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David M. Glover - One of the best experts on this subject based on the ideXlab platform.

  • APC/C-Vihar regulates centrosome activity and stability in the Drosophila germline
    2017
    Co-Authors: Alexis L. Braun, David M. Glover, Francesco Meghini, Gema Villa-fombuena, Elisa M. Fernandez, Morgane Guermont, María D. Martín-bermudo, Acaimo González-reyes, Yuu Kimata
    Abstract:

    A universal feature of metazoan reproduction is the elimination of the maternal centrosomes prior to the end of oogenesis. In animals that have a syncytial cyst stage of oocyte development, including Drosophila and mouse, the germline centrosomes undergo a migration to all reside within the oocyte. However, the functional significance of centrosome transport within the female germline and the mechanism orchestrating this event are still a mystery. The Anaphase Promoting Complex/Cyclosome (APC/C) is a multi-subunit ubiquitin ligase (E3) that temporally regulates progression of the cell cycle as well as the centrosome cycle. By altering the negative regulation of the cooperating ubiquitin conjugating enzyme (E2), Vihar/Ube2c, we show that temporal control of APC/C activity ensures centrosome stability and migration during early Drosophila oogenesis. When there is perduring APC/C activity, Polo Kinase is precociously targeted for destruction, which results in centriole instability and decreased centrosome transport to the oocyte. We show that decreased centrosome transport correlates with a decreased accumulation of pericentriolar material (PCM) proteins on the oocyte nucleus, which results in a weakening of the structural integrity of the egg chamber and loss of oocyte fate - the overall consequence being a reduction in female fertility. Considering the conserved roles of the APC/C and Polo Kinase throughout the animal kingdom and the fact that many animals have a syncytial stage of egg development, our results provide insight into the general necessity of gametic centrosome transport for female fertility.

  • Inhibition of Polo Kinase by BI2536 affects centriole separation during Drosophila male meiosis
    Cell cycle (Georgetown Tex.), 2014
    Co-Authors: Maria Riparbelli, Marco Gottardo, David M. Glover, Giuliano Callaini
    Abstract:

    Pharmacological inhibition of Drosophila Polo Kinase with BI2536 has allowed us to re-examine the requirements for Polo during Drosophila male gametogenesis. BI2536-treated spermatocytes persisted in a pro-metaphase state without dividing and had condensed chromosomes that did not separate. Centrosomes failed to recruit γ-tubulin and centrosomin (Cnn) and were not associated with microtubule arrays that were abnormal and did not form proper bipolar spindles. Centrioles, which usually separate during the anaphase of the first meiosis, remained held together in a V-shaped configuration suggesting that Polo Kinase regulates the proteolysis that breaks centriole linkage to ensure their disengagement. Despite these defects spermatid differentiation proceeds, leading to axoneme formation.

  • Correction: The Chromosomal Passenger Complex Activates Polo Kinase at Centromeres
    PLoS Biology, 2012
    Co-Authors: Mar Carmena, Xavier Pinson, Melpi Platani, Zeina Salloum, Anthony Clark, Fiona Macisaac, Hiromi Ogawa, Ulrike S. Eggert, David M. Glover
    Abstract:

    Figure S5 is a duplicate of Figure S7. Please view the correct Figure S5 here: Click here for additional data file.(3.2M, tif)

  • The Chromosomal Passenger Complex Activates Polo Kinase at Centromeres
    PLoS biology, 2012
    Co-Authors: Mar Carmena, Xavier Pinson, Melpi Platani, Zeina Salloum, Anthony Clark, Fiona Macisaac, Hiromi Ogawa, Ulrike S. Eggert, David M. Glover
    Abstract:

    The coordinated activities at centromeres of two key cell cycle Kinases, Polo and Aurora B, are critical for ensuring that the two sister kinetochores of each chromosome are attached to microtubules from opposite spindle poles prior to chromosome segregation at anaphase. Initial attachments of chromosomes to the spindle involve random interactions between kinetochores and dynamic microtubules, and errors occur frequently during early stages of the process. The balance between microtubule binding and error correction (e.g., release of bound microtubules) requires the activities of Polo and Aurora B Kinases, with Polo promoting stable attachments and Aurora B promoting detachment. Our study concerns the coordination of the activities of these two Kinases in vivo. We show that INCENP, a key scaffolding subunit of the chromosomal passenger complex (CPC), which consists of Aurora B Kinase, INCENP, Survivin, and Borealin/Dasra B, also interacts with Polo Kinase in Drosophila cells. It was known that Aurora A/Bora activates Polo at centrosomes during late G2. However, the Kinase that activates Polo on chromosomes for its critical functions at kinetochores was not known. We show here that Aurora B Kinase phosphorylates Polo on its activation loop at the centromere in early mitosis. This phosphorylation requires both INCENP and Aurora B activity (but not Aurora A activity) and is critical for Polo function at kinetochores. Our results demonstrate clearly that Polo Kinase is regulated differently at centrosomes and centromeres and suggest that INCENP acts as a platform for Kinase crosstalk at the centromere. This crosstalk may enable Polo and Aurora B to achieve a balance wherein microtubule mis-attachments are corrected, but proper attachments are stabilized allowing proper chromosome segregation.

  • Aurora B activity is required for the activation of Polo Kinase at the inner centromere.
    2012
    Co-Authors: Mar Carmena, Xavier Pinson, Melpi Platani, Zeina Salloum, Anthony Clark, Fiona Macisaac, Hiromi Ogawa, Ulrike Eggert, David M. Glover
    Abstract:

    (A) Aurora B phosphorylates Polo Kinase in vitro. Bacterially expressed HIS-Polo or HIS-PoloT182A (which is catalytically inactive and therefore unable to autophosphorylate) were incubated with (or without) Drosophila Aurora B in complex with a fragment of INCENP (residues 654–755) in presence of 32P-g-ATP, in triplicate. Reaction products were resolved by SDS-PAGE transferred to nitrocellulose and analyzed by autoradiography (AR) and anti-Polo Western blot (WB). Quantitative measurements of signals were obtained (see Materials and Methods), and the ratios were calculated for each reaction (AR/WB, A.U.: arbitrary units). Right, average values for the relative phosphorylatin of PoloWT and PoloT182A by Aurora B. Error bars, SEM. (B–D) DMel-2 cells stably expressing Polo-GFP treated with (B) DMSO or (C–D) Binucleine-2, immunostained for INCENP, Polo, and PoloT182Ph (insets: zoomed images of kinetochores). In (C–D) asterisks indicate centrosomes. Merged images show INCENP/Polo/DNA. Zoomed images in (C–D) insets show examples of kinetochore pairs showing decreased levels of PoloT182Ph. (E) Dot plot showing the quantification of INCENP/Polo/PoloT182Ph signal intensity at the kinetochore (t test: *** p

Simonetta Piatti - One of the best experts on this subject based on the ideXlab platform.

  • Silencing the spindle assembly checkpoint: Let's play Polo!
    The Journal of cell biology, 2020
    Co-Authors: Giorgia Benzi, Simonetta Piatti
    Abstract:

    Silencing of the spindle assembly checkpoint involves two protein phosphatases, PP1 and PP2A-B56, that are thought to extinguish checkpoint signaling through dephosphorylation of a checkpoint scaffold at kinetochores. In this issue, Cordeiro et al. (2020. J. Cell Biol.https://doi.org/10.1083/jcb.202002020) now show that a critical function of these phosphatases in checkpoint silencing is removal of Polo Kinase at kinetochores, which would otherwise autonomously sustain the checkpoint.

  • The RSC chromatin-remodeling complex influences mitotic exit and adaptation to the spindle assembly checkpoint by controlling the Cdc14 phosphatase
    The Journal of cell biology, 2010
    Co-Authors: Valentina Rossio, Matteo Ferrari, Achille Pellicioli, Elena Galati, Takashi Sutani, Katsuhiko Shirahige, Giovanna Lucchini, Simonetta Piatti
    Abstract:

    Upon prolonged activation of the spindle assembly checkpoint, cells escape from mitosis through a mechanism called adaptation or mitotic slippage, which is thought to underlie the resistance of cancer cells to antimitotic drugs. We show that, in budding yeast, this mechanism depends on known essential and nonessential regulators of mitotic exit, such as the Cdc14 early anaphase release (FEAR) pathway for the release of the Cdc14 phosphatase from the nucleolus in early anaphase. Moreover, the RSC (remodel the structure of chromatin) chromatin-remodeling complex bound to its accessory subunit Rsc2 is involved in this process as a novel component of the FEAR pathway. We show that Rsc2 interacts physically with the Polo Kinase Cdc5 and is required for timely phosphorylation of the Cdc14 inhibitor Net1, which is important to free Cdc14 in the active form. Our data suggest that fine-tuning regulators of mitotic exit have important functions during mitotic progression in cells treated with microtubule poisons and might be promising targets for cancer treatment.

Hongguo Yu - One of the best experts on this subject based on the ideXlab platform.

  • is cohesin required for spindle pole body centrosome cohesion
    Communicative & Integrative Biology, 2012
    Co-Authors: Martin Avey, Hongguo Yu
    Abstract:

    Centrosomes are microtubule-organizing centers that nucleate spindle microtubules during cell division. In budding yeast, the centrosome, often referred to as the spindle pole body, shares structural components with the centriole, the central core of the animal centrosome. The parental centrosome is duplicated when DNA replication takes place. Like sister chromatids tethered together by cohesin, duplicated centrosomes are linked and then separate to form the bipolar spindle necessary for chromosome segregation. Recent studies have shown that cohesin is also localized to the animal centrosome and is perhaps directly involved in engaging paired centrioles. Here we discuss the potential role of cohesin in mediating spindle-pole-body cohesion in the context of yeast meiosis. We propose that the coordination of chromosome segregation with centrosome cohesion and duplication is mediated by the antagonistic interaction between the Aurora Kinase and the Polo Kinase and that the role of cohesin in centrosome regulation appears to be indirect in budding yeast.

  • Is cohesin required for spindle-pole-body/centrosome cohesion?
    Communicative & Integrative Biology, 2012
    Co-Authors: Martin Avey, Hongguo Yu
    Abstract:

    Centrosomes are microtubule-organizing centers that nucleate spindle microtubules during cell division. In budding yeast, the centrosome, often referred to as the spindle pole body, shares structural components with the centriole, the central core of the animal centrosome. The parental centrosome is duplicated when DNA replication takes place. Like sister chromatids tethered together by cohesin, duplicated centrosomes are linked and then separate to form the bipolar spindle necessary for chromosome segregation. Recent studies have shown that cohesin is also localized to the animal centrosome and is perhaps directly involved in engaging paired centrioles. Here we discuss the potential role of cohesin in mediating spindle-pole-body cohesion in the context of yeast meiosis. We propose that the coordination of chromosome segregation with centrosome cohesion and duplication is mediated by the antagonistic interaction between the Aurora Kinase and the Polo Kinase and that the role of cohesin in centrosome regulation appears to be indirect in budding yeast.

Claudio E Sunkel - One of the best experts on this subject based on the ideXlab platform.

  • Polo regulates spindly to prevent premature stabilization of kinetochore microtubule attachments
    The EMBO Journal, 2020
    Co-Authors: Claudio E Sunkel, João Barbosa, Torcato Martins, Tanja Bange, Li Tao, Carlos Conde
    Abstract:

    Accurate chromosome segregation in mitosis requires sister kinetochores to bind to microtubules from opposite spindle poles. The stability of kinetochore-microtubule attachments is fine-tuned to prevent or correct erroneous attachments while preserving amphitelic interactions. Polo Kinase has been implicated in both stabilizing and destabilizing kinetochore-microtubule attachments. However, the mechanism underlying Polo-destabilizing activity remains elusive. Here, resorting to an RNAi screen in Drosophila for suppressors of a constitutively active Polo mutant, we identified a strong genetic interaction between Polo and the Rod-ZW10-Zwilch (RZZ) complex, whose kinetochore accumulation has been shown to antagonize microtubule stability. We find that Polo phosphorylates Spindly and impairs its ability to bind to Zwilch. This precludes dynein-mediated removal of the RZZ from kinetochores and consequently delays the formation of stable end-on attachments. We propose that high Polo-Kinase activity following mitotic entry directs the RZZ complex to minimize premature stabilization of erroneous attachments, whereas a decrease in active Polo in later mitotic stages allows the formation of stable amphitelic spindle attachments. Our findings demonstrate that Polo tightly regulates the RZZ-Spindly-dynein module during mitosis to ensure the fidelity of chromosome segregation.

  • Polo regulates Spindly to prevent premature stabilization of kinetochore–microtubule attachments
    The EMBO journal, 2019
    Co-Authors: João Barbosa, Torcato Martins, Tanja Bange, Li Tao, Carlos Conde, Claudio E Sunkel
    Abstract:

    Accurate chromosome segregation in mitosis requires sister kinetochores to bind to microtubules from opposite spindle poles. The stability of kinetochore-microtubule attachments is fine-tuned to prevent or correct erroneous attachments while preserving amphitelic interactions. Polo Kinase has been implicated in both stabilizing and destabilizing kinetochore-microtubule attachments. However, the mechanism underlying Polo-destabilizing activity remains elusive. Here, resorting to an RNAi screen in Drosophila for suppressors of a constitutively active Polo mutant, we identified a strong genetic interaction between Polo and the Rod-ZW10-Zwilch (RZZ) complex, whose kinetochore accumulation has been shown to antagonize microtubule stability. We find that Polo phosphorylates Spindly and impairs its ability to bind to Zwilch. This precludes dynein-mediated removal of the RZZ from kinetochores and consequently delays the formation of stable end-on attachments. We propose that high Polo-Kinase activity following mitotic entry directs the RZZ complex to minimize premature stabilization of erroneous attachments, whereas a decrease in active Polo in later mitotic stages allows the formation of stable amphitelic spindle attachments. Our findings demonstrate that Polo tightly regulates the RZZ-Spindly-dynein module during mitosis to ensure the fidelity of chromosome segregation.

  • The Polo Kinase is required at multiple stages during spermatogenesis in
    Chromosoma, 1998
    Co-Authors: Siegrun Herrmann, Isabel Amorim, Claudio E Sunkel
    Abstract:

    The Polo gene of Drosophila melanogaster is the founding member of the Polo-like Kinase family which is conserved among eukaryotes. Polo has been implicated in the organisation and function of the mitotic apparatus. Furthermore, Polo has been shown to be required for normal spermatogenesis. To characterize further the role of Polo in spermatogenesis, Polo mutants were analysed by immunostaining with specific antibodies and phase contrast microscopy. Immunofluorescence shows that Polo localises to the centrosomes, the centromere/kinetochore and the spindle midzone. The meiotic phenotype of various mutant allelic combinations was also studied in detail. Observation of mutant live testes indicates cytological abnormalities in all meiotic cell types, including variable DNA content and multipolar spindles. Primary spermatocytes in Polo mutant testes contain an abnormal DNA content, suggesting failure of chromosome segregation during gonial division. Immunostaining of Polo mutant cells with α-tubulin shows several abnormalities of the meiotic spindle, including a significantly reduced central spindle. Our results suggest that Polo has multiple functions during spermatogenesis.

Iain M Hagan - One of the best experts on this subject based on the ideXlab platform.

  • Centrosomal MPF triggers the mitotic and morphogenetic switches of fission yeast
    Nature cell biology, 2012
    Co-Authors: Agnes Grallert, Avinash Patel, Victor A. Tallada, Kuan Yoow Chan, Steven Bagley, Andrea Krapp, Viesturs Simanis, Iain M Hagan
    Abstract:

    The Cdc2 (also called Cdk1) Kinase is first activated at the centrosome to initiate mitosis in human cells. Hagan and colleagues demonstrate that in fission yeast, Cdc2 and Polo Kinase activation at the spindle pole body remotely controls not only mitotic commitment but also ‘new end take off’, the initiation of bipolar growth in G2.

  • the spindle pole body plays a key role in controlling mitotic commitment in the fission yeast schizosaccharomyces pombe
    Biochemical Society Transactions, 2008
    Co-Authors: Iain M Hagan
    Abstract:

    Commitment to mitosis is regulated by a conserved protein Kinase complex called MPF (mitosis-promoting factor). MPF activation triggers a positive-feedback loop that further promotes the activity of its activating phosphatase Cdc25 and is assumed to down-regulate the MPF-inhibitory Kinase Wee1. Four protein Kinases contribute to this amplification loop: MPF itself, Polo Kinase, MAPK (mitogen-activated protein Kinase) and Greatwall Kinase. The fission yeast SPB (spindle pole body) component Cut12 plays a critical role in modulating mitotic commitment. In this review, I discuss the relationship between Cut12 and the fission yeast Polo Kinase Plo1 in mitotic control. These results indicate that commitment to mitosis is co-ordinated by control networks on the spindle pole. I then describe how the Cut12/Plo1 control network links growth control signalling from TOR (target of rapamycin) and MAPK networks to the activation of MPF to regulate the timing of cell division.

  • Polo Kinase links the stress pathway to cell cycle control and tip growth in fission yeast
    Nature, 2005
    Co-Authors: Janni Petersen, Iain M Hagan
    Abstract:

    Stress-activated mitogen-activated protein Kinase cascades instigate a range of changes to enable eukaryotic cells to cope with particular insults. In Schizosaccharomyces pombe these responses include the transcription of specific gene sets and inhibition of entry into mitosis^ 1 , 2 . The S. pombe stress response pathway (SRP) also promotes commitment to mitosis in unperturbed cell cycles to allow cells to match their rate of division with nutrient availability^ 1 , 3 . The nature of this SRP function in cell cycle control is unknown. Entry into mitosis is controlled by mitosis-promoting factor (MPF; Cdc2/cyclin B) activity. Inhibitory phosphorylation of Cdc2 by Wee1 Kinase inactivates MPF until Cdc25 removes this phosphate to promote mitosis^ 4 . The balance between Wee1 and Cdc25 activities is influenced by the recruitment of Polo Kinase (Plo1) to the spindle pole body (SPB)^ 5 . The SPB component Cut12 mediates this recruitment^ 5 , 6 . Hyper-activating mutations in either cut12 or plo1 enable Cdc25-defective cells to enter mitosis^ 5 , 7 . The hyperactive cut12.s11 mutation suppresses cdc25.22 , as it promotes recruitment of active Plo1 to interphase SPBs^ 6 , 7 . Here we show that the SRP promotes phosphorylation of Plo1 on Ser 402. In unperturbed cell cycles, SRP-mediated phosphorylation of Ser 402 promotes Plo1 recruitment to SPBs and thus commitment to mitosis. Ser 402 phosphorylation also ensures efficient reinitiation of cell tip growth and cell division during recovery from particular stresses. Thus, phosphorylation of Plo1 Ser 402 not only enables SRP signalling to modulate the timing of mitotic commitment in response to nutrient status in unperturbed cycles, but also promotes the return to normal cell cycle control after stress.

  • Physical and functional interactions between Polo Kinase and the spindle pole component Cut12 regulate mitotic commitment in S. pombe
    Genes & development, 2003
    Co-Authors: Fiona H. Maciver, Kayoko Tanaka, Alasdair M. Robertson, Iain M Hagan
    Abstract:

    Commitment to mitosis is regulated by a protein Kinase complex called MPF. MPF is inhibited by Wee1-related Kinases and activated by Cdc25 phosphatase. MPF activation further boosts Cdc25 and represses Wee1. This feedback control probably involves Polo Kinase. A dominant cut12.s11 mutation in the Schizosaccharomyces pombe spindle pole body (SPB) component Cut12 both suppresses the conditional lethal mitotic commitment defect of cdc25.22 and promotes premature association of the S. pombe Polo Kinase, Plo1, with the SPB. We now show that Cut12 associated with Plo1 in two hybrid and immunoprecipitation assays. Plo1 function was required for recognition of the mitotic SPB by the phospho-specific antibody MPM-2. In vivo MPM-2 staining and in vitro Kinase assays established that the loss-of-function mutation, cut12.1, reduced mitotic activation of Plo1, whereas the gain-of-function mutation, cut12.s11, promoted higher levels of Plo1 activity than were normally seen in interphase. cut12.s11 could not promote mitotic commitment of cdc25.22 cells when Plo1 function was compromised. Expression of a constitutively active plo1 allele suppressed the mitotic commitment defect of cdc25.22. These data suggest that cut12.s11 suppresses cdc25.22 by promoting Plo1 activity. Furthermore, the delayed mitotic commitment of plo1.ts2 cells suggests that Plo1 is an integral part of the core controls that modulate MPF activation in S. pombe.