentiation.
A theoretical model of anaphase.

Open Biol. 2024 May;14(5): 230460

The Aurora B-controlled PP1/RepoMan complex determines the spatial and temporal distribution of mitotic H2B S6 phosphorylation.

Maximilian Pfisterer, Roman Robert, Vera V Saul, Amelie Pritz, Markus Seibert, Regina Feederle, M Lienhard Schmitz.

  The precise spatial and temporal control of histone phosphorylations is important for the ordered progression through the different phases of mitosis. The phosphorylation of H2B at S6 (H2B S6ph), which is crucial for chromosome segregation, reaches its maximum level during metaphase and is limited to the inner centromere. We discovered that the temporal and spatial regulation of this modification, as well as its intensity, are governed by the scaffold protein RepoMan and its associated catalytically active phosphatases, PP1α and PP1γ. Phosphatase activity is inhibited at the area of maximal H2B S6 phosphorylation at the inner centromere by site-specific Aurora B-mediated inactivation of the PP1/RepoMan complex. The motor protein Mklp2 contributes to the relocalization of Aurora B from chromatin to the mitotic spindle during anaphase, thus alleviating Aurora B-dependent repression of the PP1/RepoMan complex and enabling dephosphorylation of H2B S6. Accordingly, dysregulation of Mklp2 levels, as commonly observed in tumour cells, leads to the lack of H2B S6 dephosphorylation during early anaphase, which might contribute to chromosomal instability.

Keywords:  centromere; histone phosphorylation; mitosis; phosphatase scaffold

DOI:  https://doi.org/10.1098/rsob.230460
EMBO Rep. 2024 May 28.

Chronic spindle assembly checkpoint activation causes myelosuppression and gastrointestinal atrophy.

Gerlinde Karbon, Fabian Schuler, Vincent Z Braun, Felix Eichin, Manuel Haschka, Mathias Drach, Rocio Sotillo, Stephan Geley, Diana Cj Spierings, Andrea E Tijhuis, Floris Foijer, Andreas Villunger.

  Interference with microtubule dynamics in mitosis activates the spindle assembly checkpoint (SAC) to prevent chromosome segregation errors. The SAC induces mitotic arrest by inhibiting the anaphase-promoting complex (APC) via the mitotic checkpoint complex (MCC). The MCC component MAD2 neutralizes the critical APC cofactor, CDC20, preventing exit from mitosis. Extended mitotic arrest can promote mitochondrial apoptosis and caspase activation. However, the impact of mitotic cell death on tissue homeostasis in vivo is ill-defined. By conditional MAD2 overexpression, we observe that chronic SAC activation triggers bone marrow aplasia and intestinal atrophy in mice. While myelosuppression can be compensated for, gastrointestinal atrophy is detrimental. Remarkably, deletion of pro-apoptotic Bim/Bcl2l11 prevents gastrointestinal syndrome, while neither loss of Noxa/Pmaip or co-deletion of Bid and Puma/Bbc3 has such a protective effect, identifying BIM as rate-limiting apoptosis effector in mitotic cell death of the gastrointestinal epithelium. In contrast, only overexpression of anti-apoptotic BCL2, but none of the BH3-only protein deficiencies mentioned above, can mitigate myelosuppression. Our findings highlight tissue and cell-type-specific survival dependencies in response to SAC perturbation in vivo.

Keywords:  Apoptosis; BH3-only Proteins; MAD2; Mitosis; Spindle Assembly Checkpoint

DOI:  https://doi.org/10.1038/s44319-024-00160-3
bioRxiv. 2024 May 17. pii: 2024.05.16.594582. [Epub ahead of print]

A noncanonical GTPase signaling mechanism controls exit from mitosis in budding yeast.

Xiaoxue Zhou, Shannon Y Weng, Stephen P Bell, Angelika Amon.

In the budding yeast Saccharomyces cerevisiae , exit from mitosis is coupled to spindle position to ensure successful genome partitioning between mother and daughter cell. This coupling occurs through a GTPase signaling cascade known as the mitotic exit network (MEN). The MEN senses spindle position via a Ras-like GTPase Tem1 which primarily localizes to the spindle pole bodies (SPBs, yeast equivalent of centrosomes) during anaphase. How Tem1 couples the status of spindle position to MEN activation is not fully understood. Here, we show that Tem1 does not function as a molecular switch as its nucleotide state does not change upon MEN activation. Instead, Tem1's nucleotide state regulates its SPB localization to establish a concentration difference in the cell in response to spindle position. By artificially tethering Tem1 to the SPB, we demonstrate that the essential function of Tem1 GTP is to localize Tem1 to the SPB. Tem1 localization to the SPB primarily functions to generate a high effective concentration of Tem1 and MEN signaling can be initiated by concentrating Tem1 in the cytoplasm with genetically encoded multimeric nanoparticles. This localization/concentration-based GTPase signaling mechanism for Tem1 differs from the canonical Ras-like GTPase signaling paradigm and is likely relevant to other localization-based signaling scenarios.

DOI: https://doi.org/10.1101/2024.05.16.594582
bioRxiv. 2024 May 13. pii: 2023.11.14.567057. [Epub ahead of print]

Acentric chromosome congression and alignment on the metaphase plate via kinetochore-independent forces in Drosophila.

Hannah Vicars, Travis Karg, Alison Mills, William Sullivan.

Chromosome congression and alignment on the metaphase plate involves lateral and microtubule plus-end interactions with the kinetochore. Here we take advantage of our ability to efficiently generate a GFP-marked acentric X chromosome fragment in Drosophila neuroblasts to identify forces acting on chromosome arms that drive congression and alignment. We find acentrics efficiently align on the metaphase plate, often more rapidly than kinetochore-bearing chromosomes. Unlike intact chromosomes, the paired sister acentrics oscillate as they move to and reside on the metaphase plate in a plane distinct and significantly further from the main mass of intact chromosomes. Consequently, at anaphase onset acentrics are oriented either parallel or perpendicular to the spindle. Parallel-oriented sisters separate by sliding while those oriented perpendicularly separate via unzipping. This oscillation, together with the fact that in monopolar spindles acentrics are rapidly shunted away from the poles, indicates that distributed plus-end directed forces are primarily responsible for acentric migration. This conclusion is supported by the observation that reduction of EB1 preferentially disrupts acentric alignment. In addition, reduction of Klp3a activity, a gene required for the establishment of pole-to-pole microtubules, preferentially disrupts acentric alignment. Taken together these studies suggest that plus-end forces mediated by the outer pole-to-pole microtubules are primarily responsible for acentric metaphase alignment. Surprisingly, we find that a small fraction of sister acentrics are anti-parallel aligned indicating that the kinetochore is required to ensure parallel alignment of sister chromatids. Finally, we find induction of acentric chromosome fragments results in a global reorganization of the congressed chromosomes into a torus configuration.
Article Summary: The kinetochore serves as a site for attaching microtubules and allows for successful alignment, separation, and segregation of replicated sister chromosomes during cell division. However, previous studies have revealed that sister chromosomes without kinetochores (acentrics) often align to the metaphase plate, undergo separation and segregation, and are properly transmitted to daughter cells. In this study, we discuss the forces acting on chromosomes, independent of the kinetochore, underlying their successful alignment in early mitosis.

DOI: https://doi.org/10.1101/2023.11.14.567057
Nat Genet. 2024 May 27.

Genome folding principles uncovered in condensin-depleted mitotic chromosomes.

Han Zhao, Yinzhi Lin, En Lin, Fuhai Liu, Lirong Shu, Dannan Jing, Baiyue Wang, Manzhu Wang, Fengnian Shan, Lin Zhang, Jessica C Lam, Susannah C Midla, Belinda M Giardine, Cheryl A Keller, Ross C Hardison, Gerd A Blobel, Haoyue Zhang.

During mitosis, condensin activity is thought to interfere with interphase chromatin structures. To investigate genome folding principles in the absence of chromatin loop extrusion, we codepleted condensin I and condensin II, which triggered mitotic chromosome compartmentalization in ways similar to that in interphase. However, two distinct euchromatic compartments, indistinguishable in interphase, emerged upon condensin loss with different interaction preferences and dependencies on H3K27ac. Constitutive heterochromatin gradually self-aggregated and cocompartmentalized with facultative heterochromatin, contrasting with their separation during interphase. Notably, some cis-regulatory element contacts became apparent even in the absence of CTCF/cohesin-mediated structures. Heterochromatin protein 1 (HP1) proteins, which are thought to partition constitutive heterochromatin, were absent from mitotic chromosomes, suggesting, surprisingly, that constitutive heterochromatin can self-aggregate without HP1. Indeed, in cells traversing from M to G1 phase in the combined absence of HP1α, HP1β and HP1γ, constitutive heterochromatin compartments are normally re-established. In sum, condensin-deficient mitotic chromosomes illuminate forces of genome compartmentalization not identified in interphase cells.

DOI: https://doi.org/10.1038/s41588-024-01759-x
bioRxiv. 2024 May 14. pii: 2024.05.12.593765. [Epub ahead of print]

Temporal and anteriorly positioned mitotic zones drive asymmetric microtubule patterns needed for Left-Right Organizer development.

Yan Wu, Yiling Lan, Favour Ononiwu, Abigail Poole, Kirsten Rasmussen, Jonah Da Silva, Abdalla Wael Shamil, Li-En Jao, Heidi Hehnly.

Cellular proliferation plays a crucial role in tissue development, including the development of the Left-Right Organizer (LRO), the transient organ essential for dictating the vertebrate LR body plan. Here we investigate cell redistribution mechanisms and the dominance of specific progenitor cells in LRO formation, addressing cell lineage and cell behavior questions. Using zebrafish as a model, we mapped all LRO (Kupffer's Vesicle, KV) mitotic events, revealing an FGF-dependent, anteriorly enriched mitotic pattern. Using a KV-specific fluorescent microtubule (MT) line, we found that mitotic events align their spindle along the KV's longest axis until the rosette developmental stage, where "spinning" spindles followed by exclusion from KV occur. Daughter cells that remain are linked by cytokinetic bridges, shaping anteriorly focused MT patterns that precede apical actin recruitment. Our findings underscore the importance of spatially regulated mitotic events in establishing MT and actin pattern formation essential for LRO development.

DOI: https://doi.org/10.1101/2024.05.12.593765
Oncol Lett. 2024 Jul;28(1): 316

PLK1 inhibition leads to mitotic arrest and triggers apoptosis in cholangiocarcinoma cells.

Benchamart Moolmuang, Jittiporn Chaisaingmongkol, Pattama Singhirunnusorn, Mathuros Ruchirawat.

  Cholangiocarcinoma (CCA) is a lethal cancer originating from the epithelial cells within the bile duct and ranks as the second most prevalent form of liver cancer in Thailand. Polo-like kinase 1 (PLK1), a protein serine/threonine kinase, regulates a number of steps in cell mitosis and is upregulated in several types of cancer, including CCA. Our previous study identified PLK1 as a biomarker of the C1 subtype, correlating with poor prognosis in intrahepatic CCA. The present study aimed to examine the effect of PLK1 inhibition on CCA cells. Different CCA cell lines developed from Thai patients, HuCCA1, KKU055, KKU100 and KKU213A, were treated with two PLK1 inhibitors, BI2536 and BI6727, and were transfected with small interfering RNA, followed by analysis of cell proliferation, cell cycle distribution and cell apoptosis. It was discovered that BI2536 and BI6727 inhibited cell proliferation and caused G2/M-phase arrest in CCA cells. Furthermore, the number of total apoptotic cells was increased in PLK1 inhibitor-treated CCA cells. The expression levels of mitotic proteins, aurora kinase A, phosphorylated PLK1 (T210) and cyclin B1, were augmented in PLK1-inhibited CCA cells. Additionally, inhibition of PLK1 led to increased DNA damage, as determined by the upregulated levels of γH2AX and increased cleavage of poly (ADP-ribose) polymerase, an apoptotic marker. These results suggested that inhibiting PLK1 prolonged mitotic arrest and subsequently triggered cell apoptosis. Validation of the antiproliferative effects of PLK1 inhibition was accomplished through silencing of the PLK1 gene. In conclusion, targeting PLK1 provided promising results for further study as a potential candidate for targeted therapy in CCA.

Keywords:  BI2536; BI6727; G2/M-phase arrest; apoptosis; cholangiocarcinoma; polo-like kinase 1

DOI:  https://doi.org/10.3892/ol.2024.14449
J Biomed Res. 2024 Apr 25. 1-15

Mouse KL2 is a unique MTSE involved in chromosome-based spindle organization and regulated by multiple kinases during female meiosis.

Shi-Ya Xie, Yan-Jie Yang, Zhen Jin, Xiao-Cong Liu, Shu-Ping Zhang, Ning Su, Jia-Qi Liu, Cong-Rong Li, Dong Zhang, Lei-Lei Gao, Zhi-Xia Yang.

  Microtubule-severing enzymes (MTSEs) play important roles in mitosis and meiosis of the primitive organisms. However, no studies have assessed their roles in mammalian meiosis of females, whose abnormality accounts for over 80% of the cases of gamete-originated human reproductive disease. In the current study, we reported that katanin-like 2 (KL2) was the only MTSE concentrating at chromosomes. Furthermore, the knockdown of KL2 significantly reduced chromosome-based increase in the microtubule (MT) polymer, increased aberrant kinetochore-MT (K-MT) attachment, delayed meiosis, and severely affected normal fertility. Importantly, we demonstrated that the inhibition of aurora B, a key kinase for correcting aberrant K-MT attachment, eliminated KL2 from chromosomes completely. KL2 also interacted with phosphorylated eukaryotic elongation factor-2 kinase; they competed for chromosome binding. We also observed that the phosphorylated KL2 was localized at spindle poles, and that KL2 phosphorylation was regulated by extracellular signal-regulated kinase 1/2. In summary, our study reveals a novel function of MTSEs in mammalian female meiosis and demonstrates that multiple kinases coordinate to regulate the levels of KL2 at chromosomes.

Keywords:  KL2; MTSE; female meiosis; kinase; mouse

DOI:  https://doi.org/10.7555/JBR.37.20230290
Nature. 2024 May 29.

An alternative cell cycle coordinates multiciliated cell differentiation.

Semil P Choksi, Lauren E Byrnes, Mia J Konjikusic, Benedict W H Tsai, Rachel Deleon, Quanlong Lu, Christopher J Westlake, Jeremy F Reiter.

The canonical mitotic cell cycle coordinates DNA replication, centriole duplication and cytokinesis to generate two cells from one1. Some cells, such as mammalian trophoblast giant cells, use cell cycle variants like the endocycle to bypass mitosis2. Differentiating multiciliated cells, found in the mammalian airway, brain ventricles and reproductive tract, are post-mitotic but generate hundreds of centrioles, each of which matures into a basal body and nucleates a motile cilium3,4. Several cell cycle regulators have previously been implicated in specific steps of multiciliated cell differentiation5,6. Here we show that differentiating multiciliated cells integrate cell cycle regulators into a new alternative cell cycle, which we refer to as the multiciliation cycle. The multiciliation cycle redeploys many canonical cell cycle regulators, including cyclin-dependent kinases (CDKs) and their cognate cyclins. For example, cyclin D1, CDK4 and CDK6, which are regulators of mitotic G1-to-S progression, are required to initiate multiciliated cell differentiation. The multiciliation cycle amplifies some aspects of the canonical cell cycle, such as centriole synthesis, and blocks others, such as DNA replication. E2F7, a transcriptional regulator of canonical S-to-G2 progression, is expressed at high levels during the multiciliation cycle. In the multiciliation cycle, E2F7 directly dampens the expression of genes encoding DNA replication machinery and terminates the S phase-like gene expression program. Loss of E2F7 causes aberrant acquisition of DNA synthesis in multiciliated cells and dysregulation of multiciliation cycle progression, which disrupts centriole maturation and ciliogenesis. We conclude that multiciliated cells use an alternative cell cycle that orchestrates differentiation instead of controlling proliferation.

DOI: https://doi.org/10.1038/s41586-024-07476-z
Math Biosci. 2024 May 23. pii: S0025-5564(24)00079-8. [Epub ahead of print] 109219

A theoretical model of anaphase.

Brian D Sleeman, Iain W Stewart.

  This paper develops a theory for anaphase in cells. After a brief description of microtubules, the mitotic spindle and the centrosome, a mathematical model for anaphase is introduced and developed in the context of the cell cytoplasm and liquid crystalline structures. Prophase, prometaphase and metaphase are then briefly described in order to focus on anaphase, which is the main study of this paper. The entities involved are modelled in terms of liquid crystal defects and microtubules are represented as defect flux lines. The mathematical techniques employed make extensive use of energy considerations based on the work that was developed by Dafermos (1970) from the classical Frank-Oseen nematic liquid crystal energy (Frank, 1958; Oseen, 1933). With regard to liquid crystal theory we introduce the concept of regions of influence for defects which it is believed have important implications beyond the subject of this paper. The results of this paper align with observed biochemical phenomena and are explored in application to HeLa cells and Caenorhabditis elegans. This unified approach offers the possibility of gaining insight into various consequences of mitotic abnormalities which may result in Down syndrome, Hodgkin lymphoma, breast, prostate and various other types of cancer.

Keywords:  Anaphase; Caenorhabditis elegans; Cytoplasm; Dafermos energy; Defects; HeLa cell; Microtubules; Nematic liquid crystal; Regions of influence

DOI:  https://doi.org/10.1016/j.mbs.2024.109219