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Prakash A, Paunikar S, Webber M, McDermott E, Vellanki SH, Thompson K, Dockery P, Jahns H, Brown JAL, Hopkins AM, Bourke E. Centrosome amplification promotes cell invasion via cell-cell contact disruption and Rap-1 activation. J Cell Sci 2023; 136:jcs261150. [PMID: 37772773 PMCID: PMC10629695 DOI: 10.1242/jcs.261150] [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: 03/08/2023] [Accepted: 09/19/2023] [Indexed: 09/30/2023] Open
Abstract
Centrosome amplification (CA) is a prominent feature of human cancers linked to tumorigenesis in vivo. Here, we report mechanistic contributions of CA induction alone to tumour architecture and extracellular matrix (ECM) remodelling. CA induction in non-tumorigenic breast cells MCF10A causes cell migration and invasion, with underlying disruption of epithelial cell-cell junction integrity and dysregulation of expression and subcellular localisation of cell junction proteins. CA also elevates expression of integrin β-3, its binding partner fibronectin-1 and matrix metalloproteinase enzymes, promoting cell-ECM attachment, ECM degradation, and a migratory and invasive cell phenotype. Using a chicken embryo xenograft model for in vivo validation, we show that CA-induced (+CA) MCF10A cells invade into the chick mesodermal layer, with inflammatory cell infiltration and marked focal reactions between chorioallantoic membrane and cell graft. We also demonstrate a key role of small GTPase Rap-1 signalling through inhibition using GGTI-298, which blocked various CA-induced effects. These insights reveal that in normal cells, CA induction alone (without additional oncogenic alterations) is sufficient to confer early pro-tumorigenic changes within days, acting through Rap-1-dependent signalling to alter cell-cell contacts and ECM disruption.
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Affiliation(s)
- Anu Prakash
- Lambe Institute for Translational Research, Discipline of Pathology, Centre for Chromosome Biology, University of Galway, Galway H91 V4AY, Ireland
| | - Shishir Paunikar
- Lambe Institute for Translational Research, Discipline of Pathology, Centre for Chromosome Biology, University of Galway, Galway H91 V4AY, Ireland
| | - Mark Webber
- Lambe Institute for Translational Research, Discipline of Pathology, Centre for Chromosome Biology, University of Galway, Galway H91 V4AY, Ireland
| | - Emma McDermott
- Centre for Microscopy and Imaging, Discipline of Anatomy, School of Medicine, University of Galway, Galway H91 W5P7, Ireland
| | - Sri H. Vellanki
- Department of Surgery, Beaumont Hospital, Royal College of Surgeons in Ireland, Dublin D09 DK19, Ireland
| | - Kerry Thompson
- Centre for Microscopy and Imaging, Discipline of Anatomy, School of Medicine, University of Galway, Galway H91 W5P7, Ireland
| | - Peter Dockery
- Centre for Microscopy and Imaging, Discipline of Anatomy, School of Medicine, University of Galway, Galway H91 W5P7, Ireland
| | - Hanne Jahns
- Pathobiology Section, School of Veterinary Medicine, University College Dublin, Dublin, Ireland
| | - James A. L. Brown
- Department of Biological Sciences, University of Limerick, Limerick V94T9PX, Ireland
- Limerick Digital Cancer Research Centre (LDCRC) and Health Research Institute, University of Limerick, Limerick V94T9PX, Ireland
| | - Ann M. Hopkins
- Department of Surgery, Beaumont Hospital, Royal College of Surgeons in Ireland, Dublin D09 DK19, Ireland
| | - Emer Bourke
- Lambe Institute for Translational Research, Discipline of Pathology, Centre for Chromosome Biology, University of Galway, Galway H91 V4AY, Ireland
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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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Song H, Kim EH, Hong J, Gwon D, Kim JW, Bae GU, Jang CY. Hornerin mediates phosphorylation of the polo-box domain in Plk1 by Chk1 to induce death in mitosis. Cell Death Differ 2023; 30:2151-2166. [PMID: 37596441 PMCID: PMC10482915 DOI: 10.1038/s41418-023-01208-y] [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: 02/03/2023] [Revised: 05/25/2023] [Accepted: 06/13/2023] [Indexed: 08/20/2023] Open
Abstract
The centrosome assembles a bipolar spindle for faithful chromosome segregation during mitosis. To prevent the inheritance of DNA damage, the DNA damage response (DDR) triggers programmed spindle multipolarity and concomitant death in mitosis through a poorly understood mechanism. We identified hornerin, which forms a complex with checkpoint kinase 1 (Chk1) and polo-like kinase 1 (Plk1) to mediate phosphorylation at the polo-box domain (PBD) of Plk1, as the link between the DDR and death in mitosis. We demonstrate that hornerin mediates DDR-induced precocious centriole disengagement through a dichotomous mechanism that includes sequestration of Sgo1 and Plk1 in the cytoplasm through phosphorylation of the PBD in Plk1 by Chk1. Phosphorylation of the PBD in Plk1 abolishes the interaction with Sgo1 and phosphorylation-dependent Sgo1 translocation to the centrosome, leading to precocious centriole disengagement and spindle multipolarity. Mechanistically, hornerin traps phosphorylated Plk1 in the cytoplasm. Furthermore, PBD phosphorylation inactivates Plk1 and disrupts Cep192::Aurora A::Plk1 complex translocation to the centrosome and concurrent centrosome maturation. Remarkably, hornerin depletion leads to chemoresistance against DNA damaging agents by attenuating DDR-induced death in mitosis. These results reveal how the DDR eradicates mitotic cells harboring DNA damage to ensure genome integrity during cell division.
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Affiliation(s)
- Haiyu Song
- Research Institute of Pharmaceutical Sciences, College of Pharmacy, Sookmyung Women's University, Seoul, 04310, Republic of Korea
| | - Eun Ho Kim
- Department of Biochemistry, School of Medicine, Catholic University of Daegu, Daegu, 42472, Republic of Korea
| | - Jihee Hong
- Research Institute of Pharmaceutical Sciences, College of Pharmacy, Sookmyung Women's University, Seoul, 04310, Republic of Korea
| | - Dasom Gwon
- Research Institute of Pharmaceutical Sciences, College of Pharmacy, Sookmyung Women's University, Seoul, 04310, Republic of Korea
| | - Jee Won Kim
- Research Institute of Pharmaceutical Sciences, College of Pharmacy, Sookmyung Women's University, Seoul, 04310, Republic of Korea
| | - Gyu-Un Bae
- Research Institute of Pharmaceutical Sciences, College of Pharmacy, Sookmyung Women's University, Seoul, 04310, Republic of Korea.
| | - Chang-Young Jang
- Research Institute of Pharmaceutical Sciences, College of Pharmacy, Sookmyung Women's University, Seoul, 04310, Republic of Korea.
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4
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Bloomfield M, Cimini D. The fate of extra centrosomes in newly formed tetraploid cells: should I stay, or should I go? Front Cell Dev Biol 2023; 11:1210983. [PMID: 37576603 PMCID: PMC10413984 DOI: 10.3389/fcell.2023.1210983] [Citation(s) in RCA: 5] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/23/2023] [Accepted: 07/17/2023] [Indexed: 08/15/2023] Open
Abstract
An increase in centrosome number is commonly observed in cancer cells, but the role centrosome amplification plays along with how and when it occurs during cancer development is unclear. One mechanism for generating cancer cells with extra centrosomes is whole genome doubling (WGD), an event that occurs in over 30% of human cancers and is associated with poor survival. Newly formed tetraploid cells can acquire extra centrosomes during WGD, and a generally accepted model proposes that centrosome amplification in tetraploid cells promotes cancer progression by generating aneuploidy and chromosomal instability. Recent findings, however, indicate that newly formed tetraploid cells in vitro lose their extra centrosomes to prevent multipolar cell divisions. Rather than persistent centrosome amplification, this evidence raises the possibility that it may be advantageous for tetraploid cells to initially restore centrosome number homeostasis and for a fraction of the population to reacquire additional centrosomes in the later stages of cancer evolution. In this review, we explore the different evolutionary paths available to newly formed tetraploid cells, their effects on centrosome and chromosome number distribution in daughter cells, and their probabilities of long-term survival. We then discuss the mechanisms that may alter centrosome and chromosome numbers in tetraploid cells and their relevance to cancer progression following WGD.
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Affiliation(s)
- Mathew Bloomfield
- Department of Biological Sciences and Fralin Life Sciences Institute, Virginia Tech, Blacksburg, VA, United States
| | - Daniela Cimini
- Department of Biological Sciences and Fralin Life Sciences Institute, Virginia Tech, Blacksburg, VA, United States
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Abstract
ABSTRACT With rapid technical advances, ionizing radiation has been put into wider application in ordinary living, with the worst cytological effect on the human body being cell death. Moreover, according to the Nomenclature Committee on Cell Death, the method of radiation-induced cell death, usually classified as interphase and proliferative death, undergoes more detailed classifications oriented by its molecular mechanism. Elaborating its mode and molecular mechanism is crucial for the protection and treatment of radiation injury, as well as the radiotherapy and recovery of tumors. Varying with the changes of the radiation dose and the environment, the diverse targets and pathways of ionizing radiation result in various cell deaths. This review focuses on classifications of radiation-induced cell death and its molecular mechanism. We also examine the main characteristics of ionizing radiation-induced cell death. The modes of radiation-induced cell death can be classified as apoptosis, necrosis, autophagy-dependent cell death, pyroptosis, ferroptosis, immunogenic cell death, and non-lethal processes. Once the dose is high enough, radiation effects mostly appear as destructiveness ("destructiveness" is used to describe a situation in which cells do not have the opportunity to undergo a routine death process, in which case high-dose radiation works like a physical attack). This breaks up or even shatters cells, making it difficult to find responses of the cell itself. Due to diversities concerning cell phenotypes, phases of cell cycle, radiation dose, and even cellular subregions, various methods of cell death occur, which are difficult to identify and classify. Additionally, the existence of common initial activation and signaling molecules among all kinds of cell deaths, as well as sophisticated crossways in cellular molecules, makes it more laborious to distinguish and classify various cell deaths.
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Affiliation(s)
- Yunfei Jiao
- College of Basic Medicine, Second Military Medical University, Xiangyin Road, 200433 Shanghai, PR China
- Incubation Base for Undergraduates’ Innovation Practice, Department of Radiation Medicine, Faculty of Naval Medicine, Naval Medical University, Shanghai, China
- Department of Gastroenterology, Changhai Hospital, Naval Medical University, Shanghai, China
| | - Fangyu Cao
- College of Basic Medicine, Second Military Medical University, Xiangyin Road, 200433 Shanghai, PR China
- Incubation Base for Undergraduates’ Innovation Practice, Department of Radiation Medicine, Faculty of Naval Medicine, Naval Medical University, Shanghai, China
| | - Hu Liu
- Department of Radiation Medicine, Faculty of Naval Medicine, Second Military Medical University, Shanghai, China
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6
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Lu JY, Simon M, Zhao Y, Ablaeva J, Corson N, Choi Y, Yamada KYH, Schork NJ, Hood WR, Hill GE, Miller RA, Seluanov A, Gorbunova V. Comparative transcriptomics reveals circadian and pluripotency networks as two pillars of longevity regulation. Cell Metab 2022; 34:836-856.e5. [PMID: 35580607 PMCID: PMC9364679 DOI: 10.1016/j.cmet.2022.04.011] [Citation(s) in RCA: 27] [Impact Index Per Article: 9.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/13/2021] [Revised: 02/24/2022] [Accepted: 04/22/2022] [Indexed: 01/24/2023]
Abstract
Mammals differ more than 100-fold in maximum lifespan. Here, we conducted comparative transcriptomics on 26 species with diverse lifespans. We identified thousands of genes with expression levels negatively or positively correlated with a species' maximum lifespan (Neg- or Pos-MLS genes). Neg-MLS genes are primarily involved in energy metabolism and inflammation. Pos-MLS genes show enrichment in DNA repair, microtubule organization, and RNA transport. Expression of Neg- and Pos-MLS genes is modulated by interventions, including mTOR and PI3K inhibition. Regulatory networks analysis showed that Neg-MLS genes are under circadian regulation possibly to avoid persistent high expression, whereas Pos-MLS genes are targets of master pluripotency regulators OCT4 and NANOG and are upregulated during somatic cell reprogramming. Pos-MLS genes are highly expressed during embryogenesis but significantly downregulated after birth. This work provides targets for anti-aging interventions by defining pathways correlating with longevity across mammals and uncovering circadian and pluripotency networks as central regulators of longevity.
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Affiliation(s)
- J Yuyang Lu
- Department of Biology, University of Rochester, Rochester, NY 14627, USA
| | - Matthew Simon
- Department of Biology, University of Rochester, Rochester, NY 14627, USA
| | - Yang Zhao
- Department of Biology, University of Rochester, Rochester, NY 14627, USA
| | - Julia Ablaeva
- Department of Biology, University of Rochester, Rochester, NY 14627, USA
| | - Nancy Corson
- Department of Biology, University of Rochester, Rochester, NY 14627, USA
| | - Yongwook Choi
- Quantitative Medicine and Systems Biology Division, Translational Genomics Research Institute, Phoenix, AZ 85004, USA
| | - KayLene Y H Yamada
- Department of Biological Sciences, Auburn University, Auburn, AL 36849, USA
| | - Nicholas J Schork
- Quantitative Medicine and Systems Biology Division, Translational Genomics Research Institute, Phoenix, AZ 85004, USA
| | - Wendy R Hood
- Department of Biological Sciences, Auburn University, Auburn, AL 36849, USA
| | - Geoffrey E Hill
- Department of Biological Sciences, Auburn University, Auburn, AL 36849, USA
| | - Richard A Miller
- Department of Pathology and Geriatrics Center, University of Michigan, Ann Arbor, MI 48109, USA
| | - Andrei Seluanov
- Department of Biology, University of Rochester, Rochester, NY 14627, USA.
| | - Vera Gorbunova
- Department of Biology, University of Rochester, Rochester, NY 14627, USA.
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7
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Cosper PF, Copeland SE, Tucker JB, Weaver BA. Chromosome Missegregation as a Modulator of Radiation Sensitivity. Semin Radiat Oncol 2022; 32:54-63. [PMID: 34861996 PMCID: PMC8883596 DOI: 10.1016/j.semradonc.2021.09.002] [Citation(s) in RCA: 15] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/03/2023]
Abstract
Chromosome missegregation over the course of multiple cell divisions, termed chromosomal instability (CIN), is a hallmark of cancer. Multiple causes of CIN have been identified, including defects in the mitotic checkpoint, altered kinetochore-microtubule dynamics, centrosome amplification, and ionizing radiation. Here we review the types, mechanisms, and cellular implications of CIN. We discuss the evidence that CIN can promote tumors, suppress them, or do neither, depending on the rates of chromosome missegregration and the cellular context. Very high rates of chromosome missegregation lead to cell death due to loss of essential chromosomes; thus elevating CIN above a tolerable threshold provides a mechanistic opportunity to promote cancer cell death. Lethal rates of CIN can be achieved by a single insult or through a combination of insults. Because ionizing radiation induces CIN, additional therapies that increase CIN may serve as useful modulators of radiation sensitivity. Ultimately, quantifying the intrinsic CIN in a tumor and modulating this level pharmacologically as well as with radiation may allow for a more rational, personalized radiation therapy prescription, thereby decreasing side effects and increasing local control.
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Affiliation(s)
- Pippa F. Cosper
- Department of Human Oncology, University of Wisconsin-Madison, Madison, WI 53705, USA,University of Wisconsin Carbone Cancer Center, University of Wisconsin-Madison, Madison, WI 53705, USA
| | - Sarah E. Copeland
- Molecular & Cellular Pharmacology Graduate Training Program, University of Wisconsin-Madison, Madison, WI 53705, USA
| | - John B. Tucker
- Cancer Biology Graduate Training Program, University of Wisconsin-Madison, Madison, WI 53705, USA
| | - Beth A. Weaver
- University of Wisconsin Carbone Cancer Center, University of Wisconsin-Madison, Madison, WI 53705, USA,Department of Cellular and Regenerative Biology, University of Wisconsin-Madison, Madison, WI 53705, USA,Department of Oncology/McArdle Laboratory for Cancer Research, University of Wisconsin-Madison, Madison, WI 53705, USA,Corresponding author: Beth A. Weaver, University of Wisconsin-Madison, 1111 Highland Ave, 6109 WIMR Tower 1, Madison, WI 53705-2275, Phone: 608-263-5309, Fax: 608-265-6905,
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8
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Farina AR, Cappabianca LA, Zelli V, Sebastiano M, Mackay AR. Mechanisms involved in selecting and maintaining neuroblastoma cancer stem cell populations, and perspectives for therapeutic targeting. World J Stem Cells 2021; 13:685-736. [PMID: 34367474 PMCID: PMC8316860 DOI: 10.4252/wjsc.v13.i7.685] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/27/2021] [Revised: 03/09/2021] [Accepted: 04/14/2021] [Indexed: 02/06/2023] Open
Abstract
Pediatric neuroblastomas (NBs) are heterogeneous, aggressive, therapy-resistant embryonal tumours that originate from cells of neural crest (NC) origin and in particular neuroblasts committed to the sympathoadrenal progenitor cell lineage. Therapeutic resistance, post-therapeutic relapse and subsequent metastatic NB progression are driven primarily by cancer stem cell (CSC)-like subpopulations, which through their self-renewing capacity, intermittent and slow cell cycles, drug-resistant and reversibly adaptive plastic phenotypes, represent the most important obstacle to improving therapeutic outcomes in unfavourable NBs. In this review, dedicated to NB CSCs and the prospects for their therapeutic eradication, we initiate with brief descriptions of the unique transient vertebrate embryonic NC structure and salient molecular protagonists involved NC induction, specification, epithelial to mesenchymal transition and migratory behaviour, in order to familiarise the reader with the embryonic cellular and molecular origins and background to NB. We follow this by introducing NB and the potential NC-derived stem/progenitor cell origins of NBs, before providing a comprehensive review of the salient molecules, signalling pathways, mechanisms, tumour microenvironmental and therapeutic conditions involved in promoting, selecting and maintaining NB CSC subpopulations, and that underpin their therapy-resistant, self-renewing metastatic behaviour. Finally, we review potential therapeutic strategies and future prospects for targeting and eradication of these bastions of NB therapeutic resistance, post-therapeutic relapse and metastatic progression.
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Affiliation(s)
- Antonietta Rosella Farina
- Department of Applied Clinical and Biotechnological Sciences, University of L'Aquila, L'Aquila 67100, AQ, Italy
| | - Lucia Annamaria Cappabianca
- Department of Applied Clinical and Biotechnological Sciences, University of L'Aquila, L'Aquila 67100, AQ, Italy
| | - Veronica Zelli
- Department of Applied Clinical and Biotechnological Sciences, University of L'Aquila, L'Aquila 67100, AQ, Italy
| | - Michela Sebastiano
- Department of Applied Clinical and Biotechnological Sciences, University of L'Aquila, L'Aquila 67100, AQ, Italy
| | - Andrew Reay Mackay
- Department of Applied Clinical and Biotechnological Sciences, University of L'Aquila, L'Aquila 67100, AQ, Italy.
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9
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Etoposide Triggers Cellular Senescence by Inducing Multiple Centrosomes and Primary Cilia in Adrenocortical Tumor Cells. Cells 2021; 10:cells10061466. [PMID: 34208028 PMCID: PMC8230646 DOI: 10.3390/cells10061466] [Citation(s) in RCA: 16] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/21/2021] [Revised: 06/05/2021] [Accepted: 06/09/2021] [Indexed: 11/17/2022] Open
Abstract
Etoposide (ETO) has been used in treating adrenocortical tumor (ACT) cells. Our previous study showed that ETO inhibits ACT cell growth. In the present study, we show that ETO treatment at IC50 (10 μM) inhibited ACT cell growth by inducing cellular senescence rather than apoptosis. Several markers of cellular senescence, including enlarged nuclei, activated senescence-associated β-galactosidase activity, elevated levels of p53 and p21, and down-regulation of Lamin B1, were observed. We further found that ETO induced multiple centrosomes. The inhibition of multiple centrosomes accomplished by treating cells with either roscovitine or centrinone or through the overexpression of NR5A1/SF-1 alleviated ETO-induced senescence, suggesting that ETO triggered senescence via multiple centrosomes. Primary cilia also played a role in ETO-induced senescence. In the mechanism, DNA-PK-Chk2 signaling was activated by ETO treatment; inhibition of this signaling cascade alleviated multiple ETO-induced centrosomes and primary cilia followed by reducing cellular senescence. In addition to DNA damage signaling, autophagy was also triggered by ETO treatment for centrosomal events and senescence. Importantly, the inactivation of DNA-PK-Chk2 signaling reduced ETO-triggered autophagy; however, the inhibition of autophagy did not affect DNA-PK-Chk2 activation. Thus, ETO activated the DNA-PK-Chk2 cascade to facilitate autophagy. The activated autophagy further induced multiple centrosomes and primary cilia followed by triggering senescence.
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10
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Zhao JZ, Ye Q, Wang L, Lee SC. Centrosome amplification in cancer and cancer-associated human diseases. Biochim Biophys Acta Rev Cancer 2021; 1876:188566. [PMID: 33992724 DOI: 10.1016/j.bbcan.2021.188566] [Citation(s) in RCA: 21] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/22/2020] [Revised: 05/10/2021] [Accepted: 05/11/2021] [Indexed: 12/07/2022]
Abstract
Accumulated evidence from genetically modified cell and animal models indicates that centrosome amplification (CA) can initiate tumorigenesis with metastatic potential and enhance cell invasion. Multiple human diseases are associated with CA and carcinogenesis as well as metastasis, including infection with oncogenic viruses, type 2 diabetes, toxicosis by environmental pollution and inflammatory disease. In this review, we summarize (1) the evidence for the roles of CA in tumorigenesis and tumor cell invasion; (2) the association between diseases and carcinogenesis as well as metastasis; (3) the current knowledge of CA in the diseases; and (4) the signaling pathways of CA. We then give our own thinking and discuss perspectives relevant to CA in carcinogenesis and cancer metastasis in human diseases. In conclusion, investigations in this area might not only identify CA as a biological link between these diseases and the development of cancer but also prove the causal role of CA in cancer and progression under pathophysiological conditions, potentially taking cancer research into a new era.
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Affiliation(s)
- Ji Zhong Zhao
- Institute of Biomedical Sciences and School of Life Sciences, Jiangsu Normal University, Xuzhou, Jiangsu, PR China
| | - Qin Ye
- Institute of Biomedical Sciences and School of Life Sciences, Jiangsu Normal University, Xuzhou, Jiangsu, PR China
| | - Lan Wang
- School of Life Sciences, Shanxi University, Taiyuan, Shanxi, PR China
| | - Shao Chin Lee
- Institute of Biomedical Sciences and School of Life Sciences, Jiangsu Normal University, Xuzhou, Jiangsu, PR China.
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11
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Ma S, Rong Z, Liu C, Qin X, Zhang X, Chen Q. DNA damage promotes microtubule dynamics through a DNA-PK-AKT axis for enhanced repair. J Cell Biol 2021; 220:211656. [PMID: 33404607 PMCID: PMC7791344 DOI: 10.1083/jcb.201911025] [Citation(s) in RCA: 11] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/06/2019] [Revised: 11/01/2020] [Accepted: 12/02/2020] [Indexed: 01/09/2023] Open
Abstract
DNA double-strand breaks (DSBs) are mainly repaired by c-NHEJ and HR pathways. The enhanced DSB mobility after DNA damage is critical for efficient DSB repair. Although microtubule dynamics have been shown to regulate DSB mobility, the reverse effect of DSBs to microtubule dynamics remains elusive. Here, we uncovered a novel DSB-induced microtubule dynamics stress response (DMSR), which promotes DSB mobility and facilitates c-NHEJ repair. DMSR is accompanied by interphase centrosome maturation, which occurs in a DNA-PK-AKT-dependent manner. Depletion of PCM proteins attenuates DMSR and the mobility of DSBs, resulting in delayed c-NHEJ. Remarkably, DMSR occurs only in G1 or G0 cells and lasts around 6 h. Both inhibition of DNA-PK and depletion of 53BP1 abolish DMSR. Taken together, our study reveals a positive DNA repair mechanism in G1 or G0 cells in which DSBs actively promote microtubule dynamics and facilitate the c-NHEJ process.
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Affiliation(s)
- Shuyun Ma
- Department of Radiation and Medical Oncology, Medical Research Institute, Zhongnan Hospital of Wuhan University, Wuhan University, Wuhan, China,Frontier Science Center for Immunology and Metabolism, Medical Research Institute, Wuhan University, Wuhan, China
| | - Zeming Rong
- Department of Radiation and Medical Oncology, Medical Research Institute, Zhongnan Hospital of Wuhan University, Wuhan University, Wuhan, China,Frontier Science Center for Immunology and Metabolism, Medical Research Institute, Wuhan University, Wuhan, China
| | - Chen Liu
- Department of Radiation and Medical Oncology, Medical Research Institute, Zhongnan Hospital of Wuhan University, Wuhan University, Wuhan, China,Frontier Science Center for Immunology and Metabolism, Medical Research Institute, Wuhan University, Wuhan, China
| | - Xiaobing Qin
- Department of Radiation and Medical Oncology, Medical Research Institute, Zhongnan Hospital of Wuhan University, Wuhan University, Wuhan, China,Frontier Science Center for Immunology and Metabolism, Medical Research Institute, Wuhan University, Wuhan, China
| | - Xiaoyan Zhang
- College of Biomedicine and Health, Huazhong Agricultural University, Wuhan, China,College of Life Science and Technology, Huazhong Agricultural University, Wuhan, China
| | - Qiang Chen
- Department of Radiation and Medical Oncology, Medical Research Institute, Zhongnan Hospital of Wuhan University, Wuhan University, Wuhan, China,Frontier Science Center for Immunology and Metabolism, Medical Research Institute, Wuhan University, Wuhan, China,Correspondence to Qiang Chen:
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12
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Fan G, Sun L, Meng L, Hu C, Wang X, Shi Z, Hu C, Han Y, Yang Q, Cao L, Zhang X, Zhang Y, Song X, Xia S, He B, Zhang S, Wang C. The ATM and ATR kinases regulate centrosome clustering and tumor recurrence by targeting KIFC1 phosphorylation. Nat Commun 2021; 12:20. [PMID: 33397932 PMCID: PMC7782532 DOI: 10.1038/s41467-020-20208-x] [Citation(s) in RCA: 51] [Impact Index Per Article: 12.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/01/2020] [Accepted: 11/18/2020] [Indexed: 12/31/2022] Open
Abstract
Drug resistance and tumor recurrence are major challenges in cancer treatment. Cancer cells often display centrosome amplification. To maintain survival, cancer cells achieve bipolar division by clustering supernumerary centrosomes. Targeting centrosome clustering is therefore considered a promising therapeutic strategy. However, the regulatory mechanisms of centrosome clustering remain unclear. Here we report that KIFC1, a centrosome clustering regulator, is positively associated with tumor recurrence. Under DNA damaging treatments, the ATM and ATR kinases phosphorylate KIFC1 at Ser26 to selectively maintain the survival of cancer cells with amplified centrosomes via centrosome clustering, leading to drug resistance and tumor recurrence. Inhibition of KIFC1 phosphorylation represses centrosome clustering and tumor recurrence. This study identified KIFC1 as a prognostic tumor recurrence marker, and revealed that tumors can acquire therapeutic resistance and recurrence via triggering centrosome clustering under DNA damage stresses, suggesting that blocking KIFC1 phosphorylation may open a new vista for cancer therapy.
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Affiliation(s)
- Guangjian Fan
- Translational Medicine Center, Shanghai General Hospital, Shanghai Jiao Tong University School of Medicine, 201620, Shanghai, China
| | - Lianhui Sun
- Translational Medicine Center, Shanghai General Hospital, Shanghai Jiao Tong University School of Medicine, 201620, Shanghai, China
| | - Ling Meng
- Department of Pulmonary and Critical Care Medicine, The Second Affiliated Hospital of Shandong First Medical University, 271000, Shandong, China
| | - Chen Hu
- Translational Medicine Center, Shanghai General Hospital, Shanghai Jiao Tong University School of Medicine, 201620, Shanghai, China
| | - Xing Wang
- Translational Medicine Center, Shanghai General Hospital, Shanghai Jiao Tong University School of Medicine, 201620, Shanghai, China
| | - Zhan Shi
- Translational Medicine Center, Shanghai General Hospital, Shanghai Jiao Tong University School of Medicine, 201620, Shanghai, China
| | - Congli Hu
- Translational Medicine Center, Shanghai General Hospital, Shanghai Jiao Tong University School of Medicine, 201620, Shanghai, China
| | - Yang Han
- Translational Medicine Center, Shanghai General Hospital, Shanghai Jiao Tong University School of Medicine, 201620, Shanghai, China
| | - Qingqing Yang
- Translational Medicine Center, Shanghai General Hospital, Shanghai Jiao Tong University School of Medicine, 201620, Shanghai, China
| | - Liu Cao
- Key Laboratory of Medical Cell Biology, College of Translational Medicine, China Medical University, 110000, Shenyang, China
| | - Xiaohong Zhang
- Department of Oncology, Karmanos Cancer Institute, Wayne State University School of Medicine, 4100 John R., Detroit, MI, 48201, USA
| | - Yan Zhang
- Department of Hematology, Shanghai General Hospital, Shanghai Jiao Tong University School of Medicine, 201620, Shanghai, China
| | - Xianmin Song
- Department of Hematology, Shanghai General Hospital, Shanghai Jiao Tong University School of Medicine, 201620, Shanghai, China
| | - Shujie Xia
- Department of Urology, Shanghai General Hospital, Shanghai Jiao Tong University School of Medicine; Institute of Urology, Shanghai Jiao Tong University, 200080, Shanghai, China
| | - Baokun He
- Department of Gastroenterology, Shanghai General Hospital, Shanghai Jiao Tong University School of Medicine, 201620, Shanghai, China
| | - Shengping Zhang
- Translational Medicine Center, Shanghai General Hospital, Shanghai Jiao Tong University School of Medicine, 201620, Shanghai, China.
| | - Chuangui Wang
- Translational Medicine Center, Shanghai General Hospital, Shanghai Jiao Tong University School of Medicine, 201620, Shanghai, China.
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13
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Wang CY, Tsai SW, Chien HH, Chen TY, Sheu SY, So EC, Huang BM. Cordycepin Inhibits Human Gestational Choriocarcinoma Cell Growth by Disrupting Centrosome Homeostasis. DRUG DESIGN DEVELOPMENT AND THERAPY 2020; 14:2987-3000. [PMID: 32801639 PMCID: PMC7394508 DOI: 10.2147/dddt.s252401] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 03/04/2020] [Accepted: 07/06/2020] [Indexed: 12/19/2022]
Abstract
Introduction Human gestational choriocarcinoma, a type of gestational trophoblastic disease, occurs after miscarriage, abortion, ectopic pregnancy, or molar pregnancy. Despite recent advances in the mechanism of anticancer drugs that induce human gestational choriocarcinoma apoptosis or block its growth, new therapeutic approaches are needed to be established. Cordycepin is an active anti-cancer component extracted from Cordyceps sinensis. It prevents cell proliferation both in vitro and in vivo. Materials and Methods Here, we examined cell growth by counting cell numbers, and performing a flow cytometry assay and EdU incorporation assay. Centrosome and cytoskeleton-related structures were observed by immunofluorescence assay. The DNA damage-related signaling was examined by Western blot assay. Results Here, we showed that cordycepin inhibited human gestational choriocarcinoma cell proliferation and induced cell death. In addition, treatment with cordycepin activated DNA-PK and ERK, thus inducing centrosome amplification and aberrant mitosis. These amplified centrosomes also disrupted microtubule arrays and actin networks, thus leading to defective cell adhesion. Furthermore, cordycepin induced autophagy for triggering cell death. Conclusion Thus, our study demonstrates that cordycepin inhibits cell proliferation and disrupts the cytoskeleton by triggering centrosome amplification.
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Affiliation(s)
- Chia-Yih Wang
- Department of Cell Biology and Anatomy, College of Medicine, National Cheng Kung University, Tainan, Taiwan.,Institute of Basic Medical Sciences, College of Medicine, National Cheng Kung University, Tainan, Taiwan
| | - Shih-Wei Tsai
- Department of Obstetrics and Gynecology, An Nan Hospital, China Medical University, Tainan, Taiwan
| | - Han-Hsiang Chien
- Department of Cell Biology and Anatomy, College of Medicine, National Cheng Kung University, Tainan, Taiwan
| | - Ting-Yu Chen
- Institute of Basic Medical Sciences, College of Medicine, National Cheng Kung University, Tainan, Taiwan.,Institute of Biomedical Sciences, Academia Sinica, Taipei, Taiwan
| | - Shi-Yuan Sheu
- School of Chinese Medicine for Post-Baccalaureate, I-Shou University, Kaohsiung, Taiwan.,Department of Chinese Medicine, E-Da Cancer Hospital, Kaohsiung, Taiwan
| | - Edmund Cheung So
- Department of Anesthesia & Medical Research, An Nan Hospital, China Medical University, Tainan, Taiwan.,Graduate Institute of Medical Sciences, Chang Jung Christian University Tainan, Tainan, Taiwan
| | - Bu-Miin Huang
- Department of Cell Biology and Anatomy, College of Medicine, National Cheng Kung University, Tainan, Taiwan.,Institute of Medical Research, China Medical University, Taichung, Taiwan
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14
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Modulation of DNA Damage Response by Sphingolipid Signaling: An Interplay that Shapes Cell Fate. Int J Mol Sci 2020; 21:ijms21124481. [PMID: 32599736 PMCID: PMC7349968 DOI: 10.3390/ijms21124481] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/16/2020] [Revised: 05/07/2020] [Accepted: 05/08/2020] [Indexed: 12/11/2022] Open
Abstract
Although once considered as structural components of eukaryotic biological membranes, research in the past few decades hints at a major role of bioactive sphingolipids in mediating an array of physiological processes including cell survival, proliferation, inflammation, senescence, and death. A large body of evidence points to a fundamental role for the sphingolipid metabolic pathway in modulating the DNA damage response (DDR). The interplay between these two elements of cell signaling determines cell fate when cells are exposed to metabolic stress or ionizing radiation among other genotoxic agents. In this review, we aim to dissect the mediators of the DDR and how these interact with the different sphingolipid metabolites to mount various cellular responses.
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15
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Fujimoto M, Bo T, Yamamoto K, Yasui H, Yamamori T, Inanami O. Radiation-induced abnormal centrosome amplification and mitotic catastrophe in human cervical tumor HeLa cells and murine mammary tumor EMT6 cells. J Clin Biochem Nutr 2020; 67:240-247. [PMID: 33293764 PMCID: PMC7705082 DOI: 10.3164/jcbn.19-80] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/31/2019] [Accepted: 02/18/2020] [Indexed: 12/14/2022] Open
Abstract
Mitotic catastrophe is a form of cell death linked to aberrant mitosis caused by improper or uncoordinated mitotic progression. Abnormal centrosome amplification and mitotic catastrophe occur simultaneously, and some cells with amplified centrosomes enter aberrant mitosis, but it is not clear whether abnormal centrosome amplification triggers mitotic catastrophe. Here, to investigate whether radiation-induced abnormal centrosome amplification is essential for induction of radiation-induced mitotic catastrophe, centrinone-B, a highly selective inhibitor of polo-like kinase 4, was utilized to inhibit centrosome amplification, since polo-like kinase 4 is an essential kinase in centrosome duplication. When human cervical tumor HeLa cells and murine mammary tumor EMT6 cells were irradiated with 2.5 Gy of X-rays, cells with morphological features of mitotic catastrophe and the number of cells having >2 centrosomes increased in both cell lines. Although centrinone-B significantly inhibited radiation-induced abnormal centrosome amplification in both cell lines, such treatment did not change cell growth and significantly enhanced mitotic catastrophe in HeLa cells exposed to X-rays. In contrast, inhibition of centrosome amplification reduced cell growth and mitotic catastrophe in EMT6 cells exposed to X-rays. These results indicated that the role of radiation-induced abnormal centrosome amplification in radiation-induced mitotic catastrophe changes, depending on the cell type.
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Affiliation(s)
- Masaki Fujimoto
- Laboratory of Radiation Biology, Department of Applied Veterinary Sciences, Faculty of Veterinary Medicine, Hokkaido University, Sapporo 060-0818, Japan
| | - Tomoki Bo
- Laboratory of Radiation Biology, Department of Applied Veterinary Sciences, Faculty of Veterinary Medicine, Hokkaido University, Sapporo 060-0818, Japan
| | - Kumiko Yamamoto
- Laboratory of Radiation Biology, Department of Applied Veterinary Sciences, Faculty of Veterinary Medicine, Hokkaido University, Sapporo 060-0818, Japan
| | - Hironobu Yasui
- Laboratory of Radiation Biology, Department of Applied Veterinary Sciences, Faculty of Veterinary Medicine, Hokkaido University, Sapporo 060-0818, Japan
| | - Tohru Yamamori
- Laboratory of Radiation Biology, Department of Applied Veterinary Sciences, Faculty of Veterinary Medicine, Hokkaido University, Sapporo 060-0818, Japan
| | - Osamu Inanami
- Laboratory of Radiation Biology, Department of Applied Veterinary Sciences, Faculty of Veterinary Medicine, Hokkaido University, Sapporo 060-0818, Japan
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16
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Wang L, Han S, Zhu J, Wang X, Li Y, Wang Z, Lin E, Wang X, Molkentine DP, Blanchard P, Yang Y, Zhang R, Sahoo N, Gillin M, Zhu XR, Zhang X, Myers JN, Frank SJ. Proton versus photon radiation-induced cell death in head and neck cancer cells. Head Neck 2018; 41:46-55. [PMID: 30561022 DOI: 10.1002/hed.25357] [Citation(s) in RCA: 23] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/11/2017] [Revised: 04/04/2018] [Accepted: 05/16/2018] [Indexed: 12/17/2022] Open
Abstract
BACKGROUND Photon (X-ray) radiotherapy (XRT) kills cells via DNA damage, however, how proton radiotherapy (PRT) causes cell death in head and neck squamous cell carcinoma (HNSCC) is unclear. We investigated mechanisms of HNSCC cell death after XRT versus PRT. METHODS We assessed type of death in 2 human papillomavirus (HPV)-positive and two HPV-negative cell lines: necrosis and apoptosis (Annexin-V fluorescein isothiocyanate [FITC]); senescence (β-galactosidase); and mitotic catastrophe (γ-tubulin and diamidino-phenylindole [DAPI]). RESULTS The XRT-induced or PRT-induced cellular senescence and mitotic catastrophe in all cell lines studied suggested that PRT caused cell death to a greater extent than XRT. After PRT, mitotic catastrophe peaked in HPV-negative and HPV-positive cells at 48 and 72 hours, respectively. No obvious differences were noted in the extent of cell necrosis or apoptosis after XRT versus PRT. CONCLUSION Under the conditions and in the cell lines reported here, mitotic catastrophe and senescence were the major types of cell death induced by XRT and PRT, and PRT may be more effective.
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Affiliation(s)
- Li Wang
- Department of Experimental Radiation Oncology, The University of Texas MD Anderson Cancer Center, Houston, Texas
| | - Shichao Han
- Department of Experimental Radiation Oncology, The University of Texas MD Anderson Cancer Center, Houston, Texas.,Department of Gynecology, The Second Affiliated Hospital of Dalian Medical University, Dalian, Liaoning, China
| | - Jinming Zhu
- Department of Experimental Radiation Oncology, The University of Texas MD Anderson Cancer Center, Houston, Texas.,Department of Radiation Oncology, The Affiliated Zhongshan Hospital of Dalian University, Dalian, Liaoning, China
| | - Xiaochun Wang
- Department of Radiation Physics, The University of Texas MD Anderson Cancer Center, Houston, Texas
| | - Yuting Li
- Department of Radiation Physics, The University of Texas MD Anderson Cancer Center, Houston, Texas
| | - Zeming Wang
- Department of Radiation Physics, The University of Texas MD Anderson Cancer Center, Houston, Texas
| | - Eric Lin
- Department of Experimental Radiation Oncology, The University of Texas MD Anderson Cancer Center, Houston, Texas
| | - Xiaofang Wang
- Department of Experimental Radiation Oncology, The University of Texas MD Anderson Cancer Center, Houston, Texas
| | - David P Molkentine
- Department of Experimental Radiation Oncology, The University of Texas MD Anderson Cancer Center, Houston, Texas
| | - Pierre Blanchard
- Department of Radiation Oncology, The University of Texas MD Anderson Cancer Center, Houston, Texas.,Department of Radiation Oncology, Institut Gustave Roussy, Villejuif, France
| | - Yining Yang
- Department of Radiation Physics, The University of Texas MD Anderson Cancer Center, Houston, Texas
| | - Ruiping Zhang
- Department of Radiation Physics, The University of Texas MD Anderson Cancer Center, Houston, Texas
| | - Narayan Sahoo
- Department of Radiation Physics, The University of Texas MD Anderson Cancer Center, Houston, Texas
| | - Michael Gillin
- Department of Radiation Physics, The University of Texas MD Anderson Cancer Center, Houston, Texas
| | - Xiaorong Ronald Zhu
- Department of Radiation Physics, The University of Texas MD Anderson Cancer Center, Houston, Texas
| | - Xiaodong Zhang
- Department of Radiation Physics, The University of Texas MD Anderson Cancer Center, Houston, Texas
| | - Jeffrey N Myers
- Department of Head and Neck Surgery, The University of Texas MD Anderson Cancer Center, Houston, Texas
| | - Steven J Frank
- Department of Radiation Oncology, The University of Texas MD Anderson Cancer Center, Houston, Texas
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17
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Wang Z, Grosskurth SE, Cheung T, Petteruti P, Zhang J, Wang X, Wang W, Gharahdaghi F, Wu J, Su N, Howard RT, Mayo M, Widzowski D, Scott DA, Johannes JW, Lamb ML, Lawson D, Dry JR, Lyne PD, Tate EW, Zinda M, Mikule K, Fawell SE, Reimer C, Chen H. Pharmacological Inhibition of PARP6 Triggers Multipolar Spindle Formation and Elicits Therapeutic Effects in Breast Cancer. Cancer Res 2018; 78:6691-6702. [PMID: 30297535 DOI: 10.1158/0008-5472.can-18-1362] [Citation(s) in RCA: 24] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/02/2018] [Revised: 08/23/2018] [Accepted: 09/26/2018] [Indexed: 11/16/2022]
Abstract
: PARP proteins represent a class of post-translational modification enzymes with diverse cellular functions. Targeting PARPs has proven to be efficacious clinically, but exploration of the therapeutic potential of PARP inhibition has been limited to targeting poly(ADP-ribose) generating PARP, including PARP1/2/3 and tankyrases. The cancer-related functions of mono(ADP-ribose) generating PARP, including PARP6, remain largely uncharacterized. Here, we report a novel therapeutic strategy targeting PARP6 using the first reported PARP6 inhibitors. By screening a collection of PARP compounds for their ability to induce mitotic defects, we uncovered a robust correlation between PARP6 inhibition and induction of multipolar spindle (MPS) formation, which was phenocopied by PARP6 knockdown. Treatment with AZ0108, a PARP6 inhibitor with a favorable pharmacokinetic profile, potently induced the MPS phenotype, leading to apoptosis in a subset of breast cancer cells in vitro and antitumor effects in vivo. In addition, Chk1 was identified as a specific substrate of PARP6 and was further confirmed by enzymatic assays and by mass spectrometry. Furthermore, when modification of Chk1 was inhibited with AZ0108 in breast cancer cells, we observed marked upregulation of p-S345 Chk1 accompanied by defects in mitotic signaling. Together, these results establish proof-of-concept antitumor efficacy through PARP6 inhibition and highlight a novel function of PARP6 in maintaining centrosome integrity via direct ADP-ribosylation of Chk1 and modulation of its activity. SIGNIFICANCE: These findings describe a new inhibitor of PARP6 and identify a novel function of PARP6 in regulating activation of Chk1 in breast cancer cells.
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Affiliation(s)
- Zebin Wang
- Oncology, IMED Biotech Unit, AstraZeneca R&D Boston, Waltham, Massachusetts
| | - Shaun E Grosskurth
- Oncology, IMED Biotech Unit, AstraZeneca R&D Boston, Waltham, Massachusetts
| | - Tony Cheung
- Oncology, IMED Biotech Unit, AstraZeneca R&D Boston, Waltham, Massachusetts
| | - Philip Petteruti
- Oncology, IMED Biotech Unit, AstraZeneca R&D Boston, Waltham, Massachusetts
| | - Jingwen Zhang
- Oncology, IMED Biotech Unit, AstraZeneca R&D Boston, Waltham, Massachusetts
| | - Xin Wang
- Oncology, IMED Biotech Unit, AstraZeneca R&D Boston, Waltham, Massachusetts
| | - Wenxian Wang
- Oncology, IMED Biotech Unit, AstraZeneca R&D Boston, Waltham, Massachusetts
| | - Farzin Gharahdaghi
- Oncology, IMED Biotech Unit, AstraZeneca R&D Boston, Waltham, Massachusetts
| | - Jiaquan Wu
- Oncology, IMED Biotech Unit, AstraZeneca R&D Boston, Waltham, Massachusetts
| | - Nancy Su
- Oncology, IMED Biotech Unit, AstraZeneca R&D Boston, Waltham, Massachusetts
| | - Ryan T Howard
- Institute of Chemical Biology, Department of Chemistry, Imperial College London, London, United Kingdom
| | - Michele Mayo
- Oncology, IMED Biotech Unit, AstraZeneca R&D Boston, Waltham, Massachusetts
| | - Dan Widzowski
- Oncology, IMED Biotech Unit, AstraZeneca R&D Boston, Waltham, Massachusetts
| | - David A Scott
- Oncology, IMED Biotech Unit, AstraZeneca R&D Boston, Waltham, Massachusetts
| | - Jeffrey W Johannes
- Oncology, IMED Biotech Unit, AstraZeneca R&D Boston, Waltham, Massachusetts
| | - Michelle L Lamb
- Oncology, IMED Biotech Unit, AstraZeneca R&D Boston, Waltham, Massachusetts
| | - Deborah Lawson
- Oncology, IMED Biotech Unit, AstraZeneca R&D Boston, Waltham, Massachusetts
| | - Jonathan R Dry
- Oncology, IMED Biotech Unit, AstraZeneca R&D Boston, Waltham, Massachusetts
| | - Paul D Lyne
- Oncology, IMED Biotech Unit, AstraZeneca R&D Boston, Waltham, Massachusetts
| | - Edward W Tate
- Institute of Chemical Biology, Department of Chemistry, Imperial College London, London, United Kingdom
| | - Michael Zinda
- Oncology, IMED Biotech Unit, AstraZeneca R&D Boston, Waltham, Massachusetts
| | - Keith Mikule
- Oncology, IMED Biotech Unit, AstraZeneca R&D Boston, Waltham, Massachusetts
| | - Stephen E Fawell
- Oncology, IMED Biotech Unit, AstraZeneca R&D Boston, Waltham, Massachusetts
| | - Corinne Reimer
- Oncology, IMED Biotech Unit, AstraZeneca R&D Boston, Waltham, Massachusetts
| | - Huawei Chen
- Oncology, IMED Biotech Unit, AstraZeneca R&D Boston, Waltham, Massachusetts.
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18
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Hossain D, Ferreira Barbosa JA, Cohen ÉA, Tsang WY. HIV-1 Vpr hijacks EDD-DYRK2-DDB1 DCAF1 to disrupt centrosome homeostasis. J Biol Chem 2018; 293:9448-9460. [PMID: 29724823 PMCID: PMC6005440 DOI: 10.1074/jbc.ra117.001444] [Citation(s) in RCA: 20] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/13/2017] [Revised: 04/17/2018] [Indexed: 11/06/2022] Open
Abstract
Viruses exploit the host cell machinery for their own profit. To evade innate immune sensing and promote viral replication, HIV type 1 (HIV-1) subverts DNA repair regulatory proteins and induces G2/M arrest. The preintegration complex of HIV-1 is known to traffic along microtubules and accumulate near the microtubule-organizing center. The centrosome is the major microtubule-organizing center in most eukaryotic cells, but precisely how HIV-1 impinges on centrosome biology remains poorly understood. We report here that the HIV-1 accessory protein viral protein R (Vpr) localized to the centrosome through binding to DCAF1, forming a complex with the ubiquitin ligase EDD-DYRK2-DDB1DCAF1 and Cep78, a resident centrosomal protein previously shown to inhibit EDD-DYRK2-DDB1DCAF1 Vpr did not affect ubiquitination of Cep78. Rather, it enhanced ubiquitination of an EDD-DYRK2-DDB1DCAF1 substrate, CP110, leading to its degradation, an effect that could be overcome by Cep78 expression. The down-regulation of CP110 and elongation of centrioles provoked by Vpr were independent of G2/M arrest. Infection of T lymphocytes with HIV-1, but not with HIV-1 lacking Vpr, promoted CP110 degradation and centriole elongation. Elongated centrioles recruited more γ-tubulin to the centrosome, resulting in increased microtubule nucleation. Our results suggest that Vpr is targeted to the centrosome where it hijacks a ubiquitin ligase, disrupting organelle homeostasis, which may contribute to HIV-1 pathogenesis.
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Affiliation(s)
- Delowar Hossain
- From the Institut de recherches cliniques de Montréal, Montreal, Quebec H2W 1R7, Canada
- the Division of Experimental Medicine, McGill University, Montreal, Quebec H4A 3J1, Canada
| | | | - Éric A Cohen
- From the Institut de recherches cliniques de Montréal, Montreal, Quebec H2W 1R7, Canada
- the Division of Experimental Medicine, McGill University, Montreal, Quebec H4A 3J1, Canada
- the Department of Microbiology, Infectiology, and Immunology, Université de Montréal, Montreal, Quebec H3C 3J7, Canada, and
| | - William Y Tsang
- From the Institut de recherches cliniques de Montréal, Montreal, Quebec H2W 1R7, Canada,
- the Division of Experimental Medicine, McGill University, Montreal, Quebec H4A 3J1, Canada
- the Department of Pathology and Cell Biology, Université de Montréal, Montreal, Quebec H3C 3J7, Canada
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19
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Prakash A, Garcia-Moreno JF, Brown JAL, Bourke E. Clinically Applicable Inhibitors Impacting Genome Stability. Molecules 2018; 23:E1166. [PMID: 29757235 PMCID: PMC6100577 DOI: 10.3390/molecules23051166] [Citation(s) in RCA: 19] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/21/2018] [Revised: 04/27/2018] [Accepted: 05/01/2018] [Indexed: 12/14/2022] Open
Abstract
Advances in technology have facilitated the molecular profiling (genomic and transcriptomic) of tumours, and has led to improved stratification of patients and the individualisation of treatment regimes. To fully realize the potential of truly personalised treatment options, we need targeted therapies that precisely disrupt the compensatory pathways identified by profiling which allow tumours to survive or gain resistance to treatments. Here, we discuss recent advances in novel therapies that impact the genome (chromosomes and chromatin), pathways targeted and the stage of the pathways targeted. The current state of research will be discussed, with a focus on compounds that have advanced into trials (clinical and pre-clinical). We will discuss inhibitors of specific DNA damage responses and other genome stability pathways, including those in development, which are likely to synergistically combine with current therapeutic options. Tumour profiling data, combined with the knowledge of new treatments that affect the regulation of essential tumour signalling pathways, is revealing fundamental insights into cancer progression and resistance mechanisms. This is the forefront of the next evolution of advanced oncology medicine that will ultimately lead to improved survival and may, one day, result in many cancers becoming chronic conditions, rather than fatal diseases.
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Affiliation(s)
- Anu Prakash
- Discipline of Pathology, Lambe Institute for Translational Research, School of Medicine, National University of Ireland Galway, H91 YR71 Galway, Ireland.
| | - Juan F Garcia-Moreno
- Discipline of Surgery, Lambe Institute for Translational Research, School of Medicine, National University of Ireland Galway, H91 YR71 Galway, Ireland.
| | - James A L Brown
- Discipline of Surgery, Lambe Institute for Translational Research, School of Medicine, National University of Ireland Galway, H91 YR71 Galway, Ireland.
| | - Emer Bourke
- Discipline of Pathology, Lambe Institute for Translational Research, School of Medicine, National University of Ireland Galway, H91 YR71 Galway, Ireland.
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20
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Rogne M, Svaerd O, Madsen-Østerbye J, Hashim A, Tjønnfjord GE, Staerk J. Cytokinesis arrest and multiple centrosomes in B cell chronic lymphocytic leukaemia. J Cell Mol Med 2018. [PMID: 29516674 PMCID: PMC5908127 DOI: 10.1111/jcmm.13579] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/13/2022] Open
Abstract
Cytokinesis failure leads to the emergence of tetraploid cells and multiple centrosomes. Chronic lymphocytic leukaemia (CLL) is the most common haematological malignancy in adults and is characterized by clonal B cell expansion. Here, we show that a significant number of peripheral blood CLL cells are arrested in cytokinesis and that this event occurred after nuclear envelope reformation and before cytoplasmic abscission. mRNA expression data showed that several genes known to be crucial for cell cycle regulation, checkpoint and centromere function, such as ING4, ING5, CDKN1A and CDK4, were significantly dysregulated in CLL samples. Our results demonstrate that CLL cells exhibit difficulties in completing mitosis, which is different from but may, at least in part, explain the previously reported accumulation of CLL cells in G0/1.
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Affiliation(s)
- Marie Rogne
- Centre for Molecular Medicine Norway, Nordic European Molecular Laboratory Partnership, University of Oslo, Oslo, Norway
| | - Oksana Svaerd
- Centre for Molecular Medicine Norway, Nordic European Molecular Laboratory Partnership, University of Oslo, Oslo, Norway
| | - Julia Madsen-Østerbye
- Centre for Molecular Medicine Norway, Nordic European Molecular Laboratory Partnership, University of Oslo, Oslo, Norway
| | - Adnan Hashim
- Centre for Molecular Medicine Norway, Nordic European Molecular Laboratory Partnership, University of Oslo, Oslo, Norway
| | - Geir E Tjønnfjord
- Department of Haematology, Oslo University Hospital, Oslo, Norway.,Institute of Clinical Medicine, University of Oslo, Oslo, Norway
| | - Judith Staerk
- Centre for Molecular Medicine Norway, Nordic European Molecular Laboratory Partnership, University of Oslo, Oslo, Norway.,Department of Haematology, Oslo University Hospital, Oslo, Norway.,Norwegian Center for Stem Cell Research, Department of Immunology, Oslo University Hospital, Oslo, Norway
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21
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Tšuiko O, Jatsenko T, Parameswaran Grace LK, Kurg A, Vermeesch JR, Lanner F, Altmäe S, Salumets A. A speculative outlook on embryonic aneuploidy: Can molecular pathways be involved? Dev Biol 2018; 447:3-13. [PMID: 29391166 DOI: 10.1016/j.ydbio.2018.01.014] [Citation(s) in RCA: 25] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/17/2017] [Revised: 12/27/2017] [Accepted: 01/22/2018] [Indexed: 01/21/2023]
Abstract
The journey of embryonic development starts at oocyte fertilization, which triggers a complex cascade of events and cellular pathways that guide early embryogenesis. Recent technological advances have greatly expanded our knowledge of cleavage-stage embryo development, which is characterized by an increased rate of whole-chromosome losses and gains, mixoploidy, and atypical cleavage morphokinetics. Embryonic aneuploidy significantly contributes to implantation failure, spontaneous miscarriage, stillbirth or congenital birth defects in both natural and assisted human reproduction. Essentially, early embryo development is strongly determined by maternal factors. Owing to considerable limitations associated with human oocyte and embryo research, the use of animal models is inevitable. However, cellular and molecular mechanisms driving the error-prone early stages of development are still poorly described. In this review, we describe known events that lead to aneuploidy in mammalian oocytes and preimplantation embryos. As the processes of oocyte and embryo development are rigorously regulated by multiple signal-transduction pathways, we explore the putative role of signaling pathways in genomic integrity maintenance. Based on the existing evidence from human and animal data, we investigate whether critical early developmental pathways, like Wnt, Hippo and MAPK, together with distinct DNA damage response and DNA repair pathways can be associated with embryo genomic instability, a question that has, so far, remained largely unexplored.
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Affiliation(s)
- Olga Tšuiko
- Department of Biomedicine, Institute of Bio- and Translational Medicine, University of Tartu, Tartu 50411, Estonia; Competence Centre on Health Technologies, Tartu 50410, Estonia
| | | | - Lalit Kumar Parameswaran Grace
- Department of Women's and Children's Health, Division of Obstetrics and Gynecology, Karolinska Institutet, Karolinska University Hospital, Stockholm 17176, Sweden
| | - Ants Kurg
- Department of Biotechnology, Institute of Molecular and Cell Biology, University of Tartu, Tartu 51010, Estonia
| | - Joris Robert Vermeesch
- Laboratory of Cytogenetics and Genome Research, Center of Human Genetics, KU Leuven, Leuven 3000, Belgium
| | - Fredrik Lanner
- Department of Clinical Science, Intervention, and Technology, Karolinska Institutet, Stockholm 14186, Sweden
| | - Signe Altmäe
- Competence Centre on Health Technologies, Tartu 50410, Estonia; Department of Biochemistry and Molecular Biology, Faculty of Sciences, University of Granada, Granada 18071, Spain.
| | - Andres Salumets
- Department of Biomedicine, Institute of Bio- and Translational Medicine, University of Tartu, Tartu 50411, Estonia; Competence Centre on Health Technologies, Tartu 50410, Estonia; Department of Obstetrics and Gynecology, Institute of Clinical Medicine, University of Tartu, Tartu 51014, Estonia; Department of Obstetrics and Gynecology, University of Helsinki and Helsinki University Hospital, Helsinki 00029, Finland
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22
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Patwardhan D, Mani S, Passemard S, Gressens P, El Ghouzzi V. STIL balancing primary microcephaly and cancer. Cell Death Dis 2018; 9:65. [PMID: 29352115 PMCID: PMC5833631 DOI: 10.1038/s41419-017-0101-9] [Citation(s) in RCA: 23] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/29/2017] [Revised: 10/04/2017] [Accepted: 10/23/2017] [Indexed: 11/25/2022]
Abstract
Cell division and differentiation are two fundamental physiological processes that need to be tightly balanced to achieve harmonious development of an organ or a tissue without jeopardizing its homeostasis. The role played by the centriolar protein STIL is highly illustrative of this balance at different stages of life as deregulation of the human STIL gene expression has been associated with either insufficient brain development (primary microcephaly) or cancer, two conditions resulting from perturbations in cell cycle and chromosomal segregation. This review describes the recent advances on STIL functions in the control of centriole duplication and mitotic spindle integrity, and discusses how pathological perturbations of its finely tuned expression result in chromosomal instability in both embryonic and postnatal situations, highlighting the concept that common key factors are involved in developmental steps and tissue homeostasis.
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Affiliation(s)
- Dhruti Patwardhan
- PROTECT, INSERM, Université Paris Diderot, Sorbonne Paris Cité, Paris, France
- Centre for Neuroscience, IISC Bangalore, India
| | - Shyamala Mani
- PROTECT, INSERM, Université Paris Diderot, Sorbonne Paris Cité, Paris, France
- Curadev Pharma, B 87, Sector 83, Noida, UP, 201305,, India
| | - Sandrine Passemard
- PROTECT, INSERM, Université Paris Diderot, Sorbonne Paris Cité, Paris, France
- AP HP, Hôpital Robert Debré, Service de Génétique Clinique, Paris, France
| | - Pierre Gressens
- PROTECT, INSERM, Université Paris Diderot, Sorbonne Paris Cité, Paris, France
- Centre for the Developing Brain, Division of Imaging Sciences and Biomedical Engineering, King's College London, King's Health Partners, St. Thomas' Hospital, London, UK
| | - Vincent El Ghouzzi
- PROTECT, INSERM, Université Paris Diderot, Sorbonne Paris Cité, Paris, France.
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23
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Suzuki M, Yamamori T, Bo T, Sakai Y, Inanami O. MK-8776, a novel Chk1 inhibitor, exhibits an improved radiosensitizing effect compared to UCN-01 by exacerbating radiation-induced aberrant mitosis. Transl Oncol 2017; 10:491-500. [PMID: 28550769 PMCID: PMC5447387 DOI: 10.1016/j.tranon.2017.04.002] [Citation(s) in RCA: 23] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/24/2017] [Revised: 03/27/2017] [Accepted: 04/03/2017] [Indexed: 11/08/2022] Open
Abstract
Checkpoint kinase 1 (Chk1) is an evolutionarily conserved serine/threonine kinase that plays an important role in G2/M checkpoint signaling. Here, we evaluate the radiosensitizing effects of a novel selective Chk1 inhibitor MK-8776, comparing its efficacy with a first-generation Chk1 inhibitor UCN-01, and attempt to elucidate the mechanism of radiosensitization. In a clonogenic survival assay, MK-8776 demonstrated a more pronounced radiosensitizing effect than UCN-01, with lower cytotoxicity. Importantly, radiosensitization by MK-8776 can be achieved at doses as low as 2.5 Gy, which is a clinically applicable irradiation dose. MK-8776, but not UCN-01, exacerbated mitotic catastrophe (MC) and centrosome abnormalities, without affecting repair kinetics of DNA double strand breaks. Furthermore, live-cell imaging revealed that MK-8776 significantly abrogated the radiation-induced G2/M checkpoint, prolonged the mitotic phase, and enhanced aberrant mitosis. This suggests that Chk1 inhibition by MK-8776 activates a spindle assembly checkpoint and increases mitotic defects in irradiated EMT6 cells. In conclusion, we have shown that, at minimally toxic concentrations, MK-8776 enhances radiation-induced cell death through the enhancement of aberrant mitosis and MC, without affecting DNA damage repair.
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24
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Flanagan AM, Stavenschi E, Basavaraju S, Gaboriau D, Hoey DA, Morrison CG. Centriole splitting caused by loss of the centrosomal linker protein C-NAP1 reduces centriolar satellite density and impedes centrosome amplification. Mol Biol Cell 2017; 28:736-745. [PMID: 28100636 PMCID: PMC5349781 DOI: 10.1091/mbc.e16-05-0325] [Citation(s) in RCA: 23] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/27/2016] [Revised: 01/10/2017] [Accepted: 01/11/2017] [Indexed: 12/24/2022] Open
Abstract
Duplication of the centrosomes is a tightly regulated process. Abnormal centrosome numbers can impair cell division and cause changes in how cells migrate. Duplicated centrosomes are held together by a proteinaceous linker made up of rootletin filaments anchored to the centrioles by C-NAP1. This linker is removed in a NEK2A kinase-dependent manner as mitosis begins. To explore C-NAP1 activities in regulating centrosome activities, we used genome editing to ablate it. C-NAP1-null cells were viable and had an increased frequency of premature centriole separation, accompanied by reduced density of the centriolar satellites, with reexpression of C-NAP1 rescuing both phenotypes. We found that the primary cilium, a signaling structure that arises from the mother centriole docked to the cell membrane, was intact in the absence of C-NAP1, although components of the ciliary rootlet were aberrantly localized away from the base of the cilium. C-NAP1-deficient cells were capable of signaling through the cilium, as determined by gene expression analysis after fluid flow-induced shear stress and the relocalization of components of the Hedgehog pathway. Centrosome amplification induced by DNA damage or by PLK4 or CDK2 overexpression was markedly reduced in the absence of C-NAP1. We conclude that centriole splitting reduces the local density of key centriolar precursors to impede overduplication.
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Affiliation(s)
- Anne-Marie Flanagan
- Centre for Chromosome Biology, School of Natural Sciences, National University of Ireland Galway, Galway, Ireland
| | - Elena Stavenschi
- Trinity Centre for Bioengineering, Trinity Biomedical Sciences Institute, and
- Department of Mechanical and Manufacturing Engineering, School of Engineering, Trinity College Dublin, Dublin 2, Ireland
| | - Shivakumar Basavaraju
- Centre for Chromosome Biology, School of Natural Sciences, National University of Ireland Galway, Galway, Ireland
| | - David Gaboriau
- Centre for Chromosome Biology, School of Natural Sciences, National University of Ireland Galway, Galway, Ireland
| | - David A Hoey
- Trinity Centre for Bioengineering, Trinity Biomedical Sciences Institute, and
- Department of Mechanical and Manufacturing Engineering, School of Engineering, Trinity College Dublin, Dublin 2, Ireland
- Advanced Materials and Bioengineering Research Centre, Trinity College Dublin, and Royal College of Surgeons in Ireland, Dublin 2, Ireland
| | - Ciaran G Morrison
- Centre for Chromosome Biology, School of Natural Sciences, National University of Ireland Galway, Galway, Ireland
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25
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Liu DN, Zhou YF, Peng AF, Long XH, Chen XY, Liu ZL, Xia H. HELQ reverses the malignant phenotype of osteosarcoma cells via CHK1-RAD51 signaling pathway. Oncol Rep 2016; 37:1107-1113. [PMID: 28000895 DOI: 10.3892/or.2016.5329] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/15/2016] [Accepted: 11/03/2016] [Indexed: 11/05/2022] Open
Abstract
HELQ is a DNA helicase important for repair of DNA lesions and has been linked to several types of cancer. However, little is known about its relationship with osteosarcoma (OS) and its mechanism. In the present study, the expression of HELQ and its downstream mediators in OS cells was assayed by quantitative PCR and western blot analysis. The function of HELQ in OS cells was investigated by Transwell invasion, wound healing, CCK8 assays and Comet assay. The results demonstrated that HELQ gene and protein were expressed in OS cells. OS cell invasion, migration, proliferation and DNA damage repair were enhanced by HELQ knock-down with shRNA-lentivirus and inhibited by HELQ overexpression with lentivirus transfection. Furthermore, the antitumor activities of HELQ may be associated with upregulated expression of the DNA damage-related proteins CHK1 and RAD51. Our findings indicated that HELQ confers an anti-invasive phenotype on OS cells by activating the CHK1-RAD51 signaling pathway and suggested that HELQ could be recognized as a promising therapeutic target for OS and other types of malignant tumors.
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Affiliation(s)
- Dong Ning Liu
- Southern Medical University, Guangzhou, Guangdong, P.R. China
| | - Yun Fei Zhou
- Department of Orthopedics, The First Affiliated Hospital of Nanchang University, Nanchang, Jiangxi, P.R. China
| | - Ai Fen Peng
- Jiangxi University of Traditional Chinese Medicine, Nanchang, Jiangxi, P.R. China
| | - Xin Hua Long
- Department of Orthopedics, The First Affiliated Hospital of Nanchang University, Nanchang, Jiangxi, P.R. China
| | - Xuan Yin Chen
- Department of Orthopedics, The First Affiliated Hospital of Nanchang University, Nanchang, Jiangxi, P.R. China
| | - Zhi Li Liu
- Department of Orthopedics, The First Affiliated Hospital of Nanchang University, Nanchang, Jiangxi, P.R. China
| | - Hong Xia
- Southern Medical University, Guangzhou, Guangdong, P.R. China
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26
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Abstract
Here, we review how DNA damage affects the centrosome and how centrosomes communicate with the DNA damage response (DDR) apparatus. We discuss how several proteins of the DDR are found at centrosomes, including the ATM, ATR, CHK1 and CHK2 kinases, the BRCA1 ubiquitin ligase complex and several members of the poly(ADP-ribose) polymerase family. Stereotypical centrosome organisation, in which two centriole barrels are orthogonally arranged in a roughly toroidal pericentriolar material (PCM), is strongly affected by exposure to DNA-damaging agents. We describe the genetic dependencies and mechanisms for how the centrioles lose their close association, and the PCM both expands and distorts after DNA damage. Another consequence of genotoxic stress is that centrosomes undergo duplication outside the normal cell cycle stage, meaning that centrosome amplification is commonly seen after DNA damage. We discuss several potential mechanisms for how centrosome numbers become dysregulated after DNA damage and explore the links between the DDR and the PLK1- and separase-dependent mechanisms that drive centriole separation and reduplication. We also describe how centrosome components, such as centrin2, are directly involved in responding to DNA damage. This review outlines current questions on the involvement of centrosomes in the DDR.
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Affiliation(s)
- Lisa I Mullee
- Centre for Chromosome Biology, School of Natural Sciences, National University of Ireland Galway, Biosciences Building, Dangan, Galway, Ireland
| | - Ciaran G Morrison
- Centre for Chromosome Biology, School of Natural Sciences, National University of Ireland Galway, Biosciences Building, Dangan, Galway, Ireland.
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27
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Cosenza MR, Krämer A. Centrosome amplification, chromosomal instability and cancer: mechanistic, clinical and therapeutic issues. Chromosome Res 2016; 24:105-26. [PMID: 26645976 DOI: 10.1007/s10577-015-9505-5] [Citation(s) in RCA: 50] [Impact Index Per Article: 5.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/21/2022]
Abstract
Centrosomes, the main microtubule-organizing centers in most animal cells, are of crucial importance for the assembly of a bipolar mitotic spindle and subsequent faithful segregation of chromosomes into two daughter cells. Centrosome abnormalities can be found in virtually all cancer types and have been linked to chromosomal instability (CIN) and tumorigenesis. Although our knowledge on centrosome structure, replication, and amplification has greatly increased within recent years, still only very little is known on nature, causes, and consequences of centrosome aberrations in primary tumor tissues. In this review, we summarize our current insights into the mechanistic link between centrosome aberrations, aneuploidy, CIN and tumorigenesis. Mechanisms of induction and cellular consequences of aneuploidy, tetraploidization and CIN, as well as origin and effects of supernumerary centrosomes will be discussed. In addition, animal models for both CIN and centrosome amplification will be outlined. Finally, we describe approaches to exploit centrosome amplification, aneuploidy and CIN for novel and specific anticancer treatment strategies based on the modulation of chromosome missegregation rates.
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Affiliation(s)
- Marco Raffaele Cosenza
- Clinical Cooperation Unit Molecular Hematology/Oncology, German Cancer Research Center (DKFZ) and Department of Internal Medicine V, University of Heidelberg, Im Neuenheimer Feld 280, 69120, Heidelberg, Germany
| | - Alwin Krämer
- Clinical Cooperation Unit Molecular Hematology/Oncology, German Cancer Research Center (DKFZ) and Department of Internal Medicine V, University of Heidelberg, Im Neuenheimer Feld 280, 69120, Heidelberg, Germany.
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28
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Gouazé-Andersson V, Delmas C, Taurand M, Martinez-Gala J, Evrard S, Mazoyer S, Toulas C, Cohen-Jonathan-Moyal E. FGFR1 Induces Glioblastoma Radioresistance through the PLCγ/Hif1α Pathway. Cancer Res 2016; 76:3036-44. [PMID: 26896280 DOI: 10.1158/0008-5472.can-15-2058] [Citation(s) in RCA: 40] [Impact Index Per Article: 4.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/05/2015] [Accepted: 01/28/2016] [Indexed: 11/16/2022]
Abstract
FGF2 signaling in glioblastoma induces resistance to radiotherapy, so targeting FGF2/FGFR pathways might offer a rational strategy for tumor radiosensitization. To investigate this possibility, we evaluated a specific role for FGFR1 in glioblastoma radioresistance as modeled by U87 and LN18 glioblastomas in mouse xenograft models. Silencing FGFR1 decreased radioresistance in a manner associated with radiation-induced centrosome overduplication and mitotic cell death. Inhibiting PLCγ (PLCG1), a downstream effector signaling molecule for FGFR1, was sufficient to produce similar effects, arguing that PLCγ is an essential mediator of FGFR1-induced radioresistance. FGFR1 silencing also reduced expression of HIF1α, which in addition to its roles in hypoxic responses exerts an independent effect on radioresistance. Finally, FGFR1 silencing delayed the growth of irradiated tumor xenografts, in a manner that was associated with reduced HIF1α levels but not blood vessel alterations. Taken together, our results offer a preclinical proof of concept that FGFR1 targeting can degrade radioresistance in glioblastoma, a widespread problem in this tumor, prompting clinical investigations of the use of FGFR1 inhibitors for radiosensitization. Cancer Res; 76(10); 3036-44. ©2016 AACR.
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Affiliation(s)
- Valérie Gouazé-Andersson
- Institut National de la Santé et de la Recherche Médicale (INSERM) UMR1037/Université Toulouse III Paul Sabatier, Cancer Research Center of Toulouse (CRCT), Team 11, Toulouse, France
| | - Caroline Delmas
- Institut National de la Santé et de la Recherche Médicale (INSERM) UMR1037/Université Toulouse III Paul Sabatier, Cancer Research Center of Toulouse (CRCT), Team 11, Toulouse, France. Institut Claudius Regaud, IUCT-O, Toulouse, France
| | - Marion Taurand
- Institut National de la Santé et de la Recherche Médicale (INSERM) UMR1037/Université Toulouse III Paul Sabatier, Cancer Research Center of Toulouse (CRCT), Team 11, Toulouse, France
| | - Judith Martinez-Gala
- Institut National de la Santé et de la Recherche Médicale (INSERM) UMR1037/Université Toulouse III Paul Sabatier, Cancer Research Center of Toulouse (CRCT), Team 11, Toulouse, France. Institut Claudius Regaud, IUCT-O, Toulouse, France
| | - Solène Evrard
- Institut National de la Santé et de la Recherche Médicale (INSERM) UMR1037/Université Toulouse III Paul Sabatier, Cancer Research Center of Toulouse (CRCT), Team 11, Toulouse, France
| | - Sandrine Mazoyer
- Institut National de la Santé et de la Recherche Médicale (INSERM) UMR1037/Université Toulouse III Paul Sabatier, Cancer Research Center of Toulouse (CRCT), Team 11, Toulouse, France
| | - Christine Toulas
- Institut National de la Santé et de la Recherche Médicale (INSERM) UMR1037/Université Toulouse III Paul Sabatier, Cancer Research Center of Toulouse (CRCT), Team 11, Toulouse, France. Institut Claudius Regaud, IUCT-O, Toulouse, France.
| | - Elizabeth Cohen-Jonathan-Moyal
- Institut National de la Santé et de la Recherche Médicale (INSERM) UMR1037/Université Toulouse III Paul Sabatier, Cancer Research Center of Toulouse (CRCT), Team 11, Toulouse, France. Institut Claudius Regaud, IUCT-O, Toulouse, France.
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29
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Cellular Pathways in Response to Ionizing Radiation and Their Targetability for Tumor Radiosensitization. Int J Mol Sci 2016; 17:ijms17010102. [PMID: 26784176 PMCID: PMC4730344 DOI: 10.3390/ijms17010102] [Citation(s) in RCA: 293] [Impact Index Per Article: 32.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/29/2015] [Revised: 12/22/2015] [Accepted: 12/25/2015] [Indexed: 12/20/2022] Open
Abstract
During the last few decades, improvements in the planning and application of radiotherapy in combination with surgery and chemotherapy resulted in increased survival rates of tumor patients. However, the success of radiotherapy is impaired by two reasons: firstly, the radioresistance of tumor cells and, secondly, the radiation-induced damage of normal tissue cells located in the field of ionizing radiation. These limitations demand the development of drugs for either radiosensitization of tumor cells or radioprotection of normal tissue cells. In order to identify potential targets, a detailed understanding of the cellular pathways involved in radiation response is an absolute requirement. This review describes the most important pathways of radioresponse and several key target proteins for radiosensitization.
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30
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Daly OM, Gaboriau D, Karakaya K, King S, Dantas TJ, Lalor P, Dockery P, Krämer A, Morrison CG. Gene-targeted CEP164-deficient cells show a ciliation defect with intact DNA repair capacity. J Cell Sci 2016; 129:1769-74. [DOI: 10.1242/jcs.186221] [Citation(s) in RCA: 20] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/18/2016] [Accepted: 03/08/2016] [Indexed: 12/31/2022] Open
Abstract
Primary cilia are microtubule structures that extend from the distal end of the mature, mother centriole. CEP164 is a component of the distal appendages carried by the mother centriole that is required for primary cilium formation. Recent data have implicated CEP164 as a ciliopathy gene and suggest that CEP164 plays some roles in the DNA damage response (DDR). We used reverse genetics to test the role of CEP164 in the DDR. We found that conditional depletion of CEP164 in chicken DT40 cells using an auxin-inducible degron led to no increase in sensitivity to DNA damage induced by ionising or ultraviolet irradiation. Disruption of CEP164 in human retinal pigmented epithelial cells blocked primary cilium formation but did not affect cellular proliferation or cellular responses to ionising or ultraviolet irradiation. Furthermore, we observed no localisation of CEP164 to the nucleus using immunofluorescence microscopy and analysis of multiple tagged forms of CEP164. Our data suggest that CEP164 is not required in the DDR.
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Affiliation(s)
- Owen M. Daly
- Centre for Chromosome Biology, School of Natural Sciences, National University of Ireland Galway, Galway, Ireland
| | - David Gaboriau
- Centre for Chromosome Biology, School of Natural Sciences, National University of Ireland Galway, Galway, Ireland
| | - Kadin Karakaya
- Clinical Cooperation Unit Molecular Hematology/Oncology, German Cancer Research Center (DKFZ) and Department of Internal Medicine V, University of Heidelberg, Im Neuenheimer Feld 280, 69120, Heidelberg, Germany
| | - Sinéad King
- Centre for Chromosome Biology, School of Natural Sciences, National University of Ireland Galway, Galway, Ireland
| | - Tiago J. Dantas
- Centre for Chromosome Biology, School of Natural Sciences, National University of Ireland Galway, Galway, Ireland
| | - Pierce Lalor
- Anatomy, School of Medicine, National University of Ireland Galway, Galway, Ireland
| | - Peter Dockery
- Anatomy, School of Medicine, National University of Ireland Galway, Galway, Ireland
| | - Alwin Krämer
- Clinical Cooperation Unit Molecular Hematology/Oncology, German Cancer Research Center (DKFZ) and Department of Internal Medicine V, University of Heidelberg, Im Neuenheimer Feld 280, 69120, Heidelberg, Germany
| | - Ciaran G. Morrison
- Centre for Chromosome Biology, School of Natural Sciences, National University of Ireland Galway, Galway, Ireland
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31
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Bailey LJ, Bianchi J, Hégarat N, Hochegger H, Doherty AJ. PrimPol-deficient cells exhibit a pronounced G2 checkpoint response following UV damage. Cell Cycle 2015; 15:908-18. [PMID: 26694751 PMCID: PMC4889237 DOI: 10.1080/15384101.2015.1128597] [Citation(s) in RCA: 22] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/19/2015] [Revised: 11/26/2015] [Accepted: 11/27/2015] [Indexed: 12/04/2022] Open
Abstract
PrimPol is a recently identified member of the archaeo-eukaryote primase (AEP) family of primase-polymerases. It has been shown that this mitochondrial and nuclear localized enzyme plays roles in the maintenance of both unperturbed replication fork progression and in the bypass of lesions after DNA damage. Here, we utilized an avian (DT40) knockout cell line to further study the consequences of loss of PrimPol (PrimPol(-/-)) on the downstream maintenance of cells after UV damage. We report that PrimPol(-/-) cells are more sensitive to UV-C irradiation in colony survival assays than Pol η-deficient cells. Although this increased UV sensitivity is not evident in cell viability assays, we show that this discrepancy is due to an enhanced checkpoint arrest after UV-C damage in the absence of PrimPol. PrimPol(-/-) arrested cells become stalled in G2, where they are protected from UV-induced cell death. Despite lacking an enzyme required for the bypass and maintenance of replication fork progression in the presence of UV damage, we show that PrimPol(-/-) cells actually have an advantage in the presence of a Chk1 inhibitor due to their slow progression through S-phase.
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Affiliation(s)
- Laura J Bailey
- a Genome Damage and Stability Centre, School of Life Sciences, University of Sussex , Brighton , UK
| | - Julie Bianchi
- a Genome Damage and Stability Centre, School of Life Sciences, University of Sussex , Brighton , UK
- b Present address: Department of Oncology-Pathology, Cancer Center Karolinska, Karolinska Institutet , Stockholm , Sweden
| | - Nadia Hégarat
- a Genome Damage and Stability Centre, School of Life Sciences, University of Sussex , Brighton , UK
| | - Helfrid Hochegger
- a Genome Damage and Stability Centre, School of Life Sciences, University of Sussex , Brighton , UK
| | - Aidan J Doherty
- a Genome Damage and Stability Centre, School of Life Sciences, University of Sussex , Brighton , UK
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32
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Opposing effects of pericentrin and microcephalin on the pericentriolar material regulate CHK1 activation in the DNA damage response. Oncogene 2015; 35:2003-10. [DOI: 10.1038/onc.2015.257] [Citation(s) in RCA: 23] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/20/2014] [Revised: 05/03/2015] [Accepted: 05/26/2015] [Indexed: 12/17/2022]
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33
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Zhao B, Zhang WD, Duan YL, Lu YQ, Cun YX, Li CH, Guo K, Nie WH, Li L, Zhang R, Zheng P. Filia Is an ESC-Specific Regulator of DNA Damage Response and Safeguards Genomic Stability. Cell Stem Cell 2015; 16:684-98. [PMID: 25936915 DOI: 10.1016/j.stem.2015.03.017] [Citation(s) in RCA: 42] [Impact Index Per Article: 4.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/03/2014] [Revised: 02/16/2015] [Accepted: 03/22/2015] [Indexed: 12/20/2022]
Abstract
Pluripotent stem cells (PSCs) hold great promise in cell-based therapy, but the genomic instability seen in culture hampers their full application. A greater understanding of the factors that regulate genomic stability in PSCs could help address this issue. Here we describe the identification of Filia as a specific regulator of genomic stability in mouse embryonic stem cells (ESCs). Filia expression is induced by genotoxic stress. Filia promotes centrosome integrity and regulates the DNA damage response (DDR) through multiple pathways, including DDR signaling, cell-cycle checkpoints and damage repair, ESC differentiation, and apoptosis. Filia depletion causes ESC genomic instability, induces resistance to apoptosis, and promotes malignant transformation. As part of its role in DDR, Filia interacts with PARP1 and stimulates its enzymatic activity. Filia also constitutively resides on centrosomes and translocates to DNA damage sites and mitochondria, consistent with its multifaceted roles in regulating centrosome integrity, damage repair, and apoptosis.
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Affiliation(s)
- Bo Zhao
- State Key Laboratory of Genetic Resources and Evolution, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming, Yunnan 650223, China; Yunnan Key Laboratory of Animal Reproduction, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming, Yunnan 650223, China
| | - Wei-Dao Zhang
- State Key Laboratory of Genetic Resources and Evolution, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming, Yunnan 650223, China; Yunnan Key Laboratory of Animal Reproduction, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming, Yunnan 650223, China; Kunming College of Life Science, University of Chinese Academy of Sciences, Kunming 650204, China
| | - Ying-Liang Duan
- State Key Laboratory of Genetic Resources and Evolution, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming, Yunnan 650223, China; Yunnan Key Laboratory of Animal Reproduction, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming, Yunnan 650223, China
| | - Yong-Qing Lu
- State Key Laboratory of Genetic Resources and Evolution, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming, Yunnan 650223, China; Yunnan Key Laboratory of Animal Reproduction, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming, Yunnan 650223, China
| | - Yi-Xian Cun
- State Key Laboratory of Genetic Resources and Evolution, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming, Yunnan 650223, China; Yunnan Key Laboratory of Animal Reproduction, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming, Yunnan 650223, China
| | - Chao-Hui Li
- State Key Laboratory of Genetic Resources and Evolution, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming, Yunnan 650223, China; Yunnan Key Laboratory of Animal Reproduction, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming, Yunnan 650223, China; Kunming College of Life Science, University of Chinese Academy of Sciences, Kunming 650204, China
| | - Kun Guo
- State Key Laboratory of Genetic Resources and Evolution, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming, Yunnan 650223, China; Yunnan Key Laboratory of Animal Reproduction, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming, Yunnan 650223, China; Kunming College of Life Science, University of Chinese Academy of Sciences, Kunming 650204, China
| | - Wen-Hui Nie
- State Key Laboratory of Genetic Resources and Evolution, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming, Yunnan 650223, China
| | - Lei Li
- State Key Laboratory of Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing, 100101, China
| | - Rugang Zhang
- Gene Expression and Regulation Program, The Wistar Institute Cancer Center, The Wistar Institute, Philadelphia, PA 19104, USA
| | - Ping Zheng
- State Key Laboratory of Genetic Resources and Evolution, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming, Yunnan 650223, China; Yunnan Key Laboratory of Animal Reproduction, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming, Yunnan 650223, China.
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Filipová A, Diaz-Garcia D, Bezrouk A, Čížková D, Havelek R, Vávrová J, Dayanithi G, Řezacová M. Ionizing radiation increases primary cilia incidence and induces multiciliation in C2C12 myoblasts. Cell Biol Int 2015; 39:943-53. [DOI: 10.1002/cbin.10462] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/01/2014] [Accepted: 03/10/2015] [Indexed: 12/12/2022]
Affiliation(s)
- Alžběta Filipová
- Department of Medical Biochemistry; Faculty of Medicine, Charles University in Prague; Sokolská 581 500 05 Hradec Králové Czech Republic
| | - Daniel Diaz-Garcia
- Department of Histology and Embryology; Faculty of Medicine, Charles University in Prague; Hradec Králové Czech Republic
| | - Aleš Bezrouk
- Department of Medical Biophysics; Faculty of Medicine, Charles University in Prague; Hradec Králové Czech Republic
| | - Dana Čížková
- Department of Histology and Embryology; Faculty of Medicine, Charles University in Prague; Hradec Králové Czech Republic
| | - Radim Havelek
- Department of Medical Biochemistry; Faculty of Medicine, Charles University in Prague; Sokolská 581 500 05 Hradec Králové Czech Republic
| | - Jiřina Vávrová
- Department of Radiobiology, Faculty of Military Health Sciences; University of Defence; Hradec Králové Czech Republic
| | - Govindan Dayanithi
- Department of Molecular Neurophysiology, Institute of Experimental Medicine; Czech Academy of Sciences; Videnska 1083 142 20 Prague Czech Republic
- Institut National de la Santé et de la Recherche Médicale U1198; Université Montpellier; Montpellier France
- Ecole Pratique des Hautes Etudes-Sorbonne; Paris France
| | - Martina Řezacová
- Department of Medical Biochemistry; Faculty of Medicine, Charles University in Prague; Sokolská 581 500 05 Hradec Králové Czech Republic
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35
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Arquint C, Gabryjonczyk AM, Nigg EA. Centrosomes as signalling centres. Philos Trans R Soc Lond B Biol Sci 2015; 369:rstb.2013.0464. [PMID: 25047618 DOI: 10.1098/rstb.2013.0464] [Citation(s) in RCA: 120] [Impact Index Per Article: 12.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/14/2022] Open
Abstract
Centrosomes-as well as the related spindle pole bodies (SPBs) of yeast-have been extensively studied from the perspective of their microtubule-organizing roles. Moreover, the biogenesis and duplication of these organelles have been the subject of much attention, and the importance of centrosomes and the centriole-ciliary apparatus for human disease is well recognized. Much less developed is our understanding of another facet of centrosomes and SPBs, namely their possible role as signalling centres. Yet, many signalling components, including kinases and phosphatases, have been associated with centrosomes and spindle poles, giving rise to the hypothesis that these organelles might serve as hubs for the integration and coordination of signalling pathways. In this review, we discuss a number of selected studies that bear on this notion. We cover different processes (cell cycle control, development, DNA damage response) and organisms (yeast, invertebrates and vertebrates), but have made no attempt to be comprehensive. This field is still young and although the concept of centrosomes and SPBs as signalling centres is attractive, it remains primarily a concept-in need of further scrutiny. We hope that this review will stimulate thought and experimentation.
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Affiliation(s)
- Christian Arquint
- Biozentrum, University of Basel, Klingelbergstrasse 50/70, 4056 Basel, Switzerland
| | | | - Erich A Nigg
- Biozentrum, University of Basel, Klingelbergstrasse 50/70, 4056 Basel, Switzerland
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36
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Stellas D, Souliotis VL, Bekyrou M, Smirlis D, Kirsch-Volders M, Degrassi F, Cundari E, Kyrtopoulos SA. Benzo[a]pyrene-induced cell cycle arrest in HepG2 cells is associated with delayed induction of mitotic instability. Mutat Res 2014; 769:59-68. [PMID: 25771725 DOI: 10.1016/j.mrfmmm.2014.07.004] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/27/2014] [Revised: 06/19/2014] [Accepted: 07/11/2014] [Indexed: 06/04/2023]
Abstract
The environmental carcinogen benzo[a]pyrene (B[a]P) after being metabolised by cytochrome P450 enzymes forms DNA adducts. This abnormal situation induces changes in the cell cycle, DNA damage, chromosomal and mitotic aberrations, all of which may be related to carcinogenesis. In order to further investigate the mechanistic basis of these effects, HepG2 cells were treated with 3μM B[a]P for various time periods, followed by further incubation in the absence of B[a]P for up to 192h. B[a]P treatment led initially to S-phase arrest followed by recovery and subsequent induction of G2/M arrest, indicating activation of the corresponding DNA damage checkpoints. Immunofluorescence-based studies revealed accumulation of B[a]P-induced DNA adducts and chromosomal damage which persisted beyond mitosis and entry into a new cycle, thus giving rise to a new round of activation of the S-phase checkpoint. Prolonged further cultivation of the cells in the absence of B[a]P resulted in high frequencies of various abnormal mitotic events. Abrogation of the B[a]P-induced S-phase arrest by the Chk1 inhibitor UCN-01 triggered a strong apoptotic response but also dramatically decreased the frequency of mitotic abnormalities in the surviving cells, suggesting that events occurring during S-phase arrest contribute to the formation of delayed mitotic damage. Overall, our data demonstrate that, although S-phase arrest serves as a mechanism by which the cells reduce their load of genetic damage, its prolonged activation may also have a negative impact on the balance between cell death and heritable genetic damage.
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Affiliation(s)
- Dimitris Stellas
- Institute of Biology, Medicinal Chemistry and Biotechnology, National Hellenic Research Foundation, Athens, Greece.
| | - Vassilis L Souliotis
- Institute of Biology, Medicinal Chemistry and Biotechnology, National Hellenic Research Foundation, Athens, Greece
| | - Margarita Bekyrou
- Institute of Biology, Medicinal Chemistry and Biotechnology, National Hellenic Research Foundation, Athens, Greece
| | | | | | | | - Enrico Cundari
- Laboratory for Cell Genetics,Vrije Universiteit Brussel, Brussels, Belgium; Institute of Molecular Biology and Pathology C.N.R., Rome, Italy
| | - Soterios A Kyrtopoulos
- Institute of Biology, Medicinal Chemistry and Biotechnology, National Hellenic Research Foundation, Athens, Greece
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37
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Douthwright S, Sluder G. Link between DNA damage and centriole disengagement/reduplication in untransformed human cells. J Cell Physiol 2014; 229:1427-36. [PMID: 24532022 PMCID: PMC4122266 DOI: 10.1002/jcp.24579] [Citation(s) in RCA: 24] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/06/2014] [Accepted: 02/12/2014] [Indexed: 12/21/2022]
Abstract
The radiation and radiomimetic drugs used to treat human tumors damage DNA in both cancer cells and normal proliferating cells. Centrosome amplification after DNA damage is well established for transformed cell types but is sparsely reported and not fully understood in untransformed cells. We characterize centriole behavior after DNA damage in synchronized untransformed human cells. One hour treatment of S phase cells with the radiomimetic drug, Doxorubicin, prolongs G2 by at least 72 h, though 14% of the cells eventually go through mitosis in that time. By 72 h after DNA damage we observe a 52% incidence of centriole disengagement plus a 10% incidence of extra centrioles. We find that either APC/C or Plk activities can disengage centrioles after DNA damage, though they normally work in concert. All disengaged centrioles are associated with γ-tubulin and maturation markers and thus, should in principle be capable of reduplicating and organizing spindle poles. The low incidence of reduplication of disengaged centrioles during G2 is due to the p53-dependent expression of p21 and the consequent loss of Cdk2 activity. We find that 26% of the cells going through mitosis after DNA damage contain disengaged or extra centrioles. This could produce genomic instability through transient or persistent spindle multipolarity. Thus, for cancer patients the use of DNA damaging therapies raises the chances of genomic instability and evolution of transformed characteristics in proliferating normal cell populations.
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Affiliation(s)
- Stephen Douthwright
- Department of Cell and Developmental Biology University of Massachusetts Medical School, Worcester, Massachusetts 01655
| | - Greenfield Sluder
- Department of Cell and Developmental Biology University of Massachusetts Medical School, Worcester, Massachusetts 01655
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38
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Myung Park J, Tougeron D, Huang S, Okamoto K, Sinicrope FA. Beclin 1 and UVRAG confer protection from radiation-induced DNA damage and maintain centrosome stability in colorectal cancer cells. PLoS One 2014; 9:e100819. [PMID: 24956373 PMCID: PMC4067383 DOI: 10.1371/journal.pone.0100819] [Citation(s) in RCA: 57] [Impact Index Per Article: 5.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/05/2014] [Accepted: 05/29/2014] [Indexed: 12/15/2022] Open
Abstract
Beclin 1 interacts with UV-irradiation-resistance-associated gene (UVRAG) to form core complexes that induce autophagy. While cells with defective autophagy are prone to genomic instability that contributes to tumorigenesis, it is unknown whether Beclin1 or UVRAG can regulate the DNA damage/repair response to cancer treatment in established tumor cells. We found that siRNA knockdown of Beclin 1 or UVRAG can increase radiation-induced DNA double strand breaks (DSBs), shown by pATM and γH2Ax, and promote colorectal cancer cell death. Furthermore, knockdown of Beclin 1, UVRAG or ATG5 increased the percentage of irradiated cells with nuclear foci expressing 53BP1, a marker of nonhomologous end joining but not RAD51 (homologous recombination), compared to control siRNA. Beclin 1 siRNA was shown to attenuate UVRAG expression. Cells with a UVRAG deletion mutant defective in Beclin 1 binding showed increased radiation-induced DSBs and cell death compared to cells with ectopic wild-type UVRAG. Knockdown of Beclin 1 or UVRAG, but not ATG5, resulted in a significant increase in centrosome number (γ-tubulin staining) in irradiated cells compared to control siRNA. Taken together, these data indicate that Beclin 1 and UVRAG confer protection against radiation-induced DNA DSBs and may maintain centrosome stability in established tumor cells.
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Affiliation(s)
- Jae Myung Park
- Mayo Clinic and Mayo Clinic Cancer Center, Rochester, Minnesota, United States of America
| | - David Tougeron
- Mayo Clinic and Mayo Clinic Cancer Center, Rochester, Minnesota, United States of America
| | - Shengbing Huang
- Mayo Clinic and Mayo Clinic Cancer Center, Rochester, Minnesota, United States of America
| | - Koichi Okamoto
- Mayo Clinic and Mayo Clinic Cancer Center, Rochester, Minnesota, United States of America
| | - Frank A. Sinicrope
- Mayo Clinic and Mayo Clinic Cancer Center, Rochester, Minnesota, United States of America
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39
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Kim SH, Park ER, Joo HY, Shen YN, Hong SH, Kim CH, Singh R, Lee KH, Shin HJ. RRM1 maintains centrosomal integrity via CHK1 and CDK1 signaling during replication stress. Cancer Lett 2014; 346:249-56. [PMID: 24434653 DOI: 10.1016/j.canlet.2013.12.031] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/07/2013] [Revised: 12/12/2013] [Accepted: 12/26/2013] [Indexed: 01/14/2023]
Abstract
DNA lesion-induced centrosomal abnormalities during the replication phase are relatively unknown. Here, we report that RNAi-mediated depletion of RRM1 induces cell-cycle arrest at the replication phase, along with severe DNA damage and centrosomal amplification. Interestingly, CHK1 depletion synergistically increased RRM1-depletion-induced centrosomal amplification. In response to hydroxyurea, CHK1 was delocalized from the centrosome by RRM1 depletion. Moreover, CDK1, which functions in centrosome separation and is inhibited by CHK1, was found to be essential for RRMI1-depletion-induced centrosomal amplification. Thus, we herein demonstrate that RRM1 preserves chromosomal stability via the CHK1- and CDK1-dependent stabilization of the centrosomal integrity at the replication stage.
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Affiliation(s)
- Su-Hyeon Kim
- Division of Radiation Cancer Research, Korea Institute of Radiological & Medical Sciences, Seoul 139-706, Republic of Korea
| | - Eun-Ran Park
- Division of Radiation Cancer Research, Korea Institute of Radiological & Medical Sciences, Seoul 139-706, Republic of Korea
| | - Hyun-Yoo Joo
- Division of Radiation Cancer Research, Korea Institute of Radiological & Medical Sciences, Seoul 139-706, Republic of Korea
| | - Yan Nan Shen
- Division of Radiation Cancer Research, Korea Institute of Radiological & Medical Sciences, Seoul 139-706, Republic of Korea
| | - Sung Hee Hong
- Division of Radiation Cancer Research, Korea Institute of Radiological & Medical Sciences, Seoul 139-706, Republic of Korea
| | - Chun Ho Kim
- Division of Radiation Effect, Korea Institute of Radiological & Medical Sciences, Seoul 139-706, Republic of Korea
| | - Rachana Singh
- Division of Radiation Cancer Research, Korea Institute of Radiological & Medical Sciences, Seoul 139-706, Republic of Korea
| | - Kee-Ho Lee
- Division of Radiation Cancer Research, Korea Institute of Radiological & Medical Sciences, Seoul 139-706, Republic of Korea.
| | - Hyun-Jin Shin
- Division of Radiation Cancer Research, Korea Institute of Radiological & Medical Sciences, Seoul 139-706, Republic of Korea.
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40
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Zou J, Zhang D, Qin G, Chen X, Wang H, Zhang D. BRCA1 and FancJ cooperatively promote interstrand crosslinker induced centrosome amplification through the activation of polo-like kinase 1. Cell Cycle 2014; 13:3685-97. [PMID: 25483079 PMCID: PMC4612125 DOI: 10.4161/15384101.2014.964973] [Citation(s) in RCA: 15] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/01/2014] [Revised: 09/08/2014] [Accepted: 09/09/2014] [Indexed: 12/15/2022] Open
Abstract
DNA damage response (DDR) and the centrosome cycle are 2 of the most critical cellular processes affecting the genome stability in animal cells. Yet the cross-talks between DDR and the centrosome are poorly understood. Here we showed that deficiency of the breast cancer 1, early onset gene (BRCA1) induces centrosome amplification in non-stressed cells as previously reported while attenuating DNA damage-induced centrosome amplification (DDICA) in cells experiencing prolonged genotoxic stress. Mechanistically, the function of BRCA1 in promoting DDICA is through binding and recruiting polo-like kinase 1 (PLK1) to the centrosome. In a recent study, we showed that FancJ also suppresses centrosome amplification in non-stressed cells while promoting DDICA in both hydroxyurea and mitomycin C treated cells. FancJ is a key component of the BRCA1 B-complex. Here, we further demonstrated that, in coordination with BRCA1, FancJ promotes DDICA by recruiting both BRCA1 and PLK1 to the centrosome in the DNA damaged cells. Thus, we have uncovered a novel role of BRCA1 and FancJ in the regulation of DDICA. Dysregulation of DDR or centrosome cycle leads to aneuploidy, which is frequently seen in both solid and hematological cancers. BRCA1 and FancJ are known tumor suppressors and have well-recognized functions in DNA damage checkpoint and DNA repair. Together with our recent findings, we demonstrated here that BRCA1 and FancJ also play an important role in centrosome cycle especially in DDICA. DDICA is thought to be an alternative fail-safe mechanism to prevent cells experiencing severe DNA damage from becoming carcinogenic. Therefore, BRCA1 and FancJ are potential liaisons linking early DDR with the DDICA. We propose that together with their functions in DDR, the role of BRCA1 and FancJ in the activation of DDICA is also crucial for their tumor suppression functions in vivo.
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Key Words
- ATM, ataxia telangiectasia mutated
- ATR, ataxia telangiectasia Rad3-related
- BRCA1
- BRCA1, breast cancer gene 1
- CIN, chromosome instability
- DDICA, DNA damage induced centrosome amplification
- DDR, DNA damage response
- DNA damage response
- FancJ
- GFP, green fluorescent protein
- HR, homologous recombination
- HU, hydroxyurea
- ICL, interstrand cross-linkers
- MIN, microsatellite instability
- MMC, mitomycin C
- MT, microtubule
- PCM, pericentriolar materials
- PLK1
- PLK1, Polo-like kinase 1
- UTR, untranslated region
- WCL, whole-cell lysate
- centrosome amplification
- interstrand cross-link
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Affiliation(s)
- Jianqiu Zou
- Basic Biomedical Science Division; Sanford School of Medicine; University of South Dakota; Vermillion, SD USA
| | - Deli Zhang
- WeiFang Medical University; WeiFang, Shandong, China
| | - Guang Qin
- Department of Oncology; Central Hospital of TaiAn; TaiAn, Shandong, China
| | - Xiangming Chen
- Department of Oncology; Central Hospital of TaiAn; TaiAn, Shandong, China
| | - Hongmin Wang
- Basic Biomedical Science Division; Sanford School of Medicine; University of South Dakota; Vermillion, SD USA
| | - Dong Zhang
- Basic Biomedical Science Division; Sanford School of Medicine; University of South Dakota; Vermillion, SD USA
- Department of Biomedical Sciences; College of Osteopathic Medicine; New York Institute of Technology; Old Westbury, NY USA
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41
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Intratumoral Hypoxia as the Genesis of Genetic Instability and Clinical Prognosis in Prostate Cancer. ADVANCES IN EXPERIMENTAL MEDICINE AND BIOLOGY 2014; 772:189-204. [DOI: 10.1007/978-1-4614-5915-6_9] [Citation(s) in RCA: 26] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/26/2022]
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42
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Spontaneous slow replication fork progression elicits mitosis alterations in homologous recombination-deficient mammalian cells. Proc Natl Acad Sci U S A 2013; 111:763-8. [PMID: 24347643 DOI: 10.1073/pnas.1311520111] [Citation(s) in RCA: 61] [Impact Index Per Article: 5.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/18/2022] Open
Abstract
Homologous recombination deficient (HR(-)) mammalian cells spontaneously display reduced replication fork (RF) movement and mitotic extra centrosomes. We show here that these cells present a complex mitotic phenotype, including prolonged metaphase arrest, anaphase bridges, and multipolar segregations. We then asked whether the replication and the mitotic phenotypes are interdependent. First, we determined low doses of hydroxyurea that did not affect the cell cycle distribution or activate CHK1 phosphorylation but did slow the replication fork movement of wild-type cells to the same level than in HR(-) cells. Remarkably, these low hydroxyurea doses generated the same mitotic defects (and to the same extent) in wild-type cells as observed in unchallenged HR(-) cells. Reciprocally, supplying nucleotide precursors to HR(-) cells suppressed both their replication deceleration and mitotic extra centrosome phenotypes. Therefore, subtle replication stress that escapes to surveillance pathways and, thus, fails to prevent cells from entering mitosis alters metaphase progression and centrosome number, resulting in multipolar mitosis. Importantly, multipolar mitosis results in global unbalanced chromosome segregation involving the whole genome, even fully replicated chromosomes. These data highlight the cross-talk between chromosome replication and segregation, and the importance of HR at the interface of these two processes for protection against general genome instability.
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43
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Inanç B, Pütz M, Lalor P, Dockery P, Kuriyama R, Gergely F, Morrison CG. Abnormal centrosomal structure and duplication in Cep135-deficient vertebrate cells. Mol Biol Cell 2013; 24:2645-54. [PMID: 23864714 PMCID: PMC3756917 DOI: 10.1091/mbc.e13-03-0149] [Citation(s) in RCA: 22] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/18/2013] [Revised: 06/26/2013] [Accepted: 07/01/2013] [Indexed: 12/12/2022] Open
Abstract
Centrosomes are key microtubule-organizing centers that contain a pair of centrioles, conserved cylindrical, microtubule-based structures. Centrosome duplication occurs once per cell cycle and relies on templated centriole assembly. In many animal cells this process starts with the formation of a radially symmetrical cartwheel structure. The centrosomal protein Cep135 localizes to this cartwheel, but its role in vertebrates is not well understood. Here we examine the involvement of Cep135 in centriole function by disrupting the Cep135 gene in the DT40 chicken B-cell line. DT40 cells that lack Cep135 are viable and show no major defects in centrosome composition or function, although we note a small decrease in centriole numbers and a concomitant increase in the frequency of monopolar spindles. Furthermore, electron microscopy reveals an atypical structure in the lumen of Cep135-deficient centrioles. Centrosome amplification after hydroxyurea treatment increases significantly in Cep135-deficient cells, suggesting an inhibitory role for the protein in centrosome reduplication during S-phase delay. We propose that Cep135 is required for the structural integrity of centrioles in proliferating vertebrate cells, a role that also limits centrosome amplification in S-phase-arrested cells.
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Affiliation(s)
- Burcu Inanç
- Centre for Chromosome Biology, School of Natural Sciences, National University of Ireland Galway, Galway, Ireland
| | - Monika Pütz
- Cancer Research UK, Cambridge Research Institute, Li Ka Shing Centre, Cambridge CB2 0RE, United Kingdom
| | - Pierce Lalor
- Anatomy, School of Medicine, National University of Ireland Galway, Galway, Ireland
| | - Peter Dockery
- Anatomy, School of Medicine, National University of Ireland Galway, Galway, Ireland
| | - Ryoko Kuriyama
- Department of Genetics, Cell Biology, and Development, University of Minnesota, Minneapolis, MN 55455
| | - Fanni Gergely
- Cancer Research UK, Cambridge Research Institute, Li Ka Shing Centre, Cambridge CB2 0RE, United Kingdom
| | - Ciaran G. Morrison
- Centre for Chromosome Biology, School of Natural Sciences, National University of Ireland Galway, Galway, Ireland
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Zou J, Tian F, Li J, Pickner W, Long M, Rezvani K, Wang H, Zhang D. FancJ regulates interstrand crosslinker induced centrosome amplification through the activation of polo-like kinase 1. Biol Open 2013; 2:1022-31. [PMID: 24167712 PMCID: PMC3798185 DOI: 10.1242/bio.20135801] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/14/2013] [Accepted: 07/03/2013] [Indexed: 01/05/2023] Open
Abstract
DNA damage response (DDR) and the centrosome cycle are two of the most critical processes for maintaining a stable genome in animals. Sporadic evidence suggests a connection between these two processes. Here, we report our findings that six Fanconi Anemia (FA) proteins, including FancI and FancJ, localize to the centrosome. Intriguingly, we found that the localization of FancJ to the mother centrosome is stimulated by a DNA interstrand crosslinker, Mitomycin C (MMC). We further show that, in addition to its role in interstrand crosslinking (ICL) repair, FancJ also regulates the normal centrosome cycle as well as ICL induced centrosome amplification by activating the polo-like kinase 1 (PLK1). We have uncovered a novel function of FancJ in centrosome biogenesis and established centrosome amplification as an integral part of the ICL response.
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Affiliation(s)
- Jianqiu Zou
- Basic Biomedical Science Division, Sanford School of Medicine, University of South Dakota , Vermillion, South Dakota, 57069 , USA
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45
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Orr B, Compton DA. A double-edged sword: how oncogenes and tumor suppressor genes can contribute to chromosomal instability. Front Oncol 2013; 3:164. [PMID: 23825799 PMCID: PMC3695391 DOI: 10.3389/fonc.2013.00164] [Citation(s) in RCA: 48] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/30/2013] [Accepted: 06/06/2013] [Indexed: 12/21/2022] Open
Abstract
Most solid tumors are characterized by abnormal chromosome numbers (aneuploidy) and karyotypic profiling has shown that the majority of these tumors are heterogeneous and chromosomally unstable. Chromosomal instability (CIN) is defined as persistent mis-segregation of whole chromosomes and is caused by defects during mitosis. Large-scale genome sequencing has failed to reveal frequent mutations of genes encoding proteins involved in mitosis. On the contrary, sequencing has revealed that most mutated genes in cancer fall into a limited number of core oncogenic signaling pathways that regulate the cell cycle, cell growth, and apoptosis. This led to the notion that the induction of oncogenic signaling is a separate event from the loss of mitotic fidelity, but a growing body of evidence suggests that oncogenic signaling can deregulate cell cycle progression, growth, and differentiation as well as cause CIN. These new results indicate that the induction of CIN can no longer be considered separately from the cancer-associated driver mutations. Here we review the primary causes of CIN in mitosis and discuss how the oncogenic activation of key signal transduction pathways contributes to the induction of CIN.
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Affiliation(s)
- Bernardo Orr
- Department of Biochemistry, Geisel School of Medicine at Dartmouth , Hanover, NH , USA ; The Norris-Cotton Cancer Center, Geisel School of Medicine at Dartmouth , Hanover, NH , USA
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46
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Park MT, Oh ET, Song MJ, Lee H, Choi EK, Park HJ. NQO1 prevents radiation-induced aneuploidy by interacting with Aurora-A. Carcinogenesis 2013; 34:2470-85. [PMID: 23803694 DOI: 10.1093/carcin/bgt225] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/01/2023] Open
Abstract
Aneuploidy is the most common characteristic of human cancer cells. It also causes genomic instability, which is involved in the initiation of cancer development. Various lines of evidence indicate that nicotinamide adenine dinucleotide(P)H quinone oxidoreductase 1 (NQO1) plays an important role in cancer prevention, but the molecular mechanisms underlying this effect have not yet been fully elucidated. Here, we report that ionizing radiation (IR) induces substantial aneuploidy and centrosome amplification in NQO1-deficient cancer cells, suggesting that NQO1 plays a crucial role in preventing aneuploidy. NQO1 deficiency markedly increased the protein stability of Aurora-A in irradiated cancer cells. Small interfering RNA targeting Aurora-A effectively attenuated IR-induced centrosome amplification concerned with aneuploidy in NQO1-deficient cancer cells. Furthermore, we found that NQO1 specifically binds to Aurora-A via competing with the microtubule-binding protein, TPX2 (targeting protein for Xklp2), and contributes to the degradation of Aurora-A. Our results collectively demonstrate that NQO1 plays a key role in suppressing IR-induced centrosome amplification and aneuploidy through a direct interaction with Aurora-A.
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Affiliation(s)
- Moon-Taek Park
- Department of Microbiology, Center for Advanced Medical Education by BK21 Project, College of Medicine, Inha University, Incheon 400-712, Republic of Korea
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Petsalaki E, Zachos G. Chk1 and Mps1 jointly regulate correction of merotelic kinetochore attachments. J Cell Sci 2013; 126:1235-46. [PMID: 23321637 DOI: 10.1242/jcs.119677] [Citation(s) in RCA: 20] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/30/2023] Open
Abstract
If uncorrected, merotelic kinetochore attachments can induce mis-segregated chromosomes in anaphase. We show that checkpoint kinase 1 (Chk1) protects vertebrate cells against merotelic attachments and lagging chromosomes and is required for correction of merotelic attachments during a prolonged metaphase. Decreased Chk1 activity leads to hyper-stable kinetochore microtubules, unstable binding of MCAK, Kif2b and Mps1 to centromeres or kinetochores and reduced phosphorylation of Hec1 by Aurora-B. Phosphorylation of Aurora-B at serine 331 (Ser331) by Chk1 is high in prometaphase and decreases significantly in metaphase cells. We propose that Ser331 phosphorylation is required for optimal localization of MCAK, Kif2b and Mps1 to centromeres or kinetochores and for Hec1 phosphorylation. Furthermore, inhibition of Mps1 activity diminishes initial recruitment of MCAK and Kif2b to centromeres or kinetochores, impairs Hec1 phosphorylation and exacerbates merotelic attachments in Chk1-deficient cells. We propose that Chk1 and Mps1 jointly regulate Aurora-B, MCAK, Kif2b and Hec1 to correct merotelic attachments. These results suggest a role for Chk1 and Mps1 in error correction.
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Affiliation(s)
- Eleni Petsalaki
- Department of Biology, University of Crete, Vassilika Vouton, Heraklion 70013, Greece
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Conroy PC, Saladino C, Dantas TJ, Lalor P, Dockery P, Morrison CG. C-NAP1 and rootletin restrain DNA damage-induced centriole splitting and facilitate ciliogenesis. Cell Cycle 2013; 11:3769-78. [PMID: 23070519 DOI: 10.4161/cc.21986] [Citation(s) in RCA: 33] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/14/2022] Open
Abstract
Cilia are found on most human cells and exist as motile cilia or non-motile primary cilia. Primary cilia play sensory roles in transducing various extracellular signals, and defective ciliary functions are involved in a wide range of human diseases. Centrosomes are the principal microtubule-organizing centers of animal cells and contain two centrioles. We observed that DNA damage causes centriole splitting in non-transformed human cells, with isolated centrioles carrying the mother centriole markers CEP170 and ninein but not kizuna or cenexin. Loss of centriole cohesion through siRNA depletion of C-NAP1 or rootletin increased radiation-induced centriole splitting, with C-NAP1-depleted isolated centrioles losing mother markers. As the mother centriole forms the basal body in primary cilia, we tested whether centriole splitting affected ciliogenesis. While irradiated cells formed apparently normal primary cilia, most cilia arose from centriolar clusters, not from isolated centrioles. Furthermore, C-NAP1 or rootletin knockdown reduced primary cilium formation. Therefore, the centriole cohesion apparatus at the proximal end of centrioles may provide a target that can affect primary cilium formation as part of the DNA damage response.
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Affiliation(s)
- Pauline C Conroy
- Centre for Chromosome Biology, School of Natural Sciences, National University of Ireland Galway, Galway, Ireland
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Wang Y, Dantas TJ, Lalor P, Dockery P, Morrison CG. Promoter hijack reveals pericentrin functions in mitosis and the DNA damage response. Cell Cycle 2013; 12:635-46. [PMID: 23324397 DOI: 10.4161/cc.23516] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/19/2022] Open
Abstract
Centrosomes, the principal microtubule-organizing centers of animal somatic cells, consist of two centrioles embedded in the pericentriolar material (PCM). Pericentrin is a large PCM protein that is required for normal PCM assembly. Mutations in PCNT cause primordial dwarfism. Pericentrin has also been implicated in the control of DNA damage responses. To test how pericentrin is involved in cell cycle control after genotoxic stress, we disrupted the Pcnt locus in chicken DT40 cells. Pericentrin-deficient cells proceeded through mitosis more slowly, with a high level of monopolar spindles, and were more sensitive to spindle poisons than controls. Centriole structures appeared normal by light and electron microscopy, but the PCM did not recruit γ-tubulin efficiently. Cell cycle delays after ionizing radiation (IR) treatment were normal in pericentrin-deficient cells. However, pericentrin disruption in Mcph1-/- cells abrogated centrosome hyperamplification after IR. We conclude that pericentrin controls genomic stability by both ensuring appropriate mitotic spindle activity and centrosome regulation.
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Affiliation(s)
- Yifan Wang
- Centre for Chromosome Biology, School of Natural Sciences, National University of Ireland Galway, Galway, Ireland
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Steroidogenic factor 1 (NR5A1) maintains centrosome homeostasis in steroidogenic cells by restricting centrosomal DNA-dependent protein kinase activation. Mol Cell Biol 2012; 33:476-84. [PMID: 23166296 DOI: 10.1128/mcb.01064-12] [Citation(s) in RCA: 26] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/20/2022] Open
Abstract
Steroidogenic factor 1 (SF-1 or NR5A1) is a nuclear receptor that controls adrenogenital cell growth and differentiation. Adrenogenital primordial cells from SF-1 knockout mice die of apoptosis, but the mechanism by which SF-1 regulates cell survival is not entirely clear. Besides functioning in the nucleus, SF-1 also resides in the centrosome and controls centrosome homeostasis. Here, we show that SF-1 restricts centrosome overduplication by inhibiting aberrant activation of DNA-dependent protein kinase (DNA-PK) in the centrosome. SF-1 was found to be associated with Ku70/Ku80 only in the centrosome, sequestering them from the catalytic subunit of DNA-PK (DNA-PKcs). In the absence of SF-1, DNA-PKcs was recruited to the centrosome and activated, causing aberrant activation of centrosomal Akt and cyclin-dependent kinase 2 (CDK2)/cyclin A and leading to centrosome overduplication. Centrosome overduplication caused by SF-1 depletion was averted by the elimination of DNA-PKcs, Ku70/80, or cyclin A or by the inhibition of CDK2 or Akt. In the nucleus, SF-1 did not interact with Ku70/80, and SF-1 depletion did not activate a nuclear DNA damage response. Centriole biogenesis was also unaffected. Thus, centrosomal DNA-PK signaling triggers centrosome overduplication, and this centrosomal event, but not the nuclear DNA damage response, is controlled by SF-1.
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