bims-micesi 2022-10-23 papers

bims-micesi

Biomed News

on Mitotic cell signalling

Issue of 2022–10–23
eleven papers selected by
Valentina Piano, Max Planck Institute of Molecular Physiology

APC/CCdc20-mediated degradation of Clb4 prompts astral microtubule stabilization at anaphase onset.
Dephosphorylation in nuclear reassembly after mitosis.
Phosphosites of the yeast centrosome component Spc110 contribute to cell cycle progression and mitotic exit.
Augmin prevents merotelic attachments by promoting proper arrangement of bridging and kinetochore fibers.
Intracellular infection by symbiotic bacteria requires the mitotic kinase AURORA1.
Maize CDKA2;1a and CDKB1;1 kinases have different requirements for their activation and participate in substrate recognition.
Inhibition of MAD2L1 Mediates Pulmonary Fibrosis through Impairment of Mitochondrial Function and Induction of Cell Senescence.
Regulation of the mitotic chromosome folding machines.
RAD6 inhibition enhances paclitaxel sensitivity of triple negative breast cancer cells by aggravating mitotic spindle damage.
Spindle and kinetochore-associated complex subunit 3 could serve as a prognostic biomarker for prostate cancer.
Targeting Cdc20 for cancer therapy.

J Cell Biol. 2023 Jan 02. pii: e202203089. [Epub ahead of print]222(1):

APC/CCdc20-mediated degradation of Clb4 prompts astral microtubule stabilization at anaphase onset.

Federico Zucca, Clara Visintin, Jiaming Li, Steven P Gygi, Rosella Visintin.

Key for accurate chromosome partitioning to the offspring is the ability of mitotic spindle microtubules to respond to different molecular signals and remodel their dynamics accordingly. Spindle microtubules are conventionally divided into three classes: kinetochore, interpolar, and astral microtubules (kMTs, iMTs, and aMTs, respectively). Among all, aMT regulation remains elusive. Here, we show that aMT dynamics are tightly regulated. aMTs remain unstable up to metaphase and are stabilized at anaphase onset. This switch in aMT dynamics, important for proper spindle orientation, specifically requires the degradation of the mitotic cyclin Clb4 by the Anaphase Promoting Complex bound to its activator subunit Cdc20 (APC/CCdc20). These data highlight a unique role for mitotic cyclin Clb4 in controlling aMT regulating factors, of which Kip2 is a prime candidate, provide a framework to understand aMT regulation in vertebrates, and uncover mechanistic principles of how the APC/CCdc20 choreographs the timing of late mitotic events by sequentially impacting on the three classes of spindle microtubules.

DOI: https://doi.org/10.1083/jcb.202203089
Front Cell Dev Biol. 2022 ;10 1012768

Dephosphorylation in nuclear reassembly after mitosis.

Vincent Archambault, Jingjing Li, Virginie Emond-Fraser, Myreille Larouche.

  In most animal cell types, the interphase nucleus is largely disassembled during mitotic entry. The nuclear envelope breaks down and chromosomes are compacted into separated masses. Chromatin organization is also mostly lost and kinetochores assemble on centromeres. Mitotic protein kinases play several roles in inducing these transformations by phosphorylating multiple effector proteins. In many of these events, the mechanistic consequences of phosphorylation have been characterized. In comparison, how the nucleus reassembles at the end of mitosis is less well understood in mechanistic terms. In recent years, much progress has been made in deciphering how dephosphorylation of several effector proteins promotes nuclear envelope reassembly, chromosome decondensation, kinetochore disassembly and interphase chromatin organization. The precise roles of protein phosphatases in this process, in particular of the PP1 and PP2A groups, are emerging. Moreover, how these enzymes are temporally and spatially regulated to ensure that nuclear reassembly progresses in a coordinated manner has been partly uncovered. This review provides a global view of nuclear reassembly with a focus on the roles of dephosphorylation events. It also identifies important open questions and proposes hypotheses.

Keywords:  chromosome decondensation; kinetochore disassembly; mitosis; mitotic exit; nuclear envelope; nucleolus; nucleus; phosphatase

DOI:  https://doi.org/10.3389/fcell.2022.1012768
Biol Open. 2022 Oct 19. pii: bio.059565. [Epub ahead of print]

Phosphosites of the yeast centrosome component Spc110 contribute to cell cycle progression and mitotic exit.

Marjan Abbasi, Alexander Julner, Yan Ting Lim, Tianyun Zhao, Radoslaw Mikolaj Sobota, Victoria Menéndez-Benito.

  Spc110 is an essential component of the spindle pole body (SPB), the yeast equivalent of the centrosome, that recruits the γ-tubulin complex to the nuclear side of the SPB to produce the microtubules that form the mitotic spindle. Here we identified phosphosites S11 and S36 in maternally originated Spc110 and explored their functions in vivo. Yeast expressing non-phosphorylatable Spc110S11A had a distinct spindle phenotype characterized by higher levels of α-tubulin, which was frequently asymmetrically distributed between the two SPBs. Furthermore, expression of the double mutant Spc110S11AS36A had a delayed cell cycle progression. Specifically, the final steps of mitosis were delayed in Spc110S11AS36A cells, including expression and degradation of the mitotic cyclin Clb2, disassembling the mitotic spindle and re-localizing Cdc14 to the nucleoli, resulting in late mitotic exit and entry in G1. Thus, we propose that Spc110 phosphorylation at S11 and S36 is required to regulate timely cell cycle progression in budding yeast.

Keywords:  Cdc14; Microtubule; Mitotic spindle; Phosphorylation; Spindle pole body; Yeast

DOI:  https://doi.org/10.1242/bio.059565
Elife. 2022 Oct 21. pii: e83287. [Epub ahead of print]11

Augmin prevents merotelic attachments by promoting proper arrangement of bridging and kinetochore fibers.

Valentina Štimac, Isabella Koprivec, Martina Manenica, Juraj Simunić, Iva M Tolić.

  The human mitotic spindle is made of microtubules nucleated at centrosomes, at kinetochores, and from pre-existing microtubules by the augmin complex. However, it is unknown how the augmin-mediated nucleation affects distinct microtubule classes and thereby mitotic fidelity. Here we use superresolution microscopy to analyze the previously indistinguishable microtubule arrangements within the crowded metaphase plate area and demonstrate that augmin is vital for the formation of uniformly arranged parallel units consisting of sister kinetochore fibers connected by a bridging fiber. This ordered geometry helps both prevent and resolve merotelic attachments. Whereas augmin-nucleated bridging fibers prevent merotelic attachments by creating a nearly parallel and highly bundled microtubule arrangement unfavorable for creating additional attachments, augmin-nucleated k-fibers produce robust force required to resolve errors during anaphase. STED microscopy revealed that bridging fibers were impaired twice as much as k-fibers following augmin depletion. The complete absence of bridging fibers from a significant portion of kinetochore pairs, especially in the inner part of the spindle, resulted in the specific reduction of the interkinetochore distance. Taken together, we propose a model where augmin promotes mitotic fidelity by generating assemblies consisting of bridging and kinetochore fibers that align sister kinetochores to face opposite poles, thereby preventing erroneous attachments.

Keywords:  cancer biology; cell biology; none

DOI:  https://doi.org/10.7554/eLife.83287
Proc Natl Acad Sci U S A. 2022 Oct 25. 119(43): e2202606119

Intracellular infection by symbiotic bacteria requires the mitotic kinase AURORA1.

Jin-Peng Gao, Suyu Jiang, Yangyang Su, Ping Xu, Junjie Wang, Wenjie Liang, Cheng-Wu Liu, Jeremy D Murray.

  The subcellular events occurring in cells of legume plants as they form transcellular symbiotic-infection structures have been compared with those occurring in premitotic cells. Here, we demonstrate that Aurora kinase 1 (AUR1), a highly conserved mitotic regulator, is required for intracellular infection by rhizobia in Medicago truncatula. AUR1 interacts with microtubule-associated proteins of the TPXL and MAP65 families, which, respectively, activate and are phosphorylated by AUR1, and localizes with them within preinfection structures. MYB3R1, a rhizobia-induced mitotic transcription factor, directly regulates AUR1 through two closely spaced, mitosis-specific activator cis elements. Our data are consistent with a model in which the MYB3R1-AUR1 regulatory module serves to properly orient preinfection structures to direct the transcellular deposition of cell wall material for the growing infection thread, analogous to its role in cell plate formation. Our findings indicate that the eukaryotically conserved MYB3R1-TPXL-AUR1-MAP65 mitotic module was conscripted to support endosymbiotic infection in legumes.

Keywords:  Aurora kinase; Medicago truncatula; cell cycle; microtubule; rhizobial infection

DOI:  https://doi.org/10.1073/pnas.2202606119
FEBS J. 2022 Oct 19.

Maize CDKA2;1a and CDKB1;1 kinases have different requirements for their activation and participate in substrate recognition.

Mingyar N López-Hernández, Jorge M Vázquez-Ramos.

  Cyclin-dependent kinases (CDKs), in association with cyclins, control cell cycle progression by phosphorylating a large number of substrates. In animals, activation of CDKs regularly requires both the association to a cyclin and then phosphorylation of a highly conserved threonine residue in the CDK activation loop (the classical mechanism), mediated by a CDK-activating kinase (CAK). In addition to this typical mechanism of activation, some CDKs can also be activated by the association of a cyclin to a monomeric CDK previously phosphorylated by CAK although not all CDKs can be activated by this mechanism. In animals and yeast, cyclin, in addition to being required for CDK activation provides substrate specificity to the cyclin/CDK complex, however, in plants both the mechanisms of CDKs activation, and the relevance of the CDK-associated cyclin for substrate targeting have been poorly studied. In this work, by co-expressing proteins in E. coli, we studied maize CDKA2;1a and CDKB1;1, two of the main types of CDKs that control the cell cycle in plants. These kinases could be activated by the classical mechanism and by the association of CycD2;2a to a phosphorylated intermediate in its activation loop, a previously unproven mechanism for the activation of plant CDKs. Unlike CDKA2;1a, CDKB1;1 did not require CAK for its activation, since it autophosphorylated in its activation loop. Phosphorylation of CDKB1;1 and association of CycD2;2 was not enough for its full activation as association of maize CKS, a scaffolding protein, differentially stimulated substrate phosphorylation. Our results suggest that both CDKs participate in substrate recognition.

Keywords:  Activation mechanism; Autophosphorylation; Maize CDKs; Substrate recognition

DOI:  https://doi.org/10.1111/febs.16659
Can Respir J. 2022 ;2022 9663354

Inhibition of MAD2L1 Mediates Pulmonary Fibrosis through Impairment of Mitochondrial Function and Induction of Cell Senescence.

Lan Wang, Ruyan Wan, Xinyu Chen, Xiaoshu Guo, Zhongzheng Li, Weiming Zhao, Peishuo Yan, Guoying Yu.

Idiopathic pulmonary fibrosis (IPF) is a chronic, irreversible, and progressive interstitial lung disease characterized by recurrent alveolar epithelial cell injury, fibroblast hyperproliferation, and cumulative deposition of extracellular matrix leading to alveolar destruction in the lungs. Mitotic arrest deficient 2 like 1 (MAD2L1) is a component of the mitotic spindle assembly checkpoint that prevents the onset of anaphase until all chromosomes are properly aligned at metaphase and is a potential therapeutic target in cancers. However, the role of MAD2L1 in pulmonary fibrosis has not been explored. We analyzed the expression of MAD2L1 in lung tissues from control subjects, IPF patients, and mice with bleomycin-induced fibrosis via IHC, qRT-PCR, and Western blot analysis. We examined the roles of MAD2L1 in ROS production, mitochondrial function, cell senescence, and the establishment of a profibrotic microenvironment. We found that MAD2L1 was highly upregulated in alveolar epithelial cells in fibrotic lung tissues from both patients with IPF and mice with bleomycin-induced fibrosis. Loss of MAD2L1 expression or activity led to decreases of cell viability and proliferation in A549 cells. Subsequent mechanistic investigation demonstrated that inhibition of MAD2L1 damaged mitochondria, which led to augmented ROS production and cellular senescence, and thus promoted the establishment of a profibrotic microenvironment. Taken together, these results reveal that alleviation of alveolar epithelial cell mitochondrial damage arising from augmentation of MAD2L1 may be a novel therapeutic strategy for mitigating pulmonary fibrosis.

DOI: https://doi.org/10.1155/2022/9663354
Biochem J. 2022 Oct 28. 479(20): 2153-2173

Regulation of the mitotic chromosome folding machines.

Bastiaan Dekker, Job Dekker.

  Over the last several years enormous progress has been made in identifying the molecular machines, including condensins and topoisomerases that fold mitotic chromosomes. The discovery that condensins generate chromatin loops through loop extrusion has revolutionized, and energized, the field of chromosome folding. To understand how these machines fold chromosomes with the appropriate dimensions, while disentangling sister chromatids, it needs to be determined how they are regulated and deployed. Here, we outline the current understanding of how these machines and factors are regulated through cell cycle dependent expression, chromatin localization, activation and inactivation through post-translational modifications, and through associations with each other, with other factors and with the chromatin template itself. There are still many open questions about how condensins and topoisomerases are regulated but given the pace of progress in the chromosome folding field, it seems likely that many of these will be answered in the years ahead.

Keywords:  cell cycle; chromosomes; loop extrusion; mitotis

DOI:  https://doi.org/10.1042/BCJ20210140
BMC Cancer. 2022 Oct 18. 22(1): 1073

RAD6 inhibition enhances paclitaxel sensitivity of triple negative breast cancer cells by aggravating mitotic spindle damage.

Brittany M Haynes, Kristen Cunningham, Malathy P V Shekhar.

   BACKGROUND: Paclitaxel (PTX), a first-line therapy for triple negative breast cancers (TNBC) induces anti-tumor activity by microtubule stabilization and inhibition of cell division. Its dose-limiting toxicity and short half-life, however, pose clinical challenges underscoring the need for strategies that increase its efficiency. RAD6, a E2 ubiquitin conjugating enzyme, is associated with centrosomes at all phases of cell cycle. Constitutive overexpression of the RAD6B homolog in normal breast cells induces centrosome amplification and multipolar spindle formation, indicating its importance in centrosome regulation.
METHODS: TNBC centrosome numbers were scored by pericentrin immunostaining. PTX sensitivities and interactions with SMI#9, a RAD6-selective small molecule inhibitor, on TNBC cell survival were analyzed by MTT and colony forming assays and an isogenic MDA-MB-468 TNBC model of PTX resistance. The molecular mechanisms underlying PTX and SMI#9 induced cytotoxicity were determined by flow cytometry, immunoblot analysis of cyclin B1 and microtubule associated protein TAU, and dual immunofluorescence staining of TAU and α-tubulin.
RESULTS: Our data show aberrant centrosome numbers and that PTX sensitivities are not correlated with TNBC BRCA1 status. Combining PTX with SMI#9 synergistically enhances PTX sensitivities of BRCA1 wild-type and mutant TNBC cells. Whereas SMI#9/PTX combination treatment increased cyclin B1 levels in MDA-MB-468 cells, it induced cyclin B1 loss in HCC1937 cells with accumulation of reproductively dead giant cells, a characteristic of mitotic catastrophe. Cell cycle analysis revealed drug-induced accumulation of tetraploid cells in S and G2/M phases, and robust increases in cells with 4 N DNA content in HCC1937 cells. TAU overexpression is associated with reduced PTX efficacy. Among the six TAU isoforms, both SMI#9 and PTX downregulated 1N3R TAU in MDA-MB-468 and HCC1937 cells, suggesting a common mechanism of 1N3R regulation. Dual TAU and α-tubulin immunostaining showed that SMI#9 induces monopolar mitotic spindles. Using the isogenic model of PTX resistance, we show that SMI#9 treatment restores PTX sensitivity.
CONCLUSIONS: These data support a common mechanism of microtubule regulation by SMI#9 and PTX and suggest that combining PTX with RAD6 inhibitor may be beneficial for increasing TNBC sensitivities to PTX and alleviating toxicity. This study demonstrates a new role for RAD6 in regulating microtubule dynamics.

Keywords:  Acquired resistance; BRCA1; Cyclin B1; Paclitaxel; RAD6; TAU

DOI:  https://doi.org/10.1186/s12885-022-10119-z
Exp Hematol Oncol. 2022 Oct 20. 11(1): 76

Spindle and kinetochore-associated complex subunit 3 could serve as a prognostic biomarker for prostate cancer.

Dechao Feng, Weizhen Zhu, Xu Shi, Qiao Xiong, Dengxiong Li, Wuran Wei, Ping Han, Qiang Wei, Lu Yang.

  Spindle and kinetochore-associated complex subunit 3 (SKA3) is a microtubule-binding subcomplex of the outer kinetochore that is required for proper chromosomal segregation and cell division. However, little is known regarding the probable mechanism of SKA3, particularly in terms of prostate cancer (PCA) progression. Multiple databases, including TCGA and GTEx, were utilized to examine the expression of SKA3 in PCA patients and to shed light on the clinical significance and potential mechanism of SKA3 in the onset and progression of PCA. The biological function of SKA3 was evaluated in vitro using RT-qPCR and the CCK8 assay. For statistical analysis, the R 3.6.3 software and its associated packages were utilized. SKA3 was shown to be considerably elevated in PCA patients and was linked to a shorter progress free interval (PFI). Furthermore, we discovered that SKA3 mRNA expression was higher in PCA cells than in normal cells, and inhibition of SKA3 could clearly reduce PCA cell proliferation using the CCK8 assay. Finally, SKA3 could be used as a predictive biomarker in PCA patients.

Keywords:  Biomarker; Enrichment analysis; Prostate cancer; Spindle and kinetochore associated complex subunit 3; Targeted therapy

DOI:  https://doi.org/10.1186/s40164-022-00337-3
Biochim Biophys Acta Rev Cancer. 2022 Oct 12. pii: S0304-419X(22)00149-4. [Epub ahead of print]1877(6): 188824

Targeting Cdc20 for cancer therapy.

Seung Min Jeong, Quyen Thu Bui, Minseok Kwak, Ji Yeon Lee, Peter Chang-Whan Lee.

  The Anaphase-Promoting Complex/Cyclosome (APC/C), an E3 ubiquitin ligase, and two co-activators, Cdc20 and Cdh1, enable the ubiquitin-dependent proteasomal degradation of various critical cell cycle regulators and govern cell division in a timely and precise manner. Dysregulated cell cycle events cause uncontrolled cell proliferation, leading to tumorigenesis. Studies have shown that Cdh1 has tumor suppressive activities while Cdc20 has an oncogenic property, suggesting that Cdc20 is an emerging therapeutic target for cancer treatment. Therefore, in this review, we discussed recent findings about the essential roles of APC/C-Cdc20 in cell cycle regulation. Furthermore, we briefly summarized that the regulation of Cdc20 expression levels is strictly controlled to order cell cycle events appropriately. Finally, given the function of Cdc20 as an oncogene, therapeutic interventions targeting Cdc20 activity may be beneficial in cancer treatment.

Keywords:  Cancer; Cdc20; Cell cycle; E3 ligase; cancer therapy

DOI:  https://doi.org/10.1016/j.bbcan.2022.188824