Publications by authors named "Anthi Karaiskou"

12 Publications

  • Page 1 of 1

Insights into the antiproliferative mechanism of (C^N)-chelated half-sandwich iridium complexes.

Dalton Trans 2020 Dec;49(48):17635-17641

Institut Parisien de Chimie Moléculaire (IPCM), Sorbonne Université, CNRS, 4 place Jussieu, F-75005 Paris, France.

Transition metal-based anticancer compounds, as an alternative to platinum derivatives, are raising scientific interest as they may present distinct although poorly understood mechanisms of action. We used a structure-activity relationship-based methodology to investigate the chemical and biological features of a series of ten (C^N)-chelated half-sandwich iridiumIII complexes of the general formula [IrCp*(phox)Cl], where (phox) is a 2-phenyloxazoline ligand forming a 5-membered metallacycle. This series of compounds undergoes a fast exchange of their chlorido ligand once solubilised in DMSO. They were cytotoxic to HeLa cells with IC50 values in the micromolar range and induced a rapid activation of caspase-3, an apoptosis marker. In vitro, the oxidative power of all the complexes towards NADH was highlighted but only the complexes bearing substituents on the oxazoline ring were able to produce H2O2 at the micromolar range. However, we demonstrated using a powerful HyPer protein redox sensor-based flow cytometry assay that most complexes rapidly raised intracellular levels of H2O2. Hence, this study shows that oxidative stress can partly explain the cytotoxicity of these complexes on the HeLa cell line and gives a first entry to their mechanism of action.
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http://dx.doi.org/10.1039/d0dt03414bDOI Listing
December 2020

Regulation of the EGFR/ErbB signalling by clathrin in response to various ligands in hepatocellular carcinoma cell lines.

J Cell Mol Med 2020 07 9;24(14):8091-8102. Epub 2020 Jun 9.

INSERM UMR_S 938, Centre de Recherche Saint-Antoine (CRSA), Sorbonne Université, Paris, France.

Membrane receptor intracellular trafficking and signalling are frequently altered in cancers. Our aim was to investigate whether clathrin-dependent trafficking modulates signalling of the ErbB receptor family in response to amphiregulin (AR), EGF, heparin-binding EGF-like growth factor (HB-EGF) and heregulin-1β (HRG). Experiments were performed using three hepatocellular carcinoma (HCC) cell lines, Hep3B, HepG2 and PLC/PRF/5, expressing various levels of EGFR, ErbB2 and ErbB3. Inhibition of clathrin-mediated endocytosis (CME), by down-regulating clathrin heavy chain expression, resulted in a cell- and ligand-specific pattern of phosphorylation of the ErbB receptors and their downstream effectors. Clathrin down-regulation significantly decreased the ratio between phosphorylated EGFR (pEGFR) and total EGFR in all cell lines when stimulated with AR, EGF, HB-EGF or HRG, except in HRG-stimulated Hep3B cells in which pEGFR was not detectable. The ratio between phosphorylated ErbB2 and total ErbB2 was significantly decreased in clathrin down-regulated Hep3B cells stimulated with any of the ligands, and in HRG-stimulated PLC/PRF/5 cells. The ratio between phosphorylated ErbB3 and total ErbB3 significantly decreased in clathrin down-regulated cell lines upon stimulation with EGF or HB-EGF. STAT3 phosphorylation levels significantly increased in all cell lines irrespective of stimulation, while that of AKT remained unchanged, except in AR-stimulated Hep3B and HepG2 cells in which pAKT was significantly decreased. Finally, ERK phosphorylation was insensitive to clathrin inhibition. Altogether, our observations indicate that clathrin regulation of ErbB signalling in HCC is a complex process that likely depends on the expression of ErbB family members and on the autocrine/paracrine secretion of their ligands in the tumour environment.
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http://dx.doi.org/10.1111/jcmm.15440DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7348188PMC
July 2020

Calibrated mitotic oscillator drives motile ciliogenesis.

Science 2017 11 5;358(6364):803-806. Epub 2017 Oct 5.

Institut de Biologie de l'École Normale Supérieure (IBENS), Paris Sciences et Lettres (PSL) Research University, Paris F-75005, France.

Cell division and differentiation depend on massive and rapid organelle remodeling. The mitotic oscillator, centered on the cyclin-dependent kinase 1-anaphase-promoting complex/cyclosome (CDK1-APC/C) axis, spatiotemporally coordinates this reorganization in dividing cells. Here we discovered that nondividing cells could also implement this mitotic clocklike regulatory circuit to orchestrate subcellular reorganization associated with differentiation. We probed centriole amplification in differentiating mouse-brain multiciliated cells. These postmitotic progenitors fine-tuned mitotic oscillator activity to drive the orderly progression of centriole production, maturation, and motile ciliation while avoiding the mitosis commitment threshold. Insufficient CDK1 activity hindered differentiation, whereas excessive activity accelerated differentiation yet drove postmitotic progenitors into mitosis. Thus, postmitotic cells can redeploy and calibrate the mitotic oscillator to uncouple cytoplasmic from nuclear dynamics for organelle remodeling associated with differentiation.
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http://dx.doi.org/10.1126/science.aan8311DOI Listing
November 2017

C-Nap1 mutation affects centriole cohesion and is associated with a Seckel-like syndrome in cattle.

Nat Commun 2015 Apr 23;6:6894. Epub 2015 Apr 23.

1] Institut National de la Recherche Agronomique (INRA), Unité Mixte de Recherche 1313-Génétique Animale et Biologie Intégrative (UMR1313-GABI), F-78352 Jouy-en-Josas, France [2] Allice, Département R&D, F-75595 Paris, France.

Caprine-like Generalized Hypoplasia Syndrome (SHGC) is an autosomal-recessive disorder in Montbéliarde cattle. Affected animals present a wide range of clinical features that include the following: delayed development with low birth weight, hind limb muscular hypoplasia, caprine-like thin head and partial coat depigmentation. Here we show that SHGC is caused by a truncating mutation in the CEP250 gene that encodes the centrosomal protein C-Nap1. This mutation results in centrosome splitting, which neither affects centriole ultrastructure and duplication in dividing cells nor centriole function in cilium assembly and mitotic spindle organization. Loss of C-Nap1-mediated centriole cohesion leads to an altered cell migration phenotype. This discovery extends the range of loci that constitute the spectrum of autosomal primary recessive microcephaly (MCPH) and Seckel-like syndromes.
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http://dx.doi.org/10.1038/ncomms7894DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4423223PMC
April 2015

Metamorphosis in solitary ascidians.

Genesis 2015 Jan 14;53(1):34-47. Epub 2014 Oct 14.

Sorbonne Universités, UPMC Univ Paris 06, UMR7622-Biologie du Développement, Paris, France.

Embryonic and postembryonic development in ascidians have been studied for over a century, but it is only in the last 10 years that the complex molecular network involved in coordinating postlarval development and metamorphosis has started to emerge. In most ascidians, the transition from the larval to the sessile juvenile/adult stage, or metamorphosis, requires a combination of environmental and endogenous signals and is characterized by coordinated global morphogenetic changes that are initiated by the adhesion of the larvae. Cloney was the first to describe cellular events of ascidians' metamorphosis in 1978 and only recently elements of the molecular regulation of this crucial developmental step have been revealed. This review aims to present a thorough view of this crucial developmental step by combining recent molecular data to the already established cellular events.
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http://dx.doi.org/10.1002/dvg.22824DOI Listing
January 2015

A critical balance between Cyclin B synthesis and Myt1 activity controls meiosis entry in Xenopus oocytes.

Development 2011 Sep 27;138(17):3735-44. Epub 2011 Jul 27.

UPMC Université Paris 06, UMR7622-Biologie du Développement, 9 quai Saint Bernard, 75005 Paris, France.

In fully grown oocytes, meiosis is arrested at first prophase until species-specific initiation signals trigger maturation. Meiotic resumption universally involves early activation of M phase-promoting factor (Cdc2 kinase-Cyclin B complex, MPF) by dephosphorylation of the inhibitory Thr14/Tyr15 sites of Cdc2. However, underlying mechanisms vary. In Xenopus oocytes, deciphering the intervening chain of events has been hampered by a sensitive amplification loop involving Cdc2-Cyclin B, the inhibitory kinase Myt1 and the activating phosphatase Cdc25. In this study we provide evidence that the critical event in meiotic resumption is a change in the balance between inhibitory Myt1 activity and Cyclin B neosynthesis. First, we show that in fully grown oocytes Myt1 is essential for maintaining prophase I arrest. Second, we demonstrate that, upon upregulation of Cyclin B synthesis in response to progesterone, rapid inactivating phosphorylation of Myt1 occurs, mediated by Cdc2 and without any significant contribution of Mos/MAPK or Plx1. We propose a model in which the appearance of active MPF complexes following increased Cyclin B synthesis causes Myt1 inhibition, upstream of the MPF/Cdc25 amplification loop.
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http://dx.doi.org/10.1242/dev.063974DOI Listing
September 2011

Absence of reciprocal feedback between MPF and ERK2 MAP kinase in mitotic Xenopus laevis embryo cell-free extract.

Cell Cycle 2007 Feb 18;6(4):489-96. Epub 2007 Feb 18.

Biology and Genetics of Development, Mitosis and Meiosis Group, Institute of Genetics and Development of Rennes, University Rennes, Rennes Cedex, France.

MPF and MAP kinase ERK2 are two major M-phase kinases. They interact with each other in a complex way during meiotic maturation of Xenopus laevis oocytes. Here we study their interrelationship during first mitosis in X. laevis embryo cell-free extract perturbing the polyubiquitination pathway as a tool. Recombinant ubiquitin K48R (Ub-K48R) mutant protein arrests mitotic cyclin B degradation in the extract. This results in both increased accumulation of phosphorylated form of cyclin B2 and MPF activity as well as mitotic phosphorylation of its substrates. Ub-K48R also increased the mitotic phosphorylation of ERK2. Simultaneous addition of Ub-K48R and the proteasome inhibitor MG 132 strengthened and further prolonged MPF activity, MCM4 phosphorylation and accumulation of phosphorylated forms of cyclin B2. ERK2 phosphorylation levels increased and persisted longer than upon action of Ub-K48R alone. This shows a synergistic effect of inhibition of two different steps of ubiquitin-proteasome pathway on MPF activity and mitotic phosphorylation and ubiquitination of specific M-phase proteins. On the other hand, complete inhibition of ERK2 activation using U0126 had no effect either on MPF activity or on MCM4 phosphorylation either in control or in Ub-K48R-supplemented extracts. Experimental reduction of MPF activity by addition of recombinant p21(Cip) protein resulted in significant reduction of ERK2 phosphorylation. Thus, the reciprocal feedback observed between MPF and ERK2 in meiosis is not observed during mitotic M-phase in cell-free Xenopus embryo extracts. ERK2 phosphorylation is regulated by the levels of MPF activity, however no influence of ERK2 on MPF activity could be detected. These results show a fundamental difference in the relationship between the two major M-phase kinases in meiotic and mitotic cell cycle.
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http://dx.doi.org/10.4161/cc.6.4.3860DOI Listing
February 2007

Polo-like kinase confers MPF autoamplification competence to growing Xenopus oocytes.

Development 2004 Apr 25;131(7):1543-52. Epub 2004 Feb 25.

Laboratoire de Biologie du Développement, UMR-CNRS 7622, Equipe 'Biologie de l'ovocyte', Université Pierre et Marie Curie, boîte 24, 4 place Jussieu, 75252 Paris cedex 05, France.

During oogenesis, the Xenopus oocyte is blocked in prophase of meiosis I. It becomes competent to resume meiosis in response to progesterone at the end of its growing period (stage VI of oogenesis). Stage IV oocytes contain a store of inactive pre-MPF (Tyr15-phosphorylated Cdc2 bound to cyclin B2); the Cdc25 phosphatase that catalyzes Tyr15 dephosphorylation of Cdc2 is also present. However, the positive feedback loop that allows MPF autoamplification is not functional at this stage of oocyte growth. We report that when cyclin B is overexpressed in stage IV oocytes, MPF autoamplification does not occur and the newly formed cyclin B-Cdc2 complexes are inactivated by Tyr15 phosphorylation, indicating that Myt1 kinase remains active and that Cdc25 is prevented to be activated. Plx1 kinase (or polo-like kinase), which is required for Cdc25 activation and MPF autoamplification in full grown oocytes is not expressed at the protein level in small stage IV oocytes. In order to determine if Plx1 could be the missing regulator that prevents MPF autoamplification, polo kinase was overexpressed in stage IV oocytes. Under these conditions, the MPF-positive feedback loop was restored. Moreover, we show that acquisition of autoamplification competence does not require the Mos/MAPK pathway.
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http://dx.doi.org/10.1242/dev.01050DOI Listing
April 2004

Regulation of EDEN-dependent deadenylation of Aurora A/Eg2-derived mRNA via phosphorylation and dephosphorylation in Xenopus laevis egg extracts.

J Cell Sci 2003 Jul 13;116(Pt 13):2697-705. Epub 2003 May 13.

UMR 6061 CNRS, University of Rennes 1, Faculty of Medicine, 2 Ave. Prof. Léon Bernard, CS 34317, 35043 Rennes Cedex, France.

Deadenylation is an intimate part of the post-transcriptional regulation of maternal mRNAs in embryos. EDEN-BP is so far the only known member of a complex regulating the deadenylation of maternal mRNA in Xenopus laevis embryos in a manner that is dependent on the 3'-untranslated region called EDEN (embryo deadenylation element). In this report, we show that calcium activation of cell-free extracts triggers EDEN binding protein (EDEN-BP) dephosphorylation and concomitant deadenylation of a chimeric RNA bearing Aurora A/Eg2 EDEN sequence. Deadenylation of mRNA deprived of EDEN sequence (default deadenylation) does not change with egg activation. Kinase and phosphatase inhibitors downregulate EDEN-dependent deadenylation but they do not substantially influence default deadenylation. Using indestructible Delta90 cyclin B to revert interphase extracts to the M-phase, we show that modulation of EDEN-dependent deadenylation is independent of M-phase promoting factor (MPF) activity. These results suggest that the increase in EDEN-dependent deadenylation following egg activation is achieved, at least partially, via dephosphorylation and/or phosphorylation of regulatory proteins, including EDEN-BP dephosphorylation. This regulation proceeds in a manner independent from MPF inactivation.
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http://dx.doi.org/10.1242/jcs.00477DOI Listing
July 2003

Thr-161 phosphorylation of monomeric Cdc2. Regulation by protein phosphatase 2C in Xenopus oocytes.

J Biol Chem 2002 Aug 29;277(32):28592-600. Epub 2002 May 29.

Laboratoire de Biologie du Développement, Institut de la Recherche Agronomique/Unité Mixte de Recherche-CNRS 7622, Université Pierre et Marie Curie, Boite 24, 4 Place Jussieu, 75252 Paris cedex 05, France.

Fully grown Xenopus oocyte is arrested at prophase I of meiosis. Re-entry into meiosis depends on the activation of MPF (M-phase promoting factor or cyclin B.Cdc2 complex), triggered by progesterone. The prophase-arrested oocyte contains a store of Cdc2. Most of the protein is present as a monomer whereas a minor fraction, called pre-MPF, is found to be associated with cyclin B. Activation of Cdc2 depends on two key events: cyclin binding and an activating phosphorylation on Thr-161 residue located in the T-loop. To get new insights into the regulation of Thr-161 phosphorylation of Cdc2, monomeric Cdc2 was isolated from prophase oocytes. Based on its activation upon cyclin addition and detection by an antibody directed specifically against Cdc2 phosphorylated on Thr-161, we show for the first time that the prophase oocyte contains a significant amount of monomeric Cdc2 phosphorylated on Thr-161. PP2C, a Mg2+-dependent phosphatase, negatively controls Thr-161 phosphorylation of Cdc2. The unexpected presence of a population of free Cdc2 already phosphorylated on Thr-161 could contribute to the generation of the Cdc2 kinase activity threshold required to initiate MPF amplification.
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http://dx.doi.org/10.1074/jbc.M202742200DOI Listing
August 2002
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