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1
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Mu Z, Zheng P, Liu S, Kang Y, Xie H. Plk4 regulates centriole duplication in the embryonic development of zebrafish. Dev Biol 2025; 517:148-156. [PMID: 39304174 DOI: 10.1016/j.ydbio.2024.09.006] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/03/2024] [Revised: 08/23/2024] [Accepted: 09/16/2024] [Indexed: 09/22/2024]
Abstract
PLK4 plays a crucial role in centriole duplication, which is essential for maintaining cellular processes such as cell division, cytoskeletal stability, and cilia formation. However, the mechanisms of PLK4 remain incompletely understood, especially in the embryonic development of vertebrate species. In this study, we observed that Plk4 dysfunction led to abnormal embryonic development in zebrafish, characterized by symptoms such as dark and wrinkled skin, microphthalmia, and body axis curvature. In plk4 mutants, defects in centriole duplication led to abnormal cell division, apoptosis, and ciliogenesis defects. Moreover, overexpression of plk4 in zebrafish embryos caused excessive centrosome amplification, disrupting embryonic gastrulation through abnormal cell division and ultimately resulting in embryonic lethality. Furthermore, we identified the "cryptic" polo box (CPB) domain, consisting of two PBs (PB1 and PB2), as the critical centrosome localization domain of Plk4. Surprisingly, overexpression of these two PB domains alone was sufficient to induce embryonic lethality. Additionally, we discovered a truncated form of CPB that localizes to the centrosome without causing defects in embryonic development. Our results demonstrate that Plk4 tightly controls centriole duplication, which is essential for early embryonic development in zebrafish.
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Affiliation(s)
- Zhiyu Mu
- Institute of Evolution & Marine Biodiversity, Ocean University of China, Qingdao, 266003, China
| | - Pengfei Zheng
- Institute of Evolution & Marine Biodiversity, Ocean University of China, Qingdao, 266003, China
| | - Shuangyu Liu
- Institute of Evolution & Marine Biodiversity, Ocean University of China, Qingdao, 266003, China
| | - Yunsi Kang
- Key Laboratory of Evolution and Marine Biodiversity of the Ministry of Education, Institute of Evolution and Marine Biodiversity, Ocean University of China, Qingdao, 266003, China
| | - Haibo Xie
- Institute of Evolution & Marine Biodiversity, Ocean University of China, Qingdao, 266003, China; Key Laboratory of Evolution and Marine Biodiversity of the Ministry of Education, Institute of Evolution and Marine Biodiversity, Ocean University of China, Qingdao, 266003, China.
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2
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Kiermaier E, Stötzel I, Schapfl MA, Villunger A. Amplified centrosomes-more than just a threat. EMBO Rep 2024; 25:4153-4167. [PMID: 39285247 PMCID: PMC11467336 DOI: 10.1038/s44319-024-00260-0] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/19/2024] [Revised: 07/05/2024] [Accepted: 08/28/2024] [Indexed: 09/19/2024] Open
Abstract
Centrosomes are major organizing components of the tubulin-based cytoskeleton. In recent years, we have gained extensive knowledge about their structure, biogenesis, and function from single cells, cell-cell interactions to tissue homeostasis, including their role in human diseases. Centrosome abnormalities are linked to, among others primary microcephaly, birth defects, ciliopathies, and tumorigenesis. Centrosome amplification, a state where two or more centrosomes are present in the G1 phase of the cell cycle, correlates in cancer with karyotype alterations, clinical aggressiveness, and lymph node metastasis. However, amplified centrosomes also appear in healthy tissues and, independent of their established role, in multi-ciliation. One example is the liver where hepatocytes carry amplified centrosomes owing to whole-genome duplication events during organogenesis. More recently, amplified centrosomes have been found in neuronal progenitors and several cell types of hematopoietic origin in which they enhance cellular effector functions. These findings suggest that extra centrosomes do not necessarily pose a risk for genome integrity and are harnessed for physiological processes. Here, we compare established and emerging 'non-canonical functions' of amplified centrosomes in cancerous and somatic cells and discuss their role in cellular physiology.
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Affiliation(s)
- Eva Kiermaier
- Life and Medical Sciences Institute, Immune and Tumor Biology, University of Bonn, Bonn, Germany.
| | - Isabel Stötzel
- Life and Medical Sciences Institute, Immune and Tumor Biology, University of Bonn, Bonn, Germany
| | - Marina A Schapfl
- Institute for Developmental Immunology, Biocenter, Medical University of Innsbruck, Innsbruck, Austria
| | - Andreas Villunger
- Institute for Developmental Immunology, Biocenter, Medical University of Innsbruck, Innsbruck, Austria.
- The Research Center for Molecular Medicine (CeMM) of the Austrian Academy of Sciences, Lazarettgasse 14, 1090, Vienna, Austria.
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3
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Lee JW, Mizuno K, Watanabe H, Lee IH, Tsumita T, Hida K, Yawaka Y, Kitagawa Y, Hasebe A, Iimura T, Kong SW. Enhanced phagocytosis associated with multinucleated microglia via Pyk2 inhibition in an acute β-amyloid infusion model. J Neuroinflammation 2024; 21:196. [PMID: 39107821 PMCID: PMC11301859 DOI: 10.1186/s12974-024-03192-7] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/07/2024] [Accepted: 07/31/2024] [Indexed: 08/10/2024] Open
Abstract
Multinucleated microglia have been observed in contexts associated with infection, inflammation, and aging. Though commonly linked to pathological conditions, the larger cell size of multinucleated microglia might enhance their phagocytic functions, potentially aiding in the clearance of brain debris and suggesting a reassessment of their pathological significance. To assess the phagocytic capacity of multinucleated microglia and its implications for brain debris clearance, we induced their formation by inhibiting Pyk2 activity using the pharmacological inhibitor PF-431396, which triggers cytokinesis regression. Multinucleated microglia demonstrate enhanced phagocytic function, as evidenced by their increased capacity to engulf β-amyloid (Aβ) oligomers. Concurrently, the phosphorylation of Pyk2, induced by Aβ peptide, was diminished upon treatment with a Pyk2 inhibitor (Pyk2-Inh, PF-431396). Furthermore, the increased expression of Lamp1, a lysosomal marker, with Pyk2-inh treatment, suggests an enhancement in proteolytic activity. In vivo, we generated an acute Alzheimer's disease (AD) model by infusing Aβ into the brains of Iba-1 EGFP transgenic (Tg) mice. The administration of the Pyk2-Inh led to an increased migration of microglia toward amyloid deposits in the brains of Iba-1 EGFP Tg mice, accompanied by morphological activation, suggesting a heightened affinity for Aβ. In human microglia, lipopolysaccharide (LPS)-induced inflammatory responses showed that inhibition of Pyk2 signaling significantly reduced the transcription and protein expression of pro-inflammatory markers. These results suggest that Pyk2 inhibition can modulate microglial functions, potentially reducing neuroinflammation and aiding in the clearance of neurodegenerative disease markers. This highlights Pyk2 as a promising target for therapeutic intervention in neurodegenerative diseases.
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Affiliation(s)
- Ji-Won Lee
- Microbiology, Department of Oral Pathobiological Science, Faculty and Graduate School of Dental Medicine, Hokkaido University, Kita13 Nishi7, Kita-Ku, Sapporo, 060-8586, Japan.
| | - Kaito Mizuno
- Microbiology, Department of Oral Pathobiological Science, Faculty and Graduate School of Dental Medicine, Hokkaido University, Kita13 Nishi7, Kita-Ku, Sapporo, 060-8586, Japan
- Dentistry for Children and Disabled Persons, Department of Oral Functional Science, Faculty of Dental Medicine, Hokkaido University, Kita13 Nishi7, Kita-Ku, Sapporo, 060-8586, Japan
| | - Haruhisa Watanabe
- Department of Pharmacology, Faculty and Graduate School of Dental Medicine, Hokkaido University, Kita13 Nishi7, Kita-Ku, Sapporo, 060-8586, Japan
- Oral Diagnosis and Medicine, Department of Oral Pathobiological Science, Faculty of Dental Medicine, Hokkaido University, Kita13 Nishi7, Kita-Ku, Sapporo, 060-8586, Japan
| | - In-Hee Lee
- Computational Health and Informatics Program, Boston Children's Hospital, Boston, MA, 02215, USA
| | - Takuya Tsumita
- Department of Vascular Biology and Molecular Pathology, Faculty and Graduate School of Dental Medicine, Hokkaido University, Kita13 Nishi7, Kita-Ku, Sapporo, 060-8586, Japan
| | - Kyoko Hida
- Department of Vascular Biology and Molecular Pathology, Faculty and Graduate School of Dental Medicine, Hokkaido University, Kita13 Nishi7, Kita-Ku, Sapporo, 060-8586, Japan
| | - Yasutaka Yawaka
- Dentistry for Children and Disabled Persons, Department of Oral Functional Science, Faculty of Dental Medicine, Hokkaido University, Kita13 Nishi7, Kita-Ku, Sapporo, 060-8586, Japan
| | - Yoshimasa Kitagawa
- Oral Diagnosis and Medicine, Department of Oral Pathobiological Science, Faculty of Dental Medicine, Hokkaido University, Kita13 Nishi7, Kita-Ku, Sapporo, 060-8586, Japan
| | - Akira Hasebe
- Microbiology, Department of Oral Pathobiological Science, Faculty and Graduate School of Dental Medicine, Hokkaido University, Kita13 Nishi7, Kita-Ku, Sapporo, 060-8586, Japan
| | - Tadahiro Iimura
- Department of Pharmacology, Faculty and Graduate School of Dental Medicine, Hokkaido University, Kita13 Nishi7, Kita-Ku, Sapporo, 060-8586, Japan
| | - Sek Won Kong
- Computational Health and Informatics Program, Boston Children's Hospital, Boston, MA, 02215, USA
- Department of Pediatrics, Harvard Medical School, Boston, MA, 02115, USA
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Perrier A, Guiglielmoni N, Naquin D, Gorrichon K, Thermes C, Lameiras S, Dammermann A, Schiffer PH, Brunstein M, Canman JC, Dumont J. Maternal inheritance of functional centrioles in two parthenogenetic nematodes. Nat Commun 2024; 15:6042. [PMID: 39025889 PMCID: PMC11258339 DOI: 10.1038/s41467-024-50427-5] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/14/2023] [Accepted: 07/09/2024] [Indexed: 07/20/2024] Open
Abstract
Centrioles are the core constituent of centrosomes, microtubule-organizing centers involved in directing mitotic spindle assembly and chromosome segregation in animal cells. In sexually reproducing species, centrioles degenerate during oogenesis and female meiosis is usually acentrosomal. Centrioles are retained during male meiosis and, in most species, are reintroduced with the sperm during fertilization, restoring centriole numbers in embryos. In contrast, the presence, origin, and function of centrioles in parthenogenetic species is unknown. We found that centrioles are maternally inherited in two species of asexual parthenogenetic nematodes and identified two different strategies for maternal inheritance evolved in the two species. In Rhabditophanes diutinus, centrioles organize the poles of the meiotic spindle and are inherited by both the polar body and embryo. In Disploscapter pachys, the two pairs of centrioles remain close together and are inherited by the embryo only. Our results suggest that maternally-inherited centrioles organize the embryonic spindle poles and act as a symmetry-breaking cue to induce embryo polarization. Thus, in these parthenogenetic nematodes, centrioles are maternally-inherited and functionally replace their sperm-inherited counterparts in sexually reproducing species.
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Affiliation(s)
- Aurélien Perrier
- Université Paris Cité, CNRS, Institut Jacques Monod, F-75013, Paris, France
| | - Nadège Guiglielmoni
- Worm∼lab, Institute for Zoology, University of Cologne, Cologne, NRW, Germany
| | - Delphine Naquin
- Université Paris-Saclay, CEA, CNRS, Institute for Integrative Biology of the Cell (I2BC), 91198, Gif-sur-Yvette, France
| | - Kevin Gorrichon
- Centre de Référence, d'Innovation, d'eXpertise et de transfert (CRefIX), US 039 CEA/INRIA/INSERM, Evry, France
- Centre National de Recherche en Génomique Humaine (CNRGH), Institut de Biologie François Jacob, Direction de la Recherche Fondamentale, CEA, Evry, France
| | - Claude Thermes
- Université Paris-Saclay, CEA, CNRS, Institute for Integrative Biology of the Cell (I2BC), 91198, Gif-sur-Yvette, France
| | - Sonia Lameiras
- Institut Curie, PSL University, ICGex Next-Generation Sequencing Platform, 75005, Paris, France
| | - Alexander Dammermann
- Max Perutz Labs, Vienna Biocenter Campus (VBC), 1030, Vienna, Austria
- University of Vienna, Center for Molecular Biology, Department of Microbiology, Immunobiology and Genetics, 1030, Vienna, Austria
| | - Philipp H Schiffer
- Worm∼lab, Institute for Zoology, University of Cologne, Cologne, NRW, Germany
| | - Maia Brunstein
- Institut Pasteur, Université Paris Cité, INSERM, Institut de l'Audition, F-75012, Paris, France
| | - Julie C Canman
- Columbia University Irving Medical Center; Department of Pathology and Cell Biology, New York, NY, 10032, USA
| | - Julien Dumont
- Université Paris Cité, CNRS, Institut Jacques Monod, F-75013, Paris, France.
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5
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Wang T, Fan G, Xia Y, Zou Y, Liu Y, Wang J, Hu Y, Teng J, Huang N, Chen J. Dual roles of CCDC102A in governing centrosome duplication and cohesion. Cell Rep 2024; 43:113696. [PMID: 38280197 DOI: 10.1016/j.celrep.2024.113696] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/13/2023] [Revised: 12/03/2023] [Accepted: 01/05/2024] [Indexed: 01/29/2024] Open
Abstract
In animal cells, the dysregulation of centrosome duplication and cohesion maintenance leads to abnormal spindle assembly and chromosomal instability, contributing to developmental disorders and tumorigenesis. However, the molecular mechanisms involved in maintaining accurate centrosome number control and tethering are not fully understood. Here, we identified coiled-coil domain-containing 102A (CCDC102A) as a centrosomal protein exhibiting a barrel-like structure in the proximal regions of parent centrioles, where it prevents centrosome overduplication by restricting interactions between Cep192 and Cep152 on centrosomes, thereby ensuring bipolar spindle formation. Additionally, CCDC102A regulates the centrosome linker by recruiting and binding C-Nap1; it is removed from the centrosome after Nek2A-mediated phosphorylation at the onset of mitosis. Overall, our results indicate that CCDC102A participates in controlling centrosome number and maintaining centrosome cohesion, suggesting that a well-tuned system regulates centrosome structure and function throughout the cell cycle.
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Affiliation(s)
- Tianning Wang
- Key Laboratory of Cell Proliferation and Differentiation of the Ministry of Education, College of Life Sciences, Peking University, Beijing 100871, China; Breast Disease Diagnosis and Treatment Center/Department of Thyroid Surgery, Central Hospital Affiliated to Shandong First Medical University, Jinan 250013, China; Research Center of Translational Medicine, Central Hospital Affiliated to Shandong First Medical University, Jinan 250013, China
| | - Guiliang Fan
- Key Laboratory of Cell Proliferation and Differentiation of the Ministry of Education, College of Life Sciences, Peking University, Beijing 100871, China
| | - Yuqing Xia
- Key Laboratory of Cell Proliferation and Differentiation of the Ministry of Education, College of Life Sciences, Peking University, Beijing 100871, China
| | - Yuhong Zou
- Key Laboratory of Cell Proliferation and Differentiation of the Ministry of Education, College of Life Sciences, Peking University, Beijing 100871, China
| | - Yunjie Liu
- Key Laboratory of Cell Proliferation and Differentiation of the Ministry of Education, College of Life Sciences, Peking University, Beijing 100871, China
| | - Jiaxin Wang
- Department of Physiology and Pathophysiology, School of Basic Medical Sciences, Health Science Center, Xi'an Jiaotong University, Xi'an 710061, China; Institute of Neuroscience, Translational Medicine Institute, Health Science Center, Xi'an Jiaotong University, Xi'an 710061, China
| | - Yingchun Hu
- Key Laboratory of Cell Proliferation and Differentiation of the Ministry of Education, College of Life Sciences, Peking University, Beijing 100871, China
| | - Junlin Teng
- Key Laboratory of Cell Proliferation and Differentiation of the Ministry of Education, College of Life Sciences, Peking University, Beijing 100871, China.
| | - Ning Huang
- Department of Physiology and Pathophysiology, School of Basic Medical Sciences, Health Science Center, Xi'an Jiaotong University, Xi'an 710061, China; Institute of Neuroscience, Translational Medicine Institute, Health Science Center, Xi'an Jiaotong University, Xi'an 710061, China.
| | - Jianguo Chen
- Key Laboratory of Cell Proliferation and Differentiation of the Ministry of Education, College of Life Sciences, Peking University, Beijing 100871, China; Center for Quantitative Biology, Academy for Advanced Interdisciplinary Studies, Peking University, Beijing 100871, China.
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6
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Sullenberger C, Kong D, Avazpour P, Luvsanjav D, Loncarek J. Centrosomal organization of Cep152 provides flexibility in Plk4 and procentriole positioning. J Cell Biol 2023; 222:e202301092. [PMID: 37707473 PMCID: PMC10501443 DOI: 10.1083/jcb.202301092] [Citation(s) in RCA: 5] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/20/2023] [Revised: 06/25/2023] [Accepted: 08/17/2023] [Indexed: 09/15/2023] Open
Abstract
Centriole duplication is a high-fidelity process driven by Polo-like kinase 4 (Plk4) and a few conserved initiators. Dissecting how Plk4 and its receptors organize within centrosomes is critical to understand the centriole duplication process and biochemical and architectural differences between centrosomes of different species. Here, at nanoscale resolution, we dissect centrosomal localization of Plk4 in G1 and S phase in its catalytically active and inhibited state during centriole duplication and amplification. We build a precise distribution map of Plk4 and its receptor Cep152, as well as Cep44, Cep192, and Cep152-anchoring factors Cep57 and Cep63. We find that Cep57, Cep63, Cep44, and Cep192 localize in ninefold symmetry. However, during centriole maturation, Cep152, which we suggest is the major Plk4 receptor, develops a more complex pattern. We propose that the molecular arrangement of Cep152 creates flexibility for Plk4 and procentriole placement during centriole initiation. As a result, procentrioles form at variable positions in relation to the mother centriole microtubule triplets.
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Affiliation(s)
- Catherine Sullenberger
- Cancer Innovation Laboratory, National Institutes of Health, National Cancer Institute, Center for Cancer Research, Frederick, MD, USA
| | - Dong Kong
- Cancer Innovation Laboratory, National Institutes of Health, National Cancer Institute, Center for Cancer Research, Frederick, MD, USA
| | - Pegah Avazpour
- Cancer Innovation Laboratory, National Institutes of Health, National Cancer Institute, Center for Cancer Research, Frederick, MD, USA
| | - Delgermaa Luvsanjav
- Cancer Innovation Laboratory, National Institutes of Health, National Cancer Institute, Center for Cancer Research, Frederick, MD, USA
| | - Jadranka Loncarek
- Cancer Innovation Laboratory, National Institutes of Health, National Cancer Institute, Center for Cancer Research, Frederick, MD, USA
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7
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Dwivedi D, Harry D, Meraldi P. Mild replication stress causes premature centriole disengagement via a sub-critical Plk1 activity under the control of ATR-Chk1. Nat Commun 2023; 14:6088. [PMID: 37773176 PMCID: PMC10541884 DOI: 10.1038/s41467-023-41753-1] [Citation(s) in RCA: 6] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/07/2022] [Accepted: 09/18/2023] [Indexed: 10/01/2023] Open
Abstract
A tight synchrony between the DNA and centrosome cycle is essential for genomic integrity. Centriole disengagement, which licenses centrosomes for duplication, occurs normally during mitotic exit. We recently demonstrated that mild DNA replication stress typically seen in cancer cells causes premature centriole disengagement in untransformed mitotic human cells, leading to transient multipolar spindles that favour chromosome missegregation. How mild replication stress accelerates the centrosome cycle at the molecular level remained, however, unclear. Using ultrastructure expansion microscopy, we show that mild replication stress induces premature centriole disengagement already in G2 via the ATR-Chk1 axis of the DNA damage repair pathway. This results in a sub-critical Plk1 kinase activity that primes the pericentriolar matrix for Separase-dependent disassembly but is insufficient for rapid mitotic entry, causing premature centriole disengagement in G2. We postulate that the differential requirement of Plk1 activity for the DNA and centrosome cycles explains how mild replication stress disrupts the synchrony between both processes and contributes to genomic instability.
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Affiliation(s)
- Devashish Dwivedi
- Department of Cell Physiology and Metabolism, Faculty of Medicine, University of Geneva, 1211 Geneva 4, Geneva, Switzerland
- Translational Research Centre in Onco-hematology, Faculty of Medicine, University of Geneva, 1211 Geneva 4, Geneva, Switzerland
| | - Daniela Harry
- Department of Cell Physiology and Metabolism, Faculty of Medicine, University of Geneva, 1211 Geneva 4, Geneva, Switzerland
- Translational Research Centre in Onco-hematology, Faculty of Medicine, University of Geneva, 1211 Geneva 4, Geneva, Switzerland
| | - Patrick Meraldi
- Department of Cell Physiology and Metabolism, Faculty of Medicine, University of Geneva, 1211 Geneva 4, Geneva, Switzerland.
- Translational Research Centre in Onco-hematology, Faculty of Medicine, University of Geneva, 1211 Geneva 4, Geneva, Switzerland.
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8
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Haynes BM, Cunningham K, Shekhar MPV. RAD6 inhibition enhances paclitaxel sensitivity of triple negative breast cancer cells by aggravating mitotic spindle damage. BMC Cancer 2022; 22:1073. [PMID: 36258187 PMCID: PMC9578210 DOI: 10.1186/s12885-022-10119-z] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/09/2022] [Accepted: 09/22/2022] [Indexed: 11/28/2022] Open
Abstract
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. Supplementary Information The online version contains supplementary material available at 10.1186/s12885-022-10119-z.
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Affiliation(s)
- Brittany M Haynes
- Karmanos Cancer Institute, 4100 John R Street, Detroit, MI, 48201, USA.,Department of Oncology, Wayne State University School of Medicine, 421 E. Canfield Avenue, Detroit, MI, 48201, USA.,Present address: Office of Policy Communications, and Education, National Center for Advancing Translational Sciences, Besthesda, USA
| | - Kristen Cunningham
- Karmanos Cancer Institute, 4100 John R Street, Detroit, MI, 48201, USA.,Department of Oncology, Wayne State University School of Medicine, 421 E. Canfield Avenue, Detroit, MI, 48201, USA
| | - Malathy P V Shekhar
- Karmanos Cancer Institute, 4100 John R Street, Detroit, MI, 48201, USA. .,Department of Oncology, Wayne State University School of Medicine, 421 E. Canfield Avenue, Detroit, MI, 48201, USA. .,Department of Pathology, Wayne State University School of Medicine, 421 E. Canfield Avenue, Detroit, MI, 48201, USA.
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9
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Hoffmann I. Role of Polo-like Kinases Plk1 and Plk4 in the Initiation of Centriole Duplication-Impact on Cancer. Cells 2022; 11:786. [PMID: 35269408 PMCID: PMC8908989 DOI: 10.3390/cells11050786] [Citation(s) in RCA: 11] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/28/2021] [Revised: 02/16/2022] [Accepted: 02/22/2022] [Indexed: 02/04/2023] Open
Abstract
Centrosomes nucleate and anchor microtubules and therefore play major roles in spindle formation and chromosome segregation during mitosis. Duplication of the centrosome occurs, similar to DNA, only once during the cell cycle. Aberration of the centrosome number is common in human tumors. At the core of centriole duplication is the conserved polo-like kinase 4, Plk4, and two structural proteins, STIL and Sas-6. In this review, I summarize and discuss developments in our understanding of the first steps of centriole duplication and their regulation.
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Affiliation(s)
- Ingrid Hoffmann
- F045, Cell Cycle Control and Carcinogenesis, Im Neuenheimer Feld 242, 69115 Heidelberg, Germany
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10
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Mehta DS, Zein-Sabatto H, Ryder PV, Lee J, Lerit DA. Drosophila centrocortin is dispensable for centriole duplication but contributes to centrosome separation. G3 GENES|GENOMES|GENETICS 2022; 12:6481552. [PMID: 35100335 PMCID: PMC9210305 DOI: 10.1093/g3journal/jkab434] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 09/03/2021] [Accepted: 12/09/2021] [Indexed: 11/15/2022]
Abstract
Centrosomes are microtubule-organizing centers that duplicate exactly once to organize the bipolar mitotic spindle required for error-free mitosis. Prior work indicated that Drosophila centrocortin (cen) is required for normal centrosome separation, although a role in centriole duplication was not closely examined. Through time-lapse recordings of rapid syncytial divisions, we monitored centriole duplication and the kinetics of centrosome separation in control vs cen null embryos. Our data suggest that although cen is dispensable for centriole duplication, it contributes to centrosome separation.
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Affiliation(s)
- Dipen S Mehta
- College of Science and Mathematics, Augusta University, Augusta, GA 30912, USA
| | - Hala Zein-Sabatto
- Department of Cell Biology, Emory University School of Medicine, Atlanta, GA 30322, USA
| | - Pearl V Ryder
- Department of Cell Biology, Emory University School of Medicine, Atlanta, GA 30322, USA
- Wandrer, Atlanta, GA 30340, USA
| | - Jina Lee
- Emory College of Arts and Sciences, Emory University, Atlanta, GA 30322, USA
| | - Dorothy A Lerit
- Department of Cell Biology, Emory University School of Medicine, Atlanta, GA 30322, USA
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11
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Stötzel I, Kiermaier E. The central role of the centrosome. eLife 2022; 11:84659. [PMID: 36508246 PMCID: PMC9744438 DOI: 10.7554/elife.84659] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/14/2022] Open
Abstract
The centrosome decides which branch extending from the body of microglia will successfully engulf and clear away dead neurons.
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Affiliation(s)
- Isabel Stötzel
- Life and Medical Sciences (LIMES) Institute, Immune and Tumor Biology, University of BonnBonnGermany
| | - Eva Kiermaier
- Life and Medical Sciences (LIMES) Institute, Immune and Tumor Biology, University of BonnBonnGermany
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12
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Shin B, Kim MS, Lee Y, Jung GI, Rhee K. Generation and Fates of Supernumerary Centrioles in Dividing Cells. Mol Cells 2021; 44:699-705. [PMID: 34711687 PMCID: PMC8560585 DOI: 10.14348/molcells.2021.0220] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/23/2021] [Revised: 09/25/2021] [Accepted: 09/30/2021] [Indexed: 11/27/2022] Open
Abstract
The centrosome is a subcellular organelle from which a cilium assembles. Since centrosomes function as spindle poles during mitosis, they have to be present as a pair in a cell. How the correct number of centrosomes is maintained in a cell has been a major issue in the fields of cell cycle and cancer biology. Centrioles, the core of centrosomes, assemble and segregate in close connection to the cell cycle. Abnormalities in centriole numbers are attributed to decoupling from cell cycle regulation. Interestingly, supernumerary centrioles are commonly observed in cancer cells. In this review, we discuss how supernumerary centrioles are generated in diverse cellular conditions. We also discuss how the cells cope with supernumerary centrioles during the cell cycle.
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Affiliation(s)
- Byungho Shin
- Department of Biological Sciences, Seoul National University, Seoul 08826, Korea
| | - Myung Se Kim
- Department of Biological Sciences, Seoul National University, Seoul 08826, Korea
| | - Yejoo Lee
- Department of Biological Sciences, Seoul National University, Seoul 08826, Korea
| | - Gee In Jung
- Department of Biological Sciences, Seoul National University, Seoul 08826, Korea
| | - Kunsoo Rhee
- Department of Biological Sciences, Seoul National University, Seoul 08826, Korea
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13
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Jung GI, Rhee K. Triple deletion of TP53, PCNT, and CEP215 promotes centriole amplification in the M phase. Cell Cycle 2021; 20:1500-1517. [PMID: 34233584 DOI: 10.1080/15384101.2021.1950386] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/19/2023] Open
Abstract
Supernumerary centrioles are frequently observed in diverse types of cancer cells. In this study, we investigated the mechanism underlying the generation of supernumerary centrioles during the M phase. We generated the TP53;PCNT;CEP215 triple knockout (KO) cells and determined the configurations of the centriole during the cell cycle. The triple KO cells exhibited a precocious separation of centrioles and unscheduled centriole assembly in the M phase. Supernumerary centrioles in the triple KO cells were present throughout the cell cycle; however, among all the centrioles, only two maintained an intact composition, including CEP135, CEP192, CEP295 and CEP152. Intact centrioles were formed during the S phase and the rest of the centrioles may be generated during the M phase. M-phase-assembled centrioles lacked the ability to organize microtubules in the interphase; however, a fraction of them may acquire pericentriolar material to organize microtubules during the M phase. Taken together, our work reveals the heterogeneity of the supernumerary centrioles in the triple KO cells. .
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Affiliation(s)
- Gee In Jung
- Department of Biological Sciences, Seoul National University, Seoul, Korea
| | - Kunsoo Rhee
- Department of Biological Sciences, Seoul National University, Seoul, Korea
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14
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Sullenberger C, Vasquez-Limeta A, Kong D, Loncarek J. With Age Comes Maturity: Biochemical and Structural Transformation of a Human Centriole in the Making. Cells 2020; 9:cells9061429. [PMID: 32526902 PMCID: PMC7349492 DOI: 10.3390/cells9061429] [Citation(s) in RCA: 28] [Impact Index Per Article: 5.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/12/2020] [Revised: 05/29/2020] [Accepted: 06/04/2020] [Indexed: 12/14/2022] Open
Abstract
Centrioles are microtubule-based cellular structures present in most human cells that build centrosomes and cilia. Proliferating cells have only two centrosomes and this number is stringently maintained through the temporally and spatially controlled processes of centriole assembly and segregation. The assembly of new centrioles begins in early S phase and ends in the third G1 phase from their initiation. This lengthy process of centriole assembly from their initiation to their maturation is characterized by numerous structural and still poorly understood biochemical changes, which occur in synchrony with the progression of cells through three consecutive cell cycles. As a result, proliferating cells contain three structurally, biochemically, and functionally distinct types of centrioles: procentrioles, daughter centrioles, and mother centrioles. This age difference is critical for proper centrosome and cilia function. Here we discuss the centriole assembly process as it occurs in somatic cycling human cells with a focus on the structural, biochemical, and functional characteristics of centrioles of different ages.
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15
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Centrosome reduction in newly-generated tetraploid cancer cells obtained by separase depletion. Sci Rep 2020; 10:9152. [PMID: 32499568 PMCID: PMC7272426 DOI: 10.1038/s41598-020-65975-1] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/29/2019] [Accepted: 04/17/2020] [Indexed: 12/29/2022] Open
Abstract
Tetraploidy, a common feature in cancer, results in the presence of extra centrosomes, which has been associated with chromosome instability (CIN) and aneuploidy. Deregulation in the number of centrosomes triggers tumorigenesis. However, how supernumerary centrosomes evolve during the emergence of tetraploid cells remains yet to be elucidated. Here, generating tetraploid isogenic clones in colorectal cancer and in non-transformed cells, we show that near-tetraploid clones exhibit a significant increase in the number of centrosomes. Moreover, we find that centrosome area in near-tetraploids is twice as large as in near-diploids. To evaluate whether centrosome clustering was occurring, we next analysed the number of centrioles revealing centriole amplification. Notwithstanding, more than half of the near-tetraploids maintained in culture do not present centrosome aberrations. To test whether cells progressively lost centrioles after becoming near-tetraploid, we transiently transfected diploid cells with siRNA against ESPL1/Separase, a protease responsible for triggering anaphase, to generate newly near-tetraploid cells. Finally, using this model, we assessed the number of centrioles at different time-points after tetraploidization finding that near-tetraploids rapidly lose centrosomes over time. Taken together, these data demonstrate that although most cells reduce supernumerary centrosomes after tetraploidization, a small fraction retains extra centrioles, potentially resulting in CIN.
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16
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Shabo I, Svanvik J, Lindström A, Lechertier T, Trabulo S, Hulit J, Sparey T, Pawelek J. Roles of cell fusion, hybridization and polyploid cell formation in cancer metastasis. World J Clin Oncol 2020; 11:121-135. [PMID: 32257843 PMCID: PMC7103524 DOI: 10.5306/wjco.v11.i3.121] [Citation(s) in RCA: 52] [Impact Index Per Article: 10.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/20/2019] [Revised: 01/02/2020] [Accepted: 03/01/2020] [Indexed: 02/06/2023] Open
Abstract
Cell-cell fusion is a normal biological process playing essential roles in organ formation and tissue differentiation, repair and regeneration. Through cell fusion somatic cells undergo rapid nuclear reprogramming and epigenetic modifications to form hybrid cells with new genetic and phenotypic properties at a rate exceeding that achievable by random mutations. Factors that stimulate cell fusion are inflammation and hypoxia. Fusion of cancer cells with non-neoplastic cells facilitates several malignancy-related cell phenotypes, e.g., reprogramming of somatic cell into induced pluripotent stem cells and epithelial to mesenchymal transition. There is now considerable in vitro, in vivo and clinical evidence that fusion of cancer cells with motile leucocytes such as macrophages plays a major role in cancer metastasis. Of the many changes in cancer cells after hybridizing with leucocytes, it is notable that hybrids acquire resistance to chemo- and radiation therapy. One phenomenon that has been largely overlooked yet plays a role in these processes is polyploidization. Regardless of the mechanism of polyploid cell formation, it happens in response to genotoxic stresses and enhances a cancer cell’s ability to survive. Here we summarize the recent progress in research of cell fusion and with a focus on an important role for polyploid cells in cancer metastasis. In addition, we discuss the clinical evidence and the importance of cell fusion and polyploidization in solid tumors.
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Affiliation(s)
- Ivan Shabo
- Endocrine and Sarcoma Surgery Unit, Department of Molecular Medicine and Surgery, Karolinska Institute, Stockholm SE 171 77, Sweden
- Patient Area of Breast Cancer, Sarcoma and Endocrine Tumours, Theme Cancer, Karolinska University Hospital, Stockholm SE 171 76, Sweden
| | - Joar Svanvik
- The Transplant Institute, Sahlgrenska University Hospital, Gothenburg SE 413 45, Sweden
- Division of Surgery, Department of Biomedical and Clinical Sciences, Faculty of Medicine and Health Sciences, Linköping University, Linköping SE 581 83, Sweden
| | - Annelie Lindström
- Division of Cell Biology, Department of Biomedical and Clinical Sciences, Faculty of Medicine and Health Sciences, Linköping University, Linköping SE 581 85, Sweden
| | - Tanguy Lechertier
- Novintum Bioscience Ltd, London Bioscience Innovation Centre, London NW1 0NH, United Kingdom
| | - Sara Trabulo
- Novintum Bioscience Ltd, London Bioscience Innovation Centre, London NW1 0NH, United Kingdom
| | - James Hulit
- Novintum Bioscience Ltd, London Bioscience Innovation Centre, London NW1 0NH, United Kingdom
| | - Tim Sparey
- Novintum Bioscience Ltd, London Bioscience Innovation Centre, London NW1 0NH, United Kingdom
| | - John Pawelek
- Department of Dermatology and the Yale Cancer Center, Yale University School of Medicine, New Haven, CT 06520, United States
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17
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Riparbelli MG, Persico V, Dallai R, Callaini G. Centrioles and Ciliary Structures during Male Gametogenesis in Hexapoda: Discovery of New Models. Cells 2020; 9:E744. [PMID: 32197383 PMCID: PMC7140630 DOI: 10.3390/cells9030744] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/10/2020] [Revised: 03/08/2020] [Accepted: 03/10/2020] [Indexed: 12/12/2022] Open
Abstract
Centrioles are-widely conserved barrel-shaped organelles present in most organisms. They are indirectly involved in the organization of the cytoplasmic microtubules both in interphase and during the cell division by recruiting the molecules needed for microtubule nucleation. Moreover, the centrioles are required to assemble cilia and flagella by the direct elongation of their microtubule wall. Due to the importance of the cytoplasmic microtubules in several aspects of the cell life, any defect in centriole structure can lead to cell abnormalities that in humans may result in significant diseases. Many aspects of the centriole dynamics and function have been clarified in the last years, but little attention has been paid to the exceptions in centriole structure that occasionally appeared within the animal kingdom. Here, we focused our attention on non-canonical aspects of centriole architecture within the Hexapoda. The Hexapoda is one of the major animal groups and represents a good laboratory in which to examine the evolution and the organization of the centrioles. Although these findings represent obvious exceptions to the established rules of centriole organization, they may contribute to advance our understanding of the formation and the function of these organelles.
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Affiliation(s)
- Maria Giovanna Riparbelli
- Department of Life Sciences, University of Siena, Via Aldo Moro 2, 53100 Siena, Italy; (M.G.R.); (V.P.); (R.D.)
| | - Veronica Persico
- Department of Life Sciences, University of Siena, Via Aldo Moro 2, 53100 Siena, Italy; (M.G.R.); (V.P.); (R.D.)
| | - Romano Dallai
- Department of Life Sciences, University of Siena, Via Aldo Moro 2, 53100 Siena, Italy; (M.G.R.); (V.P.); (R.D.)
| | - Giuliano Callaini
- Department of Life Sciences, University of Siena, Via Aldo Moro 2, 53100 Siena, Italy; (M.G.R.); (V.P.); (R.D.)
- Department of Medical Biotechnologies, University of Siena, Via Aldo Moro 2, 53100 Siena, Italy
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18
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Mutation in DNA Polymerase Beta Causes Spontaneous Chromosomal Instability and Inflammation-Associated Carcinogenesis in Mice. Cancers (Basel) 2019; 11:cancers11081160. [PMID: 31412651 PMCID: PMC6721533 DOI: 10.3390/cancers11081160] [Citation(s) in RCA: 16] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/14/2019] [Revised: 08/01/2019] [Accepted: 08/08/2019] [Indexed: 12/15/2022] Open
Abstract
DNA polymerase beta (Pol β) is a key enzyme in the base excision repair (BER) pathway. Pol β is mutated in approximately 40% of human tumors in small-scale studies. The 5´-deoxyribose-5-phosphate (dRP) lyase domain of Pol β is responsible for DNA end tailoring to remove the 5’ phosphate group. We previously reported that the dRP lyase activity of Pol β is critical to maintain DNA replication fork stability and prevent cellular transformation. In this study, we tested the hypothesis that the human gastric cancer associated variant of Pol β (L22P) has the ability to promote spontaneous chromosomal instability and carcinogenesis in mice. We constructed a Pol β L22P conditional knock-in mouse model and found that L22P enhances hyperproliferation and DNA double strand breaks (DSBs) in stomach cells. Moreover, mouse embryonic fibroblasts (MEFs) derived from L22P mice frequently induce abnormal numbers of chromosomes and centrosome amplification, leading to chromosome segregation errors. Importantly, L22P mice exhibit chronic inflammation accompanied by stomach tumors. These data demonstrate that the human cancer-associated variant of Pol β can contribute to chromosomal instability and cancer development.
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19
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Abstract
The centriole is an ancient microtubule-based organelle with a conserved nine-fold symmetry. Centrioles form the core of centrosomes, which organize the interphase microtubule cytoskeleton of most animal cells and form the poles of the mitotic spindle. Centrioles can also be modified to form basal bodies, which template the formation of cilia and play central roles in cellular signaling, fluid movement, and locomotion. In this review, we discuss developments in our understanding of the biogenesis of centrioles and cilia and the regulatory controls that govern their structure and number. We also discuss how defects in these processes contribute to a spectrum of human diseases and how new technologies have expanded our understanding of centriole and cilium biology, revealing exciting avenues for future exploration.
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Affiliation(s)
- David K Breslow
- Department of Molecular, Cellular and Developmental Biology, Yale University, New Haven, Connecticut 06511, USA;
| | - Andrew J Holland
- Department of Molecular Biology and Genetics, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA;
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20
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Loncarek J, Bettencourt-Dias M. Building the right centriole for each cell type. J Cell Biol 2017; 217:823-835. [PMID: 29284667 PMCID: PMC5839779 DOI: 10.1083/jcb.201704093] [Citation(s) in RCA: 66] [Impact Index Per Article: 8.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/11/2017] [Revised: 09/14/2017] [Accepted: 11/27/2017] [Indexed: 12/22/2022] Open
Abstract
Loncarek and Bettencourt-Dias review molecular mechanisms of centriole biogenesis amongst different organisms and cell types. The centriole is a multifunctional structure that organizes centrosomes and cilia and is important for cell signaling, cell cycle progression, polarity, and motility. Defects in centriole number and structure are associated with human diseases including cancer and ciliopathies. Discovery of the centriole dates back to the 19th century. However, recent advances in genetic and biochemical tools, development of high-resolution microscopy, and identification of centriole components have accelerated our understanding of its assembly, function, evolution, and its role in human disease. The centriole is an evolutionarily conserved structure built from highly conserved proteins and is present in all branches of the eukaryotic tree of life. However, centriole number, size, and organization varies among different organisms and even cell types within a single organism, reflecting its cell type–specialized functions. In this review, we provide an overview of our current understanding of centriole biogenesis and how variations around the same theme generate alternatives for centriole formation and function.
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Affiliation(s)
- Jadranka Loncarek
- Cell Cycle Regulation Lab, Gulbenkian Institute of Science, Oeiras, Portugal
| | - Mónica Bettencourt-Dias
- Laboratory of Protein Dynamics and Signaling, National Institutes of Health/Center for Cancer Research/National Cancer Institute-Frederick, Frederick, MD
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21
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The PLK4-STIL-SAS-6 module at the core of centriole duplication. Biochem Soc Trans 2017; 44:1253-1263. [PMID: 27911707 PMCID: PMC5095913 DOI: 10.1042/bst20160116] [Citation(s) in RCA: 103] [Impact Index Per Article: 12.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/13/2016] [Revised: 06/09/2016] [Accepted: 06/24/2016] [Indexed: 11/17/2022]
Abstract
Centrioles are microtubule-based core components of centrosomes and cilia. They are duplicated exactly once during S-phase progression. Central to formation of each new (daughter) centriole is the formation of a nine-fold symmetrical cartwheel structure onto which microtubule triplets are deposited. In recent years, a module comprising the protein kinase polo-like kinase 4 (PLK4) and the two proteins STIL and SAS-6 have been shown to stay at the core of centriole duplication. Depletion of any one of these three proteins blocks centriole duplication and, conversely, overexpression causes centriole amplification. In this short review article, we summarize recent insights into how PLK4, STIL and SAS-6 co-operate in space and time to form a new centriole. These advances begin to shed light on the very first steps of centriole biogenesis.
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22
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Novak ZA, Wainman A, Gartenmann L, Raff JW. Cdk1 Phosphorylates Drosophila Sas-4 to Recruit Polo to Daughter Centrioles and Convert Them to Centrosomes. Dev Cell 2017; 37:545-57. [PMID: 27326932 PMCID: PMC4918730 DOI: 10.1016/j.devcel.2016.05.022] [Citation(s) in RCA: 46] [Impact Index Per Article: 5.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/26/2016] [Revised: 05/04/2016] [Accepted: 05/23/2016] [Indexed: 02/07/2023]
Abstract
Centrosomes and cilia are organized by a centriole pair comprising an older mother and a younger daughter. Centriole numbers are tightly regulated, and daughter centrioles (which assemble in S phase) cannot themselves duplicate or organize centrosomes until they have passed through mitosis. It is unclear how this mitotic “centriole conversion” is regulated, but it requires Plk1/Polo kinase. Here we show that in flies, Cdk1 phosphorylates the conserved centriole protein Sas-4 during mitosis. This creates a Polo-docking site that helps recruit Polo to daughter centrioles and is required for the subsequent recruitment of Asterless (Asl), a protein essential for centriole duplication and mitotic centrosome assembly. Point mutations in Sas-4 that prevent Cdk1 phosphorylation or Polo docking do not block centriole disengagement during mitosis, but block efficient centriole conversion and lead to embryonic lethality. These observations can explain why daughter centrioles have to pass through mitosis before they can duplicate and organize a centrosome.
Cdk1 phosphorylates Sas-4 to initiate Polo/Plk1 recruitment to daughter centrioles Polo recruitment promotes Asterless (Asl) incorporation into daughter centrioles Asl incorporation licenses new centrioles to duplicate and organize centrosomes These observations help explain why centriole conversion is tied to mitosis
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Affiliation(s)
- Zsofia A Novak
- Sir William Dunn School of Pathology, University of Oxford, South Parks Road, Oxford OX1 3RE, UK
| | - Alan Wainman
- Sir William Dunn School of Pathology, University of Oxford, South Parks Road, Oxford OX1 3RE, UK
| | - Lisa Gartenmann
- Sir William Dunn School of Pathology, University of Oxford, South Parks Road, Oxford OX1 3RE, UK
| | - Jordan W Raff
- Sir William Dunn School of Pathology, University of Oxford, South Parks Road, Oxford OX1 3RE, UK.
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23
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Kumar R. Separase: Function Beyond Cohesion Cleavage and an Emerging Oncogene. J Cell Biochem 2017; 118:1283-1299. [PMID: 27966791 DOI: 10.1002/jcb.25835] [Citation(s) in RCA: 16] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/11/2016] [Accepted: 12/12/2016] [Indexed: 12/22/2022]
Abstract
Proper and timely segregation of genetic endowment is necessary for survival and perpetuation of every species. Mis-segregation of chromosomes and resulting aneuploidy leads to genetic instability, which can jeopardize the survival of an individual or population as a whole. Abnormality with segregation of genetic contents has been associated with several medical consequences including cancer, sterility, mental retardation, spontaneous abortion, miscarriages, and other birth related defects. Separase, by irreversible cleavage of cohesin complex subunit, paves the way for metaphase/anaphase transition during the cell cycle. Both over or reduced expression and altered level of separase have been associated with several medical consequences including cancer, as a result separase now emerges as an important oncogene and potential molecular target for medical intervenes. Recently, separase is also found to be essential in separation and duplication of centrioles. Here, I review the role of separase in mitosis, meiosis, non-canonical roles of separase, separase regulation, as a regulator of centriole disengagement, nonproteolytic roles, diverse substrates, structural insights, and association of separase with cancer. At the ends, I proposed a model which showed that separase is active throughout the cell cycle and there is a mere increase in separase activity during metaphase contrary to the common believes that separase is inactive throughout cell cycle except for metaphase. J. Cell. Biochem. 118: 1283-1299, 2017. © 2016 Wiley Periodicals, Inc.
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Affiliation(s)
- Ravinder Kumar
- Department of Biosciences and Bioengineering, Indian Institute of Technology Bombay, Powai, Mumbai, 400 076, Maharashtra, India
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24
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Centrosome aberrations in human mammary epithelial cells driven by cooperative interactions between p16INK4a deficiency and telomere-dependent genotoxic stress. Oncotarget 2016; 6:28238-56. [PMID: 26318587 PMCID: PMC4695057 DOI: 10.18632/oncotarget.4958] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/07/2014] [Accepted: 07/07/2015] [Indexed: 01/11/2023] Open
Abstract
Virtually all human cancers display chromosome instability (CIN), a condition in which chromosomes are gained or lost at a high rate. CIN occurs early in cancer development where it may undermine the advance of the neoplastic disease. With the aim of establishing the mechanisms underlying CIN in cancer, we investigated possible links between telomere-dysfunction and centrosome defects, which were seen to coincide in early in breast carcinogenesis using human mammary epithelial cells (HMECs). In this study, we show that TP53 proficient vHMECs cells develop centrosome aberrations when telomere-dysfunction genotoxic stress is produced in the presence of a defective p16INK4a setting and in parallel with an activation of the DNA damage checkpoint response. These aberrations consist of the accumulation of centrosomes in polyploid vHMECs, plus centriole overduplication in both diploid and polyploid cells, thus reflecting that distinct mechanisms underlie the generation of centrosome aberrations in vHMECs. Transduction of vHMEC with hTERT, which rescued the telomere dysfunction phenotype and consequently reduced DNA damage checkpoint activation, led to a progressive reduction of centrosome aberrations with cell culture, both in diploid and in polyploid vHMECs. Radiation-induced DNA damage also raised centrosome aberrations in vHMEC-hTERT. Collectively, our results, using vHMECs define a model where p16INK4a deficiency along with short dysfunctional telomeres cooperatively engenders centrosome abnormalities before p53 function is compromised.
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25
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Kim M, O'Rourke BP, Soni RK, Jallepalli PV, Hendrickson RC, Tsou MFB. Promotion and Suppression of Centriole Duplication Are Catalytically Coupled through PLK4 to Ensure Centriole Homeostasis. Cell Rep 2016; 16:1195-1203. [PMID: 27425613 PMCID: PMC4972634 DOI: 10.1016/j.celrep.2016.06.069] [Citation(s) in RCA: 20] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/11/2016] [Revised: 05/19/2016] [Accepted: 06/15/2016] [Indexed: 11/26/2022] Open
Abstract
PLK4 is the major kinase driving centriole duplication. Duplication occurs only once per cell cycle, forming one new (or daughter) centriole that is tightly engaged to the preexisting (or mother) centriole. Centriole engagement is known to block the reduplication of mother centrioles, but the molecular identity responsible for the block remains unclear. Here, we show that the centriolar cartwheel, the geometric scaffold for centriole assembly, forms the identity of daughter centrioles essential for the block, ceasing further duplication of the mother centriole to which it is engaged. To ensure a steady block, we found that the cartwheel requires constant maintenance by PLK4 through phosphorylation of the same substrate that drives centriole assembly, revealing a parsimonious control in which “assembly” and “block for new assembly” are linked through the same catalytic reaction to achieve homeostasis. Our results support a recently deduced model that the cartwheel-bound PLK4 directly suppresses centriole reduplication.
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Affiliation(s)
- Minhee Kim
- Cell Biology Program, Memorial Sloan-Kettering Cancer Center, New York, NY 10065, USA; Weill Cornell Graduate School of Medical Sciences, Cornell University, New York, NY 10065, USA
| | - Brian P O'Rourke
- Cell Biology Program, Memorial Sloan-Kettering Cancer Center, New York, NY 10065, USA
| | - Rajesh Kumar Soni
- Microchemistry and Proteomics Core Facility, Memorial Sloan-Kettering Cancer Center, New York, NY 10065 USA
| | - Prasad V Jallepalli
- Molecular Biology Program, Memorial Sloan-Kettering Cancer Center, New York, NY 10065, USA
| | - Ronald C Hendrickson
- Microchemistry and Proteomics Core Facility, Memorial Sloan-Kettering Cancer Center, New York, NY 10065 USA
| | - Meng-Fu Bryan Tsou
- Cell Biology Program, Memorial Sloan-Kettering Cancer Center, New York, NY 10065, USA; Weill Cornell Graduate School of Medical Sciences, Cornell University, New York, NY 10065, USA.
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26
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Yang R, Feldman JL. SPD-2/CEP192 and CDK Are Limiting for Microtubule-Organizing Center Function at the Centrosome. Curr Biol 2015; 25:1924-31. [PMID: 26119750 DOI: 10.1016/j.cub.2015.06.001] [Citation(s) in RCA: 31] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/16/2015] [Revised: 05/18/2015] [Accepted: 06/01/2015] [Indexed: 10/23/2022]
Abstract
The centrosome acts as the microtubule-organizing center (MTOC) during mitosis in animal cells. Microtubules are nucleated and anchored by γ-tubulin ring complexes (γ-TuRCs) embedded within the centrosome's pericentriolar material (PCM). The PCM is required for the localization of γ-TuRCs, and both are steadily recruited to the centrosome, culminating in a peak in MTOC function in metaphase. In differentiated cells, the centrosome is often attenuated as an MTOC and MTOC function is reassigned to non-centrosomal sites such as the apical membrane in epithelial cells, the nuclear envelope in skeletal muscle, and down the lengths of axons and dendrites in neurons. Hyperactive MTOC function at the centrosome is associated with epithelial cancers and with invasive behavior in tumor cells. Little is known about the mechanisms that limit MTOC activation at the centrosome. Here, we find that MTOC function at the centrosome is completely inactivated during cell differentiation in C. elegans embryonic intestinal cells and MTOC function is reassigned to the apical membrane. In cells that divide after differentiation, the cellular MTOC state switches between the membrane and the centrosome. Using cell fusion experiments in live embryos, we find that the centrosome MTOC state is dominant and that the inactive MTOC state of the centrosome is malleable; fusion of a mitotic cell to a differentiated or interphase cell results in rapid reactivation of the centrosome MTOC. We show that conversion of MTOC state involves the conserved centrosome protein SPD-2/CEP192 and CDK activity from the mitotic cell.
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Affiliation(s)
- Renzhi Yang
- Department of Biology, Stanford University, Stanford, CA 94305, USA
| | - Jessica L Feldman
- Department of Biology, Stanford University, Stanford, CA 94305, USA.
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Replication stress in Mammalian cells and its consequences for mitosis. Genes (Basel) 2015; 6:267-98. [PMID: 26010955 PMCID: PMC4488665 DOI: 10.3390/genes6020267] [Citation(s) in RCA: 80] [Impact Index Per Article: 8.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/14/2015] [Revised: 05/15/2015] [Accepted: 05/18/2015] [Indexed: 12/23/2022] Open
Abstract
The faithful transmission of genetic information to daughter cells is central to maintaining genomic stability and relies on the accurate and complete duplication of genetic material during each cell cycle. However, the genome is routinely exposed to endogenous and exogenous stresses that can impede the progression of replication. Such replication stress can be an early cause of cancer or initiate senescence. Replication stress, which primarily occurs during S phase, results in consequences during mitosis, jeopardizing chromosome segregation and, in turn, genomic stability. The traces of replication stress can be detected in the daughter cells during G1 phase. Alterations in mitosis occur in two types: 1) local alterations that correspond to breaks, rearrangements, intertwined DNA molecules or non-separated sister chromatids that are confined to the region of the replication dysfunction; 2) genome-wide chromosome segregation resulting from centrosome amplification (although centrosomes do not contain DNA), which amplifies the local replication stress to the entire genome. Here, we discuss the endogenous causes of replication perturbations, the mechanisms of replication fork restart and the consequences for mitosis, chromosome segregation and genomic stability.
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Katanin p80 regulates human cortical development by limiting centriole and cilia number. Neuron 2015; 84:1240-57. [PMID: 25521379 DOI: 10.1016/j.neuron.2014.12.017] [Citation(s) in RCA: 79] [Impact Index Per Article: 7.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 12/03/2014] [Indexed: 11/20/2022]
Abstract
Katanin is a microtubule-severing complex whose catalytic activities are well characterized, but whose in vivo functions are incompletely understood. Human mutations in KATNB1, which encodes the noncatalytic regulatory p80 subunit of katanin, cause severe microlissencephaly. Loss of Katnb1 in mice confirms essential roles in neurogenesis and cell survival, while loss of zebrafish katnb1 reveals specific roles for katnin p80 in early and late developmental stages. Surprisingly, Katnb1 null mutant mouse embryos display hallmarks of aberrant Sonic hedgehog signaling, including holoprosencephaly. KATNB1-deficient human cells show defective proliferation and spindle structure, while Katnb1 null fibroblasts also demonstrate a remarkable excess of centrioles, with supernumerary cilia but deficient Hedgehog signaling. Our results reveal unexpected functions for KATNB1 in regulating overall centriole, mother centriole, and cilia number, and as an essential gene for normal Hedgehog signaling during neocortical development.
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Fırat-Karalar EN, Stearns T. The centriole duplication cycle. Philos Trans R Soc Lond B Biol Sci 2015; 369:rstb.2013.0460. [PMID: 25047614 DOI: 10.1098/rstb.2013.0460] [Citation(s) in RCA: 112] [Impact Index Per Article: 11.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/12/2022] Open
Abstract
Centrosomes are the main microtubule-organizing centre of animal cells and are important for many critical cellular and developmental processes from cell polarization to cell division. At the core of the centrosome are centrioles, which recruit pericentriolar material to form the centrosome and act as basal bodies to nucleate formation of cilia and flagella. Defects in centriole structure, function and number are associated with a variety of human diseases, including cancer, brain diseases and ciliopathies. In this review, we discuss recent advances in our understanding of how new centrioles are assembled and how centriole number is controlled. We propose a general model for centriole duplication control in which cooperative binding of duplication factors defines a centriole 'origin of duplication' that initiates duplication, and passage through mitosis effects changes that license the centriole for a new round of duplication in the next cell cycle. We also focus on variations on the general theme in which many centrioles are created in a single cell cycle, including the specialized structures associated with these variations, the deuterosome in animal cells and the blepharoplast in lower plant cells.
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Affiliation(s)
| | - Tim Stearns
- Department of Biology, Stanford University, Stanford, CA 94305-5020, USA Department of Genetics, Stanford University Medical School, Stanford, CA 94305-5120, USA
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NR5A1 prevents centriole splitting by inhibiting centrosomal DNA-PK activation and β-catenin accumulation. Cell Commun Signal 2014; 12:55. [PMID: 25421435 PMCID: PMC4262199 DOI: 10.1186/s12964-014-0055-9] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/17/2014] [Accepted: 08/31/2014] [Indexed: 11/16/2022] Open
Abstract
Background Adrenogonadal cell growth and differentiation are controlled by nuclear receptor NR5A1 (Ad4BP/SF-1) that regulates the expression of adrenal and gonadal genes. In addition, SF-1 also resides in the centrosome and controls centrosome homeostasis by restricting the activity of centrosomal DNA-PK and CDK2/cyclin A. Results Here we show that SF-1 depletion resulted in centriole splitting and amplification due to aberrant activation of DNA-PK in the centrosome of mouse adrenocortical Y1 cells. In the absence of SF-1, GSK3β was aberrantly phosphorylated during G1 phase and β-catenin was accumulated in the centrosome, but not in the nucleus. DNA-PK inhibitor vanillin reversed these phenomena. SF-1 overexpression led to inhibition of centrosomal DNA-PK activation caused by SF-1 depletion. Both full-length SF-1 and truncated SF-1 devoid of its DNA-binding domain rescued the multiple centrosome phenotype caused by SF-1 depletion, indicating that the effect of SF-1 in the centrosome is not contributed by its DNA-binding domain. Furthermore, SF-1 interacted with cyclin A in the centrosome, but not in the nucleus. Depletion of SF-1 also resulted in centriole splitting, genomic instability and reduced growth of mouse testicular Leydig MA10 cells. Conclusion Centrosomal DNA-PK signaling triggers the accumulation of β-catenin, leading to centrosome over-duplication and centriole splitting. This cascade of centrosomal events results in genomic instability and reduced cell numbers.
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Nam HJ, Naylor RM, van Deursen JM. Centrosome dynamics as a source of chromosomal instability. Trends Cell Biol 2014; 25:65-73. [PMID: 25455111 DOI: 10.1016/j.tcb.2014.10.002] [Citation(s) in RCA: 61] [Impact Index Per Article: 5.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/23/2014] [Revised: 10/14/2014] [Accepted: 10/15/2014] [Indexed: 01/03/2023]
Abstract
Accurate segregation of duplicated chromosomes between two daughter cells depends on bipolar spindle formation, a metaphase state in which sister kinetochores are attached to microtubules emanating from opposite spindle poles. To ensure bi-orientation, cells possess surveillance systems that safeguard against microtubule-kinetochore attachment defects, including the spindle assembly checkpoint and the error correction machinery. However, recent developments have identified centrosome dynamics--that is, centrosome disjunction and poleward movement of duplicated centrosomes--as a central target for deregulation of bi-orientation in cancer cells. In this review, we discuss novel insights into the mechanisms that underlie centrosome dynamics and discuss how these mechanisms are perturbed in cancer cells to drive chromosome mis-segregation and advance neoplastic transformation.
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Affiliation(s)
- Hyun-Ja Nam
- Department of Pediatric and Adolescent Medicine, Mayo Clinic, Rochester, MN, USA
| | - Ryan M Naylor
- Department of Biochemistry and Molecular Biology, Mayo Clinic, Rochester, MN, USA
| | - Jan M van Deursen
- Department of Pediatric and Adolescent Medicine, Mayo Clinic, Rochester, MN, USA; Department of Biochemistry and Molecular Biology, Mayo Clinic, Rochester, MN, USA.
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32
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Lu Y, Roy R. Centrosome/Cell cycle uncoupling and elimination in the endoreduplicating intestinal cells of C. elegans. PLoS One 2014; 9:e110958. [PMID: 25360893 PMCID: PMC4215990 DOI: 10.1371/journal.pone.0110958] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/11/2013] [Accepted: 09/28/2014] [Indexed: 01/14/2023] Open
Abstract
The centrosome cycle is most often coordinated with mitotic cell division through the activity of various essential cell cycle regulators, consequently ensuring that the centriole is duplicated once, and only once, per cell cycle. However, this coupling can be altered in specific developmental contexts; for example, multi-ciliated cells generate hundreds of centrioles without any S-phase requirement for their biogenesis, while Drosophila follicle cells eliminate their centrosomes as they begin to endoreduplicate. In order to better understand how the centrosome cycle and the cell cycle are coordinated in a developmental context we use the endoreduplicating intestinal cell lineage of C. elegans to address how novel variations of the cell cycle impact this important process. In C. elegans, the larval intestinal cells undergo one nuclear division without subsequent cytokinesis, followed by four endocycles that are characterized by successive rounds of S-phase. We monitored the levels of centriolar/centrosomal markers and found that centrosomes lose their pericentriolar material following the nuclear division that occurs during the L1 stage and is thereafter never re-gained. The centrioles then become refractory to S phase regulators that would normally promote duplication during the first endocycle, after which they are eliminated during the L2 stage. Furthermore, we show that SPD-2 plays a central role in the numeral regulation of centrioles as a potential target of CDK activity. On the other hand, the phosphorylation on SPD-2 by Polo-like kinase, the transcriptional regulation of genes that affect centriole biogenesis, and the ubiquitin/proteasome degradation pathway, contribute collectively to the final elimination of the centrioles during the L2 stage.
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Affiliation(s)
- Yu Lu
- Department of Biology, The Developmental Biology Research Initiative, McGill University, Montreal, Quebec, Canada
| | - Richard Roy
- Department of Biology, The Developmental Biology Research Initiative, McGill University, Montreal, Quebec, Canada
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33
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Abstract
This paper describes the inner workings of centrioles (a pair of small organelles adjacent to the nucleus) as they create cell electropolarity, engage in cell division (mitosis), but in going awry, also promote the development of cancers. The electropolarity arises from vibrations of microtubules composing the centrioles. Mitosis begins as each centrioles duplicates itself by growing a daughter centriole on its side. If during duplication more than one daughter is grown, cancer can occur and the cells divide uncontrollably. Cancer cells with supernumerary centrioles have high electropolarity which can serve as an attractor for charged therapeutic nanoparticles.
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Affiliation(s)
- Ronald L. Huston
- Life Fellow ASME Department of Mechanical and Materials Engineering, University of Cincinnati, P.O. Box 210072, Cincinnati, OH 45221-0072 e-mail:
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34
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Increased centrosome amplification in aged stem cells of the Drosophila midgut. Biochem Biophys Res Commun 2014; 450:961-5. [DOI: 10.1016/j.bbrc.2014.06.085] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/02/2014] [Accepted: 06/17/2014] [Indexed: 11/23/2022]
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35
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Gottardo M, Callaini G, Riparbelli MG. Procentriole assembly without centriole disengagement - a paradox of male gametogenesis. J Cell Sci 2014; 127:3434-9. [PMID: 24938597 DOI: 10.1242/jcs.152843] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/18/2022] Open
Abstract
Disengagement of parent centrioles represents the licensing process to restrict centriole duplication exactly once during the cell cycle. However, we provide compelling evidence that this general rule is overridden in insect gametogenesis, when distinct procentrioles are generated during prophase of the first meiosis while parent centrioles are still engaged. Moreover, the number of procentrioles increases during the following meiotic divisions, and up to four procentrioles were found at the base of each mother centriole. However, procentrioles fail to organize a complete set of A-tubules and are thus unable to function as a template for centriole formation. Such a system, in which procentrioles form but halt growth, represents a unique model to analyze the process of cartwheel assembly and procentriole formation.
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Affiliation(s)
- Marco Gottardo
- Department of Life Sciences, University of Siena, Via A. Moro 4, 53100 Siena, Italy
| | - Giuliano Callaini
- Department of Life Sciences, University of Siena, Via A. Moro 4, 53100 Siena, Italy
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36
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Abstract
The belief that cohesin complexes link mother to daughter centrioles has received substantial experimental support. New studies challenge the primacy of cohesin in centriole engagement and provide a more nuanced view into the mechanisms for centriole disengagement in anaphase.
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Affiliation(s)
- Greenfield Sluder
- Department of Cell and Developmental Biology, University of Massachusetts Medical School, Worcester, MA 01655, USA.
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37
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Pihan GA. Centrosome dysfunction contributes to chromosome instability, chromoanagenesis, and genome reprograming in cancer. Front Oncol 2013; 3:277. [PMID: 24282781 PMCID: PMC3824400 DOI: 10.3389/fonc.2013.00277] [Citation(s) in RCA: 100] [Impact Index Per Article: 8.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/29/2013] [Accepted: 10/28/2013] [Indexed: 12/19/2022] Open
Abstract
The unique ability of centrosomes to nucleate and organize microtubules makes them unrivaled conductors of important interphase processes, such as intracellular payload traffic, cell polarity, cell locomotion, and organization of the immunologic synapse. But it is in mitosis that centrosomes loom large, for they orchestrate, with clockmaker's precision, the assembly and functioning of the mitotic spindle, ensuring the equal partitioning of the replicated genome into daughter cells. Centrosome dysfunction is inextricably linked to aneuploidy and chromosome instability, both hallmarks of cancer cells. Several aspects of centrosome function in normal and cancer cells have been molecularly characterized during the last two decades, greatly enhancing our mechanistic understanding of this tiny organelle. Whether centrosome defects alone can cause cancer, remains unanswered. Until recently, the aggregate of the evidence had suggested that centrosome dysfunction, by deregulating the fidelity of chromosome segregation, promotes and accelerates the characteristic Darwinian evolution of the cancer genome enabled by increased mutational load and/or decreased DNA repair. Very recent experimental work has shown that missegregated chromosomes resulting from centrosome dysfunction may experience extensive DNA damage, suggesting additional dimensions to the role of centrosomes in cancer. Centrosome dysfunction is particularly prevalent in tumors in which the genome has undergone extensive structural rearrangements and chromosome domain reshuffling. Ongoing gene reshuffling reprograms the genome for continuous growth, survival, and evasion of the immune system. Manipulation of molecular networks controlling centrosome function may soon become a viable target for specific therapeutic intervention in cancer, particularly since normal cells, which lack centrosome alterations, may be spared the toxicity of such therapies.
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Affiliation(s)
- German A Pihan
- Department of Pathology and Laboratory Medicine, Beth Israel Deaconess Medical Center, Harvard Medical School , Boston, MA , USA
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38
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Mbom BC, Nelson WJ, Barth A. β-catenin at the centrosome: discrete pools of β-catenin communicate during mitosis and may co-ordinate centrosome functions and cell cycle progression. Bioessays 2013; 35:804-9. [PMID: 23804296 DOI: 10.1002/bies.201300045] [Citation(s) in RCA: 40] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/10/2022]
Abstract
Beta-catenin is a multifunctional protein with critical roles in cell-cell adhesion, Wnt-signaling and the centrosome cycle. Whereas the roles of β-catenin in cell-cell adhesion and Wnt-signaling have been studied extensively, the mechanism(s) involving β-catenin in centrosome functions are poorly understood. β-Catenin localizes to centrosomes and promotes mitotic progression. NIMA-related protein kinase 2 (Nek2), which stimulates centrosome separation, binds to and phosphorylates β-catenin. β-Catenin interacting proteins involved in Wnt signaling such as adenomatous polyposis coli, Axin, and GSK3β, are also localized at centrosomes and play roles in promoting mitotic progression. Additionally, proteins associated with cell-cell adhesion sites, such as dynein, regulate mitotic spindle positioning. These roles of proteins at the cell cortex and Wnt signaling that involve β-catenin indicate a cross-talk between different sub-cellular sites in the cell at mitosis, and that different pools of β-catenin may co-ordinate centrosome functions and cell cycle progression.
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Affiliation(s)
- Bertrade C Mbom
- Department of Biology, Stanford University, Stanford, CA, USA
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39
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Schvarzstein M, Pattabiraman D, Bembenek JN, Villeneuve AM. Meiotic HORMA domain proteins prevent untimely centriole disengagement during Caenorhabditis elegans spermatocyte meiosis. Proc Natl Acad Sci U S A 2013; 110:E898-907. [PMID: 23401519 PMCID: PMC3593872 DOI: 10.1073/pnas.1213888110] [Citation(s) in RCA: 22] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/11/2022] Open
Abstract
In many species where oocytes lack centrosomes, sperm contribute both genetic material and centriole(s) to the zygote. Correct centriole organization during male meiosis is critical to guarantee a normal bipolar mitotic spindle in the zygote. During Caenorhabditis elegans male meiosis, centrioles normally undergo two rounds of duplication, resulting in haploid sperm each containing a single tightly engaged centriole pair. Here we identify an unanticipated role for C. elegans HORMA (Hop1/Rev7/Mad2) domain proteins HTP-1/2 and HIM-3 in regulating centriole disengagement during spermatocyte meiosis. In him-3 and htp-1 htp-2 mutants, centrioles separate inappropriately during meiosis II, resulting in spermatids with disengaged centrioles. Moreover, extra centrosomes are detected in a subset of zygotes. Together, these data implicate HIM-3 and HTP-1/2 in preventing centriole disengagement during meiosis II. We showed previously that HTP-1/2 prevents premature loss of sister chromatid cohesion during the meiotic divisions by inhibiting removal of meiotic cohesin complexes containing the REC-8 subunit. Worms lacking REC-8, or expressing a mutant separase protein with elevated local concentration at centrosomes and in sperm, likewise exhibit inappropriate centriole separation during spermatocyte meiosis. These observations are consistent with HIM-3 and HTP-1/2 preventing centriole disengagement by inhibiting separase-dependent cohesin removal. Our data suggest that the same specialized meiotic mechanisms that function to prevent premature release of sister chromatid cohesion during meiosis I in C. elegans also function to inhibit centriole separation at meiosis II, thereby ensuring that the zygote inherits the appropriate complement of chromosomes and centrioles.
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Affiliation(s)
- Mara Schvarzstein
- Departments of Developmental Biology and Genetics, Stanford University School of Medicine, Stanford, CA 94305; and
| | - Divya Pattabiraman
- Departments of Developmental Biology and Genetics, Stanford University School of Medicine, Stanford, CA 94305; and
| | - Joshua N. Bembenek
- Department of Biochemistry, Cellular, and Molecular Biology, University of Tennessee, Knoxville, TN 37916
| | - Anne M. Villeneuve
- Departments of Developmental Biology and Genetics, Stanford University School of Medicine, Stanford, CA 94305; and
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40
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Brownlee CW, Rogers GC. Show me your license, please: deregulation of centriole duplication mechanisms that promote amplification. Cell Mol Life Sci 2013; 70:1021-34. [PMID: 22892665 PMCID: PMC11113234 DOI: 10.1007/s00018-012-1102-6] [Citation(s) in RCA: 34] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/31/2012] [Revised: 07/13/2012] [Accepted: 07/17/2012] [Indexed: 12/13/2022]
Abstract
Centrosomes are organelles involved in generating and organizing the interphase microtubule cytoskeleton, mitotic spindles and cilia. At the centrosome core are a pair of centrioles, structures that act as the duplicating elements of this organelle. Centrioles function to recruit and organize pericentriolar material which nucleates microtubules. While centrioles are relatively simple in construction, the mechanics of centriole biogenesis remain an important yet poorly understood process. More mysterious still are the regulatory mechanisms that oversee centriole assembly. The fidelity of centriole duplication is critical as defects in either the assembly or number of centrioles promote aneuploidy, primary microcephaly, birth defects, ciliopathies and tumorigenesis. In addition, some pathogens employ mechanisms to promote centriole overduplication to the detriment of the host cell. This review summarizes our current understanding of this important topic, highlighting the need for further study if new therapeutics are to be developed to treat diseases arising from defects of centrosome duplication.
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Affiliation(s)
- Christopher W. Brownlee
- Department of Cellular and Molecular Medicine, Arizona Cancer Center, University of Arizona, Tucson, AZ 85724 USA
| | - Gregory C. Rogers
- Department of Cellular and Molecular Medicine, Arizona Cancer Center, University of Arizona, Tucson, AZ 85724 USA
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41
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Kodani A, Salomé Sirerol-Piquer M, Seol A, Garcia-Verdugo JM, Reiter JF. Kif3a interacts with Dynactin subunit p150 Glued to organize centriole subdistal appendages. EMBO J 2013; 32:597-607. [PMID: 23386061 DOI: 10.1038/emboj.2013.3] [Citation(s) in RCA: 68] [Impact Index Per Article: 5.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/19/2012] [Accepted: 12/04/2012] [Indexed: 01/17/2023] Open
Abstract
Formation of cilia, microtubule-based structures that function in propulsion and sensation, requires Kif3a, a subunit of Kinesin II essential for intraflagellar transport (IFT). We have found that, Kif3a is also required to organize centrioles. In the absence of Kif3a, the subdistal appendages of centrioles are disorganized and lack p150(Glued) and Ninein. Consequently, microtubule anchoring, centriole cohesion and basal foot formation are abrogated by loss of Kif3a. Kif3a localizes to the mother centriole and interacts with the Dynactin subunit p150(Glued). Depletion of p150(Glued) phenocopies the effects of loss of Kif3a, indicating that Kif3a recruitment of p150(Glued) is critical for subdistal appendage formation. The transport functions of Kif3a are dispensable for subdistal appendage organization as mutant forms of Kif3a lacking motor activity or the motor domain can restore p150(Glued) localization. Comparison to cells lacking Ift88 reveals that the centriolar functions of Kif3a are independent of IFT. Thus, in addition to its ciliogenic roles, Kif3a recruits p150(Glued) to the subdistal appendages of mother centrioles, critical for centrosomes to function as microtubule-organizing centres.
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Affiliation(s)
- Andrew Kodani
- Department of Biochemistry and Biophysics, Cardiovascular Research Institute, University of California, San Francisco, CA 94158-9001, USA
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42
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A novel role of human holliday junction resolvase GEN1 in the maintenance of centrosome integrity. PLoS One 2012; 7:e49687. [PMID: 23166748 PMCID: PMC3500319 DOI: 10.1371/journal.pone.0049687] [Citation(s) in RCA: 15] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/21/2012] [Accepted: 10/11/2012] [Indexed: 02/07/2023] Open
Abstract
The maintenance of genomic stability requires accurate genome replication, repair of DNA damage, and the precise segregation of chromosomes in mitosis. GEN1 possesses Holliday junction resolvase activity in vitro and presumably functions in homology driven repair of DNA double strand breaks. However, little is currently known about the cellular functions of human GEN1. In the present study we demonstrate that GEN1 is a novel centrosome associated protein and we characterize the various phenotypes associated with GEN1 deficiency. We identify an N-terminal centrosome localization signal in GEN1, which is required and sufficient for centrosome localization. We report that GEN1 depletion results in aberrant centrosome numbers associated with the formation of multiple spindle poles in mitosis, an increased number of cells with multi-nuclei, increased apoptosis and an elevated level of spontaneous DNA damage. We find homologous recombination severely impaired in GEN1 deficient cells, suggesting that GEN1 functions as a Holliday junction resolvase in vivo as well as in vitro. Complementation of GEN1 depleted cells with various GEN1 constructs revealed that centrosome association but not catalytic activity of GEN1 is required for preventing centrosome hyper-amplification, formation of multiple mitotic spindles, and multi-nucleation. Our findings provide novel insight into the biological functions of GEN1 by uncovering an important role of GEN1 in the regulation of centrosome integrity.
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Ruan K, Ye F, Li C, Liou YC, Lin SC, Lin SY. PLK1 interacts and phosphorylates Axin that is essential for proper centrosome formation. PLoS One 2012; 7:e49184. [PMID: 23155463 PMCID: PMC3498349 DOI: 10.1371/journal.pone.0049184] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/23/2012] [Accepted: 10/04/2012] [Indexed: 11/18/2022] Open
Abstract
Abnormal amplification of centrosomes could lead to improper chromosome segregation and aneuploidy and is implicated in cancer development. Here, we demonstrate that Axin, a scaffolding protein in Wnt signaling, is phosphorylated by PLK1 during mitosis. Phosphorylation of Axin Ser-157 by PLK1 abolished Axin association with γ-tubulin, while substitution of Ser-157 with alanine exhibited sustained interaction with γ-tubulin. In addition, overexpression of Axin-S157A significantly increased the number of cells with multi-centrosomes. These results suggest that the phosphorylation status of Axin, mediated by PLK1, dynamically regulates its association with γ-tubulin and centrosome formation and segregation.
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Affiliation(s)
- Ka Ruan
- State Key Laboratory of Cellular Stress Biology, School of Life Sciences, Xiamen University, Xiamen, Fujian, China
| | - Fan Ye
- Department of Biological Science, National University of Singapore, Singapore, Republic of Singapore
| | - Chenyu Li
- Department of Biological Science, National University of Singapore, Singapore, Republic of Singapore
| | - Yih-Cherng Liou
- Department of Biological Science, National University of Singapore, Singapore, Republic of Singapore
| | - Sheng-Cai Lin
- State Key Laboratory of Cellular Stress Biology, School of Life Sciences, Xiamen University, Xiamen, Fujian, China
| | - Shu-Yong Lin
- State Key Laboratory of Cellular Stress Biology, School of Life Sciences, Xiamen University, Xiamen, Fujian, China
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Hatano T, Sluder G. The interrelationship between APC/C and Plk1 activities in centriole disengagement. Biol Open 2012; 1:1153-60. [PMID: 23213396 PMCID: PMC3507193 DOI: 10.1242/bio.20122626] [Citation(s) in RCA: 21] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/23/2012] [Accepted: 08/06/2012] [Indexed: 11/20/2022] Open
Abstract
Mother–daughter centriole disengagement, the necessary first step in centriole duplication, involves Plk1 activity in early mitosis and separase activity after APC/C activity mediates securin degradation. Plk1 activity is thought to be essential and sufficient for centriole disengagement with separase activity playing a supporting but non-essential role. In separase null cells, however, centriole disengagement is substantially delayed. The ability of APC/C activity alone to mediate centriole disengagement has not been directly tested. We investigate the interrelationship between Plk1 and APC/C activities in disengaging centrioles in S or G2 HeLa and RPE1 cells, cell types that do not reduplicate centrioles when arrested in S phase. Knockdown of the interphase APC/C inhibitor Emi1 leads to centriole disengagement and reduplication of the mother centrioles, though this is slow. Strong inhibition of Plk1 activity, if any, during S does not block centriole disengagement and mother centriole reduplication in Emi1 depleted cells. Centriole disengagement depends on APC/C–Cdh1 activity, not APC/C–Cdc20 activity. Also, Plk1 and APC/C–Cdh1 activities can independently promote centriole disengagement in G2 arrested cells. Thus, Plk1 and APC/C–Cdh1 activities are independent but slow pathways for centriole disengagement. By having two slow mechanisms for disengagement working together, the cell ensures that centrioles will not prematurely separate in late G2 or early mitosis, thereby risking multipolar spindle assembly, but rather disengage in a timely fashion only late in mitosis.
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Affiliation(s)
- Toshiyuki Hatano
- Department of Cell Biology, University of Massachusetts Medical School , Worcester, MA 01605 , USA
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Dantas TJ, Daly OM, Morrison CG. Such small hands: the roles of centrins/caltractins in the centriole and in genome maintenance. Cell Mol Life Sci 2012; 69:2979-97. [PMID: 22460578 PMCID: PMC11114748 DOI: 10.1007/s00018-012-0961-1] [Citation(s) in RCA: 42] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/13/2011] [Revised: 02/20/2012] [Accepted: 03/12/2012] [Indexed: 01/11/2023]
Abstract
Centrins are small, highly conserved members of the EF-hand superfamily of calcium-binding proteins that are found throughout eukaryotes. They play a major role in ensuring the duplication and appropriate functioning of the ciliary basal bodies in ciliated cells. They have also been localised to the centrosome, which is the major microtubule organising centre in animal somatic cells. We describe the identification, cloning and characterisation of centrins in multiple eukaryotic species. Although centrins have been implicated in centriole biogenesis, recent results have indicated that centrosome duplication can, in fact, occur in the absence of centrins. We discuss these data and the non-centrosomal functions that are emerging for the centrins. In particular, we discuss the involvement of centrins in nucleotide excision repair, a process that repairs the DNA lesions that are induced primarily by ultraviolet irradiation. We discuss how centrin may be involved in these diverse processes and contribute to nuclear and cytoplasmic events.
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Affiliation(s)
- Tiago J. Dantas
- Centre for Chromosome Biology, School of Natural Sciences, National University of Ireland Galway, University Road, Galway, Ireland
| | - Owen M. Daly
- Centre for Chromosome Biology, School of Natural Sciences, National University of Ireland Galway, University Road, Galway, Ireland
| | - Ciaran G. Morrison
- Centre for Chromosome Biology, School of Natural Sciences, National University of Ireland Galway, University Road, Galway, Ireland
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Hossain M, Stillman B. Meier-Gorlin syndrome mutations disrupt an Orc1 CDK inhibitory domain and cause centrosome reduplication. Genes Dev 2012; 26:1797-810. [PMID: 22855792 DOI: 10.1101/gad.197178.112] [Citation(s) in RCA: 55] [Impact Index Per Article: 4.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/24/2022]
Abstract
Like DNA replication, centrosomes are licensed to duplicate once per cell division cycle to ensure genetic stability. In addition to regulating DNA replication, the Orc1 subunit of the human origin recognition complex controls centriole and centrosome copy number. Here we report that Orc1 harbors a PACT centrosome-targeting domain and a separate domain that differentially inhibits the protein kinase activities of Cyclin E-CDK2 and Cyclin A-CDK2. A cyclin-binding motif (Cy motif) is required for Orc1 to bind Cyclin A and inhibit Cyclin A-CDK2 kinase activity but has no effect on Cyclin E-CDK2 kinase activity. In contrast, Orc1 inhibition of Cyclin E-CDK2 kinase activity occurs by a different mechanism that is affected by Orc1 mutations identified in Meier-Gorlin syndrome patients. The cyclin/CDK2 kinase inhibitory domain of Orc1, when tethered to the PACT domain, localizes to centrosomes and blocks centrosome reduplication. Meier-Gorlin syndrome mutations that disrupt Cyclin E-CDK2 kinase inhibition also allow centrosome reduplication. Thus, Orc1 contains distinct domains that control centrosome copy number and DNA replication. We suggest that the Orc1 mutations present in some Meier-Gorlin syndrome patients contribute to the pronounced microcephaly and dwarfism observed in these individuals by altering centrosome duplication in addition to DNA replication defects.
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Affiliation(s)
- Manzar Hossain
- Cold Spring Harbor Laboratory, Cold Spring Harbor, New York 11724, USA
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Glaser N, Stopper H. Patulin: Mechanism of genotoxicity. Food Chem Toxicol 2012; 50:1796-801. [DOI: 10.1016/j.fct.2012.02.096] [Citation(s) in RCA: 81] [Impact Index Per Article: 6.2] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/18/2012] [Revised: 02/27/2012] [Accepted: 02/28/2012] [Indexed: 11/30/2022]
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Matsuo K, Ohsumi K, Iwabuchi M, Kawamata T, Ono Y, Takahashi M. Kendrin is a novel substrate for separase involved in the licensing of centriole duplication. Curr Biol 2012; 22:915-21. [PMID: 22542101 DOI: 10.1016/j.cub.2012.03.048] [Citation(s) in RCA: 89] [Impact Index Per Article: 6.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/07/2011] [Revised: 02/13/2012] [Accepted: 03/16/2012] [Indexed: 11/17/2022]
Abstract
The centrosome, consisting of a pair of centrioles surrounded by pericentriolar material, directs the formation of bipolar spindles during mitosis. Aberrant centrosome number can promote chromosome instability, which is implicated in tumorigenesis. Thus, centrosome duplication needs to be tightly regulated to occur only once per cell cycle. Separase, a cysteine protease that triggers sister chromatid separation, is involved in centriole disengagement, which licenses centrosomes for the next round of duplication. However, at least two questions remain unsolved: what is the substrate relevant to the disengagement, and how does separase, activated at anaphase onset, act on the disengagement that occurs during late mitosis. Here, we show that kendrin, also named pericentrin, is cleaved by activated separase at a consensus site in vivo and in vitro, and this leads to the delayed release of kendrin from the centrosome later in mitosis. Furthermore, we demonstrate that expression of a noncleavable kendrin mutant suppresses centriole disengagement and subsequent centriole duplication. Based on these results, we propose that kendrin is a novel and crucial substrate for separase at the centrosome, protecting the engaged centrioles from premature disengagement and thereby blocking reduplication until the cell passes through mitosis.
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Affiliation(s)
- Kazuhiko Matsuo
- Faculty of Pharmaceutical Science, Teikyo Heisei University, Ichihara, Japan
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Loss of cyclin-dependent kinase 2 (CDK2) inhibitory phosphorylation in a CDK2AF knock-in mouse causes misregulation of DNA replication and centrosome duplication. Mol Cell Biol 2012; 32:1421-32. [PMID: 22331465 DOI: 10.1128/mcb.06721-11] [Citation(s) in RCA: 18] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/14/2022] Open
Abstract
Cyclin-dependent kinase 1 (CDK1) inhibitory phosphorylation controls the onset of mitosis and is essential for the checkpoint pathways that prevent the G(2)- to M-phase transition in cells with unreplicated or damaged DNA. To address whether CDK2 inhibitory phosphorylation plays a similar role in cell cycle regulation and checkpoint responses at the start of the S phase, we constructed a mouse strain in which the two CDK2 inhibitory phosphorylation sites, threonine 14 and tyrosine 15, were changed to alanine and phenylalanine, respectively (CDK2AF). This approach showed that inhibitory phosphorylation of CDK2 had a major role in controlling cyclin E-associated kinase activity and thus both determined the timing of DNA replication in a normal cell cycle and regulated centrosome duplication. Further, DNA damage in G(1) CDK2AF cells did not downregulate cyclin E-CDK2 activity when the CDK inhibitor p21 was also knocked down. We were surprised to find that this was insufficient to cause cells to bypass the checkpoint and enter the S phase. This led to the discovery of two previously unrecognized pathways that control the activity of cyclin A at the G(1) DNA damage checkpoint and may thereby prevent S-phase entry even when cyclin E-CDK2 activity is deregulated.
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50
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Bahtz R, Seidler J, Arnold M, Haselmann-Weiss U, Antony C, Lehmann WD, Hoffmann I. GCP6 is a substrate of Plk4 and required for centriole duplication. J Cell Sci 2012; 125:486-96. [PMID: 22302995 DOI: 10.1242/jcs.093930] [Citation(s) in RCA: 66] [Impact Index Per Article: 5.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
Centriole duplication occurs once per cell cycle and requires Plk4, a member of the Polo-like kinase family. A key component of the centrosome is the γ-tubulin ring complex (γ-TuRC) that nucleates microtubules. GCP6 is a member of the γ-TuRC, but its role in human cells and the regulation of its functions remain unclear. Here we report that depletion of human GCP6 prevents assembly of the γ-TuRC and induces a high percentage of monopolar spindles. These spindles are characterized by a loss of centrosomal γ-tubulin and reduced centriole numbers. We found that GCP6 is localized in the pericentriolar material but also at distal portions of centrioles. In addition, GCP6 is required for centriole duplication and Plk4-induced centriole overduplication. GCP6 interacts with and is phosphorylated by Plk4. Moreover, we find that Plk4-dependent phosphorylation of GCP6 regulates centriole duplication. These data suggest that GCP6 is a target of Plk4 in centriole biogenesis.
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Affiliation(s)
- Ramona Bahtz
- Cell Cycle Control and Carcinogenesis, German Cancer Research Center (DKFZ), 69120 Heidelberg, Germany
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