84 results on '"Renata Basto"'
Search Results
2. A catalog of numerical centrosome defects in epithelial ovarian cancers
- Author
-
Jean‐Philippe Morretton, Anthony Simon, Aurélie Herbette, Jorge Barbazan, Carlos Pérez‐González, Camille Cosson, Bassirou Mboup, Aurélien Latouche, Tatiana Popova, Yann Kieffer, Anne‐Sophie Macé, Pierre Gestraud, Guillaume Bataillon, Véronique Becette, Didier Meseure, André Nicolas, Odette Mariani, Anne Vincent‐Salomon, Marc‐Henri Stern, Fatima Mechta‐Grigoriou, Sergio Roman Roman, Danijela Matic Vignjevic, Roman Rouzier, Xavier Sastre‐Garau, Oumou Goundiam, and Renata Basto
- Subjects
centrosomes ,ovarian cancers ,centrosome number alterations ,Medicine (General) ,R5-920 ,Genetics ,QH426-470 - Abstract
Abstract Centrosome amplification, the presence of more than two centrosomes in a cell is a common feature of most human cancer cell lines. However, little is known about centrosome numbers in human cancers and whether amplification or other numerical aberrations are frequently present. To address this question, we have analyzed a large cohort of primary human epithelial ovarian cancers (EOCs) from 100 patients. We found that rigorous quantitation of centrosome number in tumor samples was extremely challenging due to tumor heterogeneity and extensive tissue disorganization. Interestingly, even if centrosome clusters could be identified, the incidence of centrosome amplification was not comparable to what has been described in cultured cancer cells. Surprisingly, centrosome loss events where a few or many nuclei were not associated with centrosomes were clearly noticed and overall more frequent than centrosome amplification. Our findings highlight the difficulty of characterizing centrosome numbers in human tumors, while revealing a novel paradigm of centrosome number defects in EOCs.
- Published
- 2022
- Full Text
- View/download PDF
3. Topical Delivery of Niacinamide to Skin Using Hybrid Nanogels Enhances Photoprotection Effect
- Author
-
Renata Basto, Raquel Andrade, Cláudia Nunes, Sofia A. Costa Lima, and Salette Reis
- Subjects
in vitro release ,jojoba oil ,oleic acid ,permeation enhancers ,transethosomes ,tween 80 ,Pharmacy and materia medica ,RS1-441 - Abstract
Niacinamide (NIA) has been widely used in halting the features of ageing by acting as an antioxidant and preventing dehydration. NIA’s physicochemical properties suggest difficulties in surpassing the barrier imposed by the stratum corneum layer to reach the target in the skin. To improve cutaneous delivery of NIA, a hybrid nanogel was designed using carrageenan and polyvinylpyrrolidone polymers combined with jojoba oil as a permeation enhancer. Three different types of transethosomes were prepared by the thin-film hydration method, made distinct by the presence of either an edge activator or a permeation enhancer, to allow for a controlled delivery of NIA. Formulations were characterized by measurements of size, polydispersity index, zeta potential, encapsulation efficiency, and loading capacity, and by evaluating their chemical interactions and morphology. Skin permeation assays were performed using Franz diffusion cells. The hybrid hydrogels exhibited robust, porous, and highly aligned macrostructures, and when present, jojoba oil changed their morphology. Skin permeation studies with transethosomes-loaded hydrogels showed that nanogels per se exhibit a more controlled and enhanced permeation, in particular when jojoba oil was present in the transethosomes. These promising nanogels protected the human keratinocytes from UV radiation, and thus can be added to sunscreens or after-sun lotions to improve skin protection.
- Published
- 2021
- Full Text
- View/download PDF
4. CEP19 cooperates with FOP and CEP350 to drive early steps in the ciliogenesis programme
- Author
-
Bahareh A. Mojarad, Gagan D. Gupta, Monica Hasegan, Oumou Goudiam, Renata Basto, Anne-Claude Gingras, and Laurence Pelletier
- Subjects
centrosomes ,cilia ,centrioles ,ciliopathies ,super-resolution microscopy ,Biology (General) ,QH301-705.5 - Abstract
Primary cilia are microtubule-based sensory organelles necessary for efficient transduction of extracellular cues. To initiate cilia formation, ciliary vesicles (CVs) are transported to the vicinity of the centrosome where they dock to the distal end of the mother centriole and fuse to initiate cilium assembly. However, to this date, the early steps in cilia formation remain incompletely understood. Here, we demonstrate functional interplay between CEP19, FOP and CEP350 in ciliogenesis. Using three-dimensional structured-illumination microscopy (3D-SIM) imaging, we mapped the relative spatial distribution of these proteins at the distal end of the mother centriole and show that CEP350/FOP act upstream of CEP19 in their recruitment hierarchy. We demonstrate that CEP19 CRISPR KO cells are severely impaired in their ability to form cilia, analogous to the loss of function of CEP19 binding partners FOP and CEP350. Notably, in the absence of CEP19 microtubule anchoring at centromes is similar in manner to its interaction partners FOP and CEP350. Using GFP-tagged deletion constructs of CEP19, we show that the C-terminus of CEP19 is required for both its localization to centrioles and for its function in ciliogenesis. Critically, this region also mediates the interaction between CEP19 and FOP/CEP350. Interestingly, a morbid-obesity-associated R82* truncated mutant of CEP19 cannot ciliate nor interact with FOP and CEP350, indicative of a putative role for CEP19 in ciliopathies. Finally, analysis of CEP19 KO cells using thin-section electron microscopy revealed marked defects in the docking of CVs to the distal end of the mother centrioles. Together, these data demonstrate a role for the CEP19, FOP and CEP350 module in ciliogenesis and the possible effect of disrupting their functions in ciliopathies.
- Published
- 2017
- Full Text
- View/download PDF
5. Bug22 influences cilium morphology and the post-translational modification of ciliary microtubules
- Author
-
Teresa Mendes Maia, Delphine Gogendeau, Carole Pennetier, Carsten Janke, and Renata Basto
- Subjects
Basal bodies ,Cilia ,Sperm individualization ,Spermatogenesis ,Tubulin post translation modifications ,Science ,Biology (General) ,QH301-705.5 - Abstract
Summary Cilia and flagella are organelles essential for motility and sensing of environmental stimuli. Depending on the cell type, cilia acquire a defined set of functions and, accordingly, are built with an appropriate length and molecular composition. Several ciliary proteins display a high degree of conservation throughout evolution and mutations in ciliary genes are associated with various diseases such as ciliopathies and infertility. Here, we describe the role of the highly conserved ciliary protein, Bug22, in Drosophila. Previous studies in unicellular organisms have shown that Bug22 is required for proper cilia function, but its exact role in ciliogenesis has not been investigated yet. Null Bug22 mutant flies display cilia-associated phenotypes and nervous system defects. Furthermore, sperm differentiation is blocked at the individualization stage, due to impaired migration of the individualization machinery. Tubulin post-translational modifications (PTMs) such as polyglycylation, polyglutamylation or acetylation, are determinants of microtubule (MT) functions and stability in centrioles, cilia and neurons. We found defects in the timely incorporation of polyglycylation in sperm axonemal MTs of Bug22 mutants. In addition, we found that depletion of human Bug22 in RPE1 cells resulted in the appearance of longer cilia and reduced axonemal polyglutamylation. Our work identifies Bug22 as a protein that plays a conserved role in the regulation of PTMs of the ciliary axoneme.
- Published
- 2014
- Full Text
- View/download PDF
6. Experimental Approaches to Generate and Isolate Human Tetraploid Cells
- Author
-
Sara Vanessa Bernhard, Simon Gemble, Renata Basto, and Zuzana Storchova
- Published
- 2023
7. Illuminati: a form of gene expression plasticity in Drosophila neural stem cells
- Author
-
Alix Goupil, Jan Peter Heinen, Riham Salame, Fabrizio Rossi, Jose Reina, Carole Pennetier, Anthony Simon, Patricia Skorski, Anxela Louzao, Allison J. Bardin, Renata Basto, and Cayetano Gonzalez
- Subjects
DNA-Binding Proteins ,Drosophila melanogaster ,Neural Stem Cells ,Mutation ,Animals ,Gene Expression ,Drosophila Proteins ,Drosophila ,Molecular Biology ,Developmental Biology - Abstract
While testing for genome instability in Drosophila as reported by unscheduled upregulation of UAS-GFP in cells that co-express GAL80 and GAL4, we noticed that, as expected, background levels were low in most developing tissues. However, GFP-positive clones were frequent in the larval brain. Most of these clones originated from central brain neural stem cells. Using imaging-based approaches and genome sequencing, we show that these unscheduled clones do not result from chromosome loss or mutations in GAL80. We have named this phenomenon ‘Illuminati’. Illuminati is strongly enhanced in brat tumors and is also sensitive to environmental conditions such as food content and temperature. Illuminati is suppressed by Su(var)2-10, but it is not significantly affected by several modifiers of position effect variegation or Gal4::UAS variegation. We conclude that Illuminati identifies a previously unknown type of functional instability that may have important implications in development and disease.
- Published
- 2022
8. Microvilli-derived Extracellular Vesicles Govern Morphogenesis in Drosophila wing epithelium
- Author
-
Graça Raposo, Laurent Ruel, Gisela D’Angelo, Anne-Sophie Macé, Renata Basto, Maryse Romao, Lucie Sengmanivong, Ilse Hurbain, Pascal P. Thérond, Centre de recherche de l'Institut Curie [Paris], Institut Curie [Paris], Structure and Membrane Compartments [Paris], Biologie Cellulaire et Cancer, Institut Curie [Paris]-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS)-Institut Curie [Paris]-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS), BioImaging Cell and Tissue Core Facility (PICT-IBiSA), Institut de signalisation, biologie du développement et cancer (ISBDC), Centre National de la Recherche Scientifique (CNRS)-Université Nice Sophia Antipolis (... - 2019) (UNS), COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-Université Côte d'Azur (UCA), Institut de Biologie Valrose (IBV), Université Nice Sophia Antipolis (... - 2019) (UNS), COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Côte d'Azur (UCA)-Centre National de la Recherche Scientifique (CNRS), Institut Curie [Paris]-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS), COMUE Université Côte d'Azur (2015 - 2019) (COMUE UCA)-COMUE Université Côte d'Azur (2015 - 2019) (COMUE UCA)-Université Côte d'Azur (UCA), Compartimentation et dynamique cellulaires (CDC), Centre National de la Recherche Scientifique (CNRS)-Institut Curie [Paris]-Université Pierre et Marie Curie - Paris 6 (UPMC), COMUE Université Côte d'Azur (2015 - 2019) (COMUE UCA)-COMUE Université Côte d'Azur (2015 - 2019) (COMUE UCA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université Côte d'Azur (UCA), COMUE Université Côte d'Azur (2015 - 2019) (COMUE UCA)-COMUE Université Côte d'Azur (2015 - 2019) (COMUE UCA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Côte d'Azur (UCA)-Centre National de la Recherche Scientifique (CNRS), COMUE Université Côte d'Azur (2015 - 2019) (COMUE UCA)-COMUE Université Côte d'Azur (2015 - 2019) (COMUE UCA)-Centre National de la Recherche Scientifique (CNRS)-Université Côte d'Azur (UCA), and Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut Curie [Paris]-Centre National de la Recherche Scientifique (CNRS)
- Subjects
0303 health sciences ,animal structures ,Chemistry ,[SDV]Life Sciences [q-bio] ,Morphogenesis ,Extracellular vesicle ,[SDV.BC]Life Sciences [q-bio]/Cellular Biology ,Epithelium ,Cell biology ,03 medical and health sciences ,0302 clinical medicine ,medicine.anatomical_structure ,embryonic structures ,medicine ,Secretion ,Cytoskeleton ,Hedgehog ,030217 neurology & neurosurgery ,Biogenesis ,030304 developmental biology ,Morphogen - Abstract
The regulation and coordination of developmental processes involves the secretion of morphogens and membrane carriers, including extracellular vesicles, which facilitate their transport over long distance. The long-range activity of the Hedgehog morphogen is conveyed by extracellular vesicles. However, the site and the molecular basis of their biogenesis remains unknown. By combining fluorescence and electron microscopy combined with genetics and cell biology approaches, we investigated the origin and the cellular mechanisms underlying extracellular vesicle biogenesis, and their contribution to Drosophila wing disc development, exploiting Hedgehog as a long-range morphogen. We show that microvilli of Drosophila wing disc epithelium are the site of generation of small extracellular vesicles that transport Hedgehog across the tissue. This process requires the Prominin-like protein, whose activity, together with interacting cytoskeleton components and lipids, is critical for maintaining microvilli integrity and function in secretion. Our results provide the first evidence that microvilli-derived extracellular vesicles contribute to Hedgehog long-range signaling activity highlighting their physiological significance in tissue development in vivo.
- Published
- 2021
9. Illuminati, a novel form of gene expression plasticity in Drosophila neural stem cells
- Author
-
Skorski P, Bardin A, Carole Pennetier, Lauzao A, Anthony Simon, Fabrizio Rossi, Alix Goupil, Cayetano Gonzalez, Heinen Jp, Salame R, Renata Basto, Centre de recherche de l'Institut Curie [Paris], and Institut Curie [Paris]
- Subjects
0303 health sciences ,Transgene ,[SDV]Life Sciences [q-bio] ,fungi ,Biology ,Cell cycle ,Neural stem cell ,Green fluorescent protein ,Cell biology ,03 medical and health sciences ,0302 clinical medicine ,Neuroblast ,Gene expression ,Gene ,Psychological repression ,030217 neurology & neurosurgery ,030304 developmental biology - Abstract
With the aim of developing a genetic instability (GI) sensor in vivo we used the well-established Gal80/Gal4-UAS system combined with a visual GFP marker in Drosophila. We generated a collection of 25 Drosophila lines carrying GAL80 transgenes in different locations in all major chromosomes (X, Y, II, and III). We found low rates of GFP cells in epithelial tissues such as wing discs. In contrast, in larval brains, GFP positive clusters containing neural stem cells- also called neuroblasts (NBs)- and their offspring, were highly frequent. Using genetic and imaging-based approaches, we show that GFP NBs do not result from aneuploidy or mutations in the GAL80 gene, but rather by stochastic repression of GAL80 expression. We named this novel type of gene expression instability Illuminati. Importantly, Illuminati frequency is influenced by environmental and stress conditions. Further, we found that once established, Illuminati can be propagated over many cell cycles.
- Published
- 2021
10. Genetic instability from a single S-phase after whole genome duplication
- Author
-
Simon Gemble, René Wardenaar, Kristina Keuper, Nishit Srivastava, Maddalena Nano, Anne-Sophie Macé, Andréa E. Tijhuis, Sara Vanessa Bernhard, Diana C.J. Spierings, Anthony Simon, Oumou Goundiam, Helfrid Hochegger, Matthieu Piel, Floris Foijer, Zuzana Storchová, and Renata Basto
- Subjects
0303 health sciences ,DNA damage ,DNA replication ,Chromosome ,Biology ,Cell biology ,03 medical and health sciences ,0302 clinical medicine ,Centrosome ,Chromosome instability ,Interphase ,Ploidy ,Mitosis ,030217 neurology & neurosurgery ,030304 developmental biology - Abstract
Doubling of the full chromosome content -whole genome duplications (WGDs)- is frequently found in human cancers and is responsible for the rapid evolution of genetically unstable karyotypes 1–3. It has previously been established that WGDs fuel chromosome instability due to abnormal mitosis owing to the presence of extra centrosomes and extra chromosomes 4–8. Tolerance to ploidy changes has been identified in different model organisms and cell types 5,6,9–12, revealing long term cellular adaptations that accommodate ploidy increase. Importantly, however, the immediate consequences of WGDs as cells become tetraploid are not known. It also remains unknown whether WGD triggers other events leading to genetic instability (GIN), independently of mitosis. In this study, we induced tetraploidy in diploid genetically stable RPE-1 cells and monitored the first interphase. We found that newly born tetraploids undergo high rates of DNA damage during DNA replication. Using DNA combing and single cell sequencing, we show that replication forks are unstable, perturbing DNA replication dynamics and generating under- and over-replicated regions at the end of S-phase. Mechanistically, we found that these defects result from lack of protein mass scaling up at the G1/S transition, which impairs the fidelity of DNA replication. This work shows that within a single interphase, unscheduled tetraploid cells can accumulate highly abnormal karyotypes. These findings provide an explanation for the GIN landscape that favors tumorigenesis after tetraploidization.
- Published
- 2021
11. Author Correction: Genetic instability from a single S phase after whole-genome duplication
- Author
-
Simon Gemble, René Wardenaar, Kristina Keuper, Nishit Srivastava, Maddalena Nano, Anne-Sophie Macé, Andréa E. Tijhuis, Sara Vanessa Bernhard, Diana C. J. Spierings, Anthony Simon, Oumou Goundiam, Helfrid Hochegger, Matthieu Piel, Floris Foijer, Zuzana Storchová, and Renata Basto
- Subjects
Multidisciplinary - Published
- 2022
12. Genetic instability from a single S phase after whole-genome duplication
- Author
-
Simon Gemble, René Wardenaar, Kristina Keuper, Nishit Srivastava, Maddalena Nano, Anne-Sophie Macé, Andréa E. Tijhuis, Sara Vanessa Bernhard, Diana C. J. Spierings, Anthony Simon, Oumou Goundiam, Helfrid Hochegger, Matthieu Piel, Floris Foijer, Zuzana Storchová, Renata Basto, Stem Cell Aging Leukemia and Lymphoma (SALL), Damage and Repair in Cancer Development and Cancer Treatment (DARE), and Restoring Organ Function by Means of Regenerative Medicine (REGENERATE)
- Subjects
DNA Replication ,EXPRESSION ,DYNAMICS ,Multidisciplinary ,Karyotype ,DNA-REPLICATION ,Mitosis ,POLYPLOIDY ,S Phase ,Tetraploidy ,E2F ,Chromosomal Instability ,Gene Duplication ,CYTOKINESIS ,G1 ,Humans ,CYCLE ,TRANSCRIPTION ,DNA Damage - Abstract
Diploid and stable karyotypes are associated with health and fitness in animals. By contrast, whole-genome duplications—doublings of the entire complement of chromosomes—are linked to genetic instability and frequently found in human cancers1–3. It has been established that whole-genome duplications fuel chromosome instability through abnormal mitosis4–8; however, the immediate consequences of tetraploidy in the first interphase are not known. This is a key question because single whole-genome duplication events such as cytokinesis failure can promote tumorigenesis9 and DNA double-strand breaks10. Here we find that human cells undergo high rates of DNA damage during DNA replication in the first S phase following induction of tetraploidy. Using DNA combing and single-cell sequencing, we show that DNA replication dynamics is perturbed, generating under- and over-replicated regions. Mechanistically, we find that these defects result from a shortage of proteins during the G1/S transition, which impairs the fidelity of DNA replication. This work shows that within a single interphase, unscheduled tetraploid cells can acquire highly abnormal karyotypes. These findings provide an explanation for the genetic instability landscape that favours tumorigenesis after tetraploidization.
- Published
- 2022
13. Centrosomes in disease: how the same music can sound so different?
- Author
-
Oumou Goundiam, Renata Basto, and Institute Curie
- Subjects
Tumor suppressor gene ,[SDV]Life Sciences [q-bio] ,Cell ,Mitosis ,Biology ,03 medical and health sciences ,0302 clinical medicine ,Structural Biology ,medicine ,Animals ,Humans ,Molecular Biology ,Tissue homeostasis ,ComputingMilieux_MISCELLANEOUS ,030304 developmental biology ,Centrosome ,0303 health sciences ,Cell Cycle ,Cell migration ,Microtubule organizing center ,Cell cycle ,Cell biology ,medicine.anatomical_structure ,Microtubule-Organizing Center ,Music ,030217 neurology & neurosurgery - Abstract
Centrosomes are the major microtubule organizing center of animal cells. Centrosomes contribute to timely bipolar spindle assembly during mitosis and participate in the regulation of other processes such as polarity establishment and cell migration. Centrosome numbers are tightly controlled during the cell cycle to ensure that mitosis is initiated with only two centrosomes. Deviations in centrosome number or structure are known to impact cell or tissue homeostasis and can impact different processes as diverse as proliferation, death or disease. Interestingly, defects in centrosome number seem to culminate with common responses, which depend on p53 activation even in different contexts such as development or cancer. p53 is a tumor suppressor gene with essential roles in the maintenance of genetic stability normally stimulated by various cellular stresses. Here, we review current knowledge and discuss how defects in centrosome structure and number can lead to different human pathologies.
- Published
- 2021
14. Drosophilaneural stem cells show a unique dynamic pattern of gene expression that is influenced by environmental factors
- Author
-
Anthony Simon, Allison J. Bardin, Alix Goupil, Carole Pennetier, Patricia Skorski, and Renata Basto
- Subjects
biology ,In vivo ,Gene expression ,medicine ,Aneuploidy ,Gene silencing ,Epigenetics ,biology.organism_classification ,medicine.disease ,Drosophila ,Neural stem cell ,Green fluorescent protein ,Cell biology - Abstract
With the aim of developing a sensor for chromosome lossin vivo, we used the well-established GAL4/GAL80 system combined with a visual GFP marker inDrosophila. We show a low frequency of green cells in mostDrosophilatissues, suggesting low aneuploidy levels. Unexpectedly, in the brain, GFP positive cells are more frequent, but in this case, they do not represent chromosome loss. Using genetic manipulations, RNA FISH and time-lapse microscopy, we uncovered a dynamic and reversible silencing ofGAL80that occurs inDrosophilaneural stem cells. Further, we showed that this novel gene expression regulation is influenced by environmental changes such as temperature variations or food composition. These results have important implications for theDrosophilacommunity, namely the possible interpretation of false positive cells in clonal experiments. Additionally, they also highlight a level of mosaicism and plasticity in the brain, consistent with possible epigenetic regulation of fly chromosomes, which is different from other organs and tissues.
- Published
- 2020
15. Microvilli-derived extracellular vesicles carry Hedgehog morphogenic signals for Drosophila wing imaginal disc development
- Author
-
Ilse Hurbain, Anne-Sophie Macé, Maryse Romao, Elodie Prince, Lucie Sengmanivong, Laurent Ruel, Renata Basto, Pascal P. Thérond, Graça Raposo, Gisela D’Angelo, Biologie Cellulaire et Cancer, Institut Curie [Paris]-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS), Structure and Membrane Compartments [Paris], Institut Curie [Paris]-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS)-Institut Curie [Paris]-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS), Institut de Biologie Valrose (IBV), Université Nice Sophia Antipolis (... - 2019) (UNS), COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Côte d'Azur (UCA)-Centre National de la Recherche Scientifique (CNRS), Institut de signalisation, biologie du développement et cancer (ISBDC), Centre National de la Recherche Scientifique (CNRS)-Université Nice Sophia Antipolis (... - 2019) (UNS), and COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-Université Côte d'Azur (UCA)
- Subjects
0303 health sciences ,Microvilli ,[SDV.BC]Life Sciences [q-bio]/Cellular Biology ,General Biochemistry, Genetics and Molecular Biology ,Extracellular Vesicles ,03 medical and health sciences ,Drosophila melanogaster ,0302 clinical medicine ,Imaginal Discs ,[SDV.BC.IC]Life Sciences [q-bio]/Cellular Biology/Cell Behavior [q-bio.CB] ,Morphogenesis ,Animals ,Drosophila Proteins ,Wings, Animal ,Drosophila ,Hedgehog Proteins ,AC133 Antigen ,General Agricultural and Biological Sciences ,[SDV.BDD]Life Sciences [q-bio]/Development Biology ,ComputingMilieux_MISCELLANEOUS ,030217 neurology & neurosurgery ,030304 developmental biology - Abstract
Morphogens are secreted molecules that regulate and coordinate major developmental processes, such as cell differentiation and tissue morphogenesis. Depending on the mechanisms of secretion and the nature of their carriers, morphogens act at short and long range. We investigated the paradigmatic long-range activity of Hedgehog (Hh), a well-known morphogen, and its contribution to the growth and patterning of the Drosophila wing imaginal disc. Extracellular vesicles (EVs) contribute to Hh long-range activity; however, the nature, the site, and the mechanisms underlying the biogenesis of these vesicular carriers remain unknown. Here, through the analysis of mutants and a series of Drosophila RNAi-depleted wing imaginal discs using fluorescence and live-imaging electron microscopy, including tomography and 3D reconstruction, we demonstrate that microvilli of the wing imaginal disc epithelium are the site of generation of small EVs that transport Hh across the tissue. Further, we show that the Prominin-like (PromL) protein is critical for microvilli integrity. Together with actin cytoskeleton and membrane phospholipids, PromL maintains microvilli architecture that is essential to promote its secretory function. Importantly, the distribution of Hh to microvilli and its release via these EVs contribute to the proper morphogenesis of the wing imaginal disc. Our results demonstrate that microvilli-derived EVs are carriers for Hh long-range signaling in vivo. By establishing that members of the Prominin protein family are key determinants of microvilli formation and integrity, our findings support the view that microvilli-derived EVs conveying Hh may provide a means for exchanging signaling cues of high significance in tissue development and cancer.
- Published
- 2022
16. Centrosomes: The good and the bad for brain development
- Author
-
Renata Basto, Véronique Marthiens, Centre de recherche de l'Institut Curie [Paris], and Institut Curie [Paris]
- Subjects
Brain development ,[SDV]Life Sciences [q-bio] ,Mitosis ,Spindle Apparatus ,Biology ,03 medical and health sciences ,0302 clinical medicine ,Chromosome Segregation ,Organelle ,Animals ,Humans ,Organism ,ComputingMilieux_MISCELLANEOUS ,030304 developmental biology ,Centrosome ,0303 health sciences ,Brain ,Structural integrity ,Cell Biology ,General Medicine ,Neural stem cell ,Cell biology ,Brain growth ,Neurodevelopmental Disorders ,Brain size ,030217 neurology & neurosurgery - Abstract
Centrosomes nucleate and organise the microtubule cytoskeleton in animal cells. These membraneless organelles are key structures for tissue organisation, polarity and growth. Centrosome dysfunction, defined as deviation in centrosome numbers and/or structural integrity, has major impact on brain size and functionality, as compared with other tissues of the organism. In this review, we discuss the contribution of centrosomes to brain growth during development. We discuss in particular the impact of centrosome dysfunction in Drosophila and mammalian neural stem cell division and fitness, which ultimately underlie brain growth defects.
- Published
- 2020
17. Chromosomes function as a barrier to mitotic spindle bipolarity in polyploid cells
- Author
-
Carole Pennetier, Oumou Goundiam, Frances Edwards, Gaëlle Letort, Delphine Gogendeau, Maddalena Nano, Simon Gemble, Alix Goupil, Renata Basto, Centre de recherche de l'Institut Curie [Paris], Institut Curie [Paris], Biologie Cellulaire et Cancer, Institut Curie [Paris]-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS), Centre interdisciplinaire de recherche en biologie (CIRB), Labex MemoLife, École normale supérieure - Paris (ENS-PSL), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Collège de France (CdF (institution))-Ecole Superieure de Physique et de Chimie Industrielles de la Ville de Paris (ESPCI Paris), Université Paris sciences et lettres (PSL)-École normale supérieure - Paris (ENS-PSL), Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Basto, Renata, École normale supérieure - Paris (ENS Paris), Université Paris sciences et lettres (PSL)-École normale supérieure - Paris (ENS Paris), and Curie Institute
- Subjects
[SDV]Life Sciences [q-bio] ,Spindle Apparatus ,[SDV.BC.BC]Life Sciences [q-bio]/Cellular Biology/Subcellular Processes [q-bio.SC] ,Biology ,Spindle pole body ,Article ,Polyploidy ,03 medical and health sciences ,0302 clinical medicine ,Polyploid ,Chromosome instability ,[SDV.BC.BC] Life Sciences [q-bio]/Cellular Biology/Subcellular Processes [q-bio.SC] ,Animals ,Humans ,Mitosis ,Cells, Cultured ,ComputingMilieux_MISCELLANEOUS ,030304 developmental biology ,Cancer ,Centrosome ,0303 health sciences ,fungi ,Chromosome ,food and beverages ,Cell Biology ,Spindle apparatus ,Cell biology ,HEK293 Cells ,nervous system ,Drosophila ,Female ,Ploidy ,biological phenomena, cell phenomena, and immunity ,030217 neurology & neurosurgery ,Cell Cycle and Division - Abstract
Polyploid cells with extra centrosomes and extra DNA divide in a multipolar manner. Using Drosophila and cancer cells challenged in different ways and in silico modeling, Goupil et al. show that chromosomes act as a barrier inhibiting spindle pole coalescence and favoring multipolar spindle assembly., Ploidy variations such as genome doubling are frequent in human tumors and have been associated with genetic instability favoring tumor progression. How polyploid cells deal with increased centrosome numbers and DNA content remains unknown. Using Drosophila neuroblasts and human cancer cells to study mitotic spindle assembly in polyploid cells, we found that most polyploid cells divide in a multipolar manner. We show that even if an initial centrosome clustering step can occur at mitotic entry, the establishment of kinetochore-microtubule attachments leads to spatial chromosome configurations, whereby the final coalescence of supernumerary poles into a bipolar array is inhibited. Using in silico approaches and various spindle and DNA perturbations, we show that chromosomes act as a physical barrier blocking spindle pole coalescence and bipolarity. Importantly, microtubule stabilization suppressed multipolarity by improving both centrosome clustering and pole coalescence. This work identifies inhibitors of bipolar division in polyploid cells and provides a rationale to understand chromosome instability typical of polyploid cancer cells.
- Published
- 2020
18. Cell-Cycle Asynchrony Generates DNA Damage at Mitotic Entry in Polyploid Cells
- Author
-
Vincent Fraisier, Carole Pennetier, Anthony Simon, Renata Basto, Maddalena Nano, Véronique Marthiens, Simon Gemble, Lipides - Nutrition - Cancer (U866) (LNC), Université de Bourgogne (UB)-Institut National de la Santé et de la Recherche Médicale (INSERM)-AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Ecole Nationale Supérieure de Biologie Appliquée à la Nutrition et à l'Alimentation de Dijon (ENSBANA), Physico-Chimie-Curie (PCC), Centre National de la Recherche Scientifique (CNRS)-Institut Curie [Paris]-Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut de Chimie du CNRS (INC), Compartimentation et dynamique cellulaires (CDC), and Centre National de la Recherche Scientifique (CNRS)-Institut Curie [Paris]-Université Pierre et Marie Curie - Paris 6 (UPMC)
- Subjects
0301 basic medicine ,Male ,DNA damage ,[SDV]Life Sciences [q-bio] ,Mitosis ,Biology ,General Biochemistry, Genetics and Molecular Biology ,Polyploidy ,03 medical and health sciences ,Mice ,0302 clinical medicine ,Neuroblast ,Polyploid ,Neural Stem Cells ,Cell Line, Tumor ,Gene duplication ,Animals ,Drosophila Proteins ,Humans ,ComputingMilieux_MISCELLANEOUS ,Cytokinesis ,Cell Cycle ,Chromosome ,Cell cycle ,Cell biology ,Mice, Inbred C57BL ,030104 developmental biology ,Drosophila melanogaster ,Female ,General Agricultural and Biological Sciences ,030217 neurology & neurosurgery ,DNA Damage - Abstract
Summary Polyploidy arises from the gain of complete chromosome sets [ 1 ], and it is known to promote cancer genome evolution. Recent evidence suggests that a large proportion of human tumors experience whole-genome duplications (WGDs), which might favor the generation of highly abnormal karyotypes within a short time frame, rather than in a stepwise manner [ 2 , 3 , 4 , 5 , 6 ]. However, the molecular mechanisms linking whole-genome duplication to genetic instability remain poorly understood. Using repeated cytokinesis failure to induce polyploidization of Drosophila neural stem cells (NSCs) (also called neuroblasts [NBs]), we investigated the consequences of polyploidy in vivo. Surprisingly, we found that DNA damage is generated in a subset of nuclei of polyploid NBs during mitosis. Importantly, our observations in flies were confirmed in mouse NSCs (mNSCs) and human cancer cells after acute cytokinesis inhibition. Interestingly, DNA damage occurs in nuclei that were not ready to enter mitosis but were forced to do so when exposed to the mitotic environment of neighboring nuclei within the same cell. Additionally, we found that polyploid cells are cell-cycle asynchronous and forcing cell-cycle synchronization was sufficient to lower the levels of DNA damage generated during mitosis. Overall, this work supports a model in which DNA damage at mitotic entry can generate DNA structural abnormalities that might contribute to the onset of genetic instability.
- Published
- 2019
19. Topical Delivery of Niacinamide to Skin Using Hybrid Nanogels Enhances Photoprotection Effect
- Author
-
Salette Reis, Sofia A. Costa Lima, Cláudia Nunes, Renata Basto, and Raquel G D Andrade
- Subjects
integumentary system ,Polyvinylpyrrolidone ,Jojoba oil ,Chemistry ,in vitro release ,jojoba oil ,permeation enhancers ,Pharmaceutical Science ,Permeation ,tween 80 ,Article ,UV radiation ,RS1-441 ,Pharmacy and materia medica ,medicine.anatomical_structure ,oleic acid ,transethosomes ,Niacinamide ,Self-healing hydrogels ,Stratum corneum ,medicine ,Zeta potential ,Biophysics ,medicine.drug ,Nanogel - Abstract
Niacinamide (NIA) has been widely used in halting the features of ageing by acting as an antioxidant and preventing dehydration. NIA’s physicochemical properties suggest difficulties in surpassing the barrier imposed by the stratum corneum layer to reach the target in the skin. To improve cutaneous delivery of NIA, a hybrid nanogel was designed using carrageenan and polyvinylpyrrolidone polymers combined with jojoba oil as a permeation enhancer. Three different types of transethosomes were prepared by the thin-film hydration method, made distinct by the presence of either an edge activator or a permeation enhancer, to allow for a controlled delivery of NIA. Formulations were characterized by measurements of size, polydispersity index, zeta potential, encapsulation efficiency, and loading capacity, and by evaluating their chemical interactions and morphology. Skin permeation assays were performed using Franz diffusion cells. The hybrid hydrogels exhibited robust, porous, and highly aligned macrostructures, and when present, jojoba oil changed their morphology. Skin permeation studies with transethosomes-loaded hydrogels showed that nanogels per se exhibit a more controlled and enhanced permeation, in particular when jojoba oil was present in the transethosomes. These promising nanogels protected the human keratinocytes from UV radiation, and thus can be added to sunscreens or after-sun lotions to improve skin protection.
- Published
- 2021
20. Plk4 Regulates Centriole Asymmetry and Spindle Orientation in Neural Stem Cells
- Author
-
Anthony Simon, Yuu Kimata, Maddalena Nano, Victor Racine, Daniel W. Buster, Alix Goupil, John M. Ryniawec, Carole Pennetier, Davide Gambarotto, Renata Basto, Delphine Gogendeau, Gregory C. Rogers, Damien Blanc, Centre de recherche de l'Institut Curie [Paris], Institut Curie [Paris], Department of Cell Biology, Sciences III, University of Geneva, Geneva, Switzerland, Compartimentation et dynamique cellulaires (CDC), Centre National de la Recherche Scientifique (CNRS)-Institut Curie [Paris]-Université Pierre et Marie Curie - Paris 6 (UPMC), Centre de génétique moléculaire (CGM), Centre National de la Recherche Scientifique (CNRS), Physico-Chimie-Curie (PCC), Centre National de la Recherche Scientifique (CNRS)-Institut Curie [Paris]-Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut de Chimie du CNRS (INC), Institute of Molecular and Cell Biology - Molecular controls of Morphogenesis and Tumor Progression, and Institute of Molecular and Cell Biology
- Subjects
Male ,PLK4 ,Centriole ,[SDV]Life Sciences [q-bio] ,Spindle Apparatus ,Protein Serine-Threonine Kinases ,centrosome positioning ,Biology ,symmetry breaking ,Cdh1 Proteins ,Article ,General Biochemistry, Genetics and Molecular Biology ,03 medical and health sciences ,0302 clinical medicine ,Neural Stem Cells ,spindle orientation ,Animals ,Drosophila Proteins ,Phosphorylation ,centrosomes ,Molecular Biology ,Cells, Cultured ,ComputingMilieux_MISCELLANEOUS ,Centrioles ,030304 developmental biology ,Centrosome ,0303 health sciences ,Astral microtubule nucleation ,Apical cortex ,Cell Cycle ,Cell Biology ,Spd2 ,Neural stem cell ,Cell biology ,Drosophila melanogaster ,Plk4 ,Female ,Stem cell ,Developmental biology ,030217 neurology & neurosurgery ,Developmental Biology - Abstract
Summary Defects in mitotic spindle orientation (MSO) disrupt the organization of stem cell niches impacting tissue morphogenesis and homeostasis. Mutations in centrosome genes reduce MSO fidelity, leading to tissue dysplasia and causing several diseases such as microcephaly, dwarfism, and cancer. Whether these mutations perturb spindle orientation solely by affecting astral microtubule nucleation or whether centrosome proteins have more direct functions in regulating MSO is unknown. To investigate this question, we analyzed the consequences of deregulating Plk4 (the master centriole duplication kinase) activity in Drosophila asymmetrically dividing neural stem cells. We found that Plk4 functions upstream of MSO control, orchestrating centriole symmetry breaking and consequently centrosome positioning. Mechanistically, we show that Plk4 acts through Spd2 phosphorylation, which induces centriole release from the apical cortex. Overall, this work not only reveals a role for Plk4 in regulating centrosome function but also links the centrosome biogenesis machinery with the MSO apparatus., Graphical Abstract, Highlights • Drosophila Plk4 mutant NSCs show defects in centriole asymmetry and spindle positioning • Apical centriole anchoring requires the PCM protein Spd-2 and the APC/C activator Fzr • Movement of the centriole toward the basal side of the cell requires Plk4 activity • At the mother centriole, Plk4 phosphorylates Spd2 to trigger PCM shedding and Fzr loss, Mitotic spindle orientation is tightly regulated during development and adulthood to maintain tissue organization and homeostasis. Spindle orientation requires the coordination between centrosomes and cortical cues. Gambarotto et al. report that the centrosome components Plk4 and Spd2 regulate centrosome asymmetry in interphase to influence spindle positioning in mitosis.
- Published
- 2019
21. Low centrosome numbers correlate with higher aggressivity in ovarian cancer
- Author
-
Anne Vincent-Salomon, Xavier Sastre-Garau, Guillaume Bataillon, Fatima Mechta-Grigoriou, Pierre Gestraud, B. Mboup, Odette Mariani, Fariba Nemati, Didier Decaudin, Véronique Becette, J.-P. Morretton, S. Roman Roman, Claire Bonneau, Renata Basto, Marc-Henri Stern, Aurélie Herbette, Tatiana Popova, Didier Meseure, Oumou Goundiam, Roman Rouzier, André Nicolas, C. Cosson, Jorge Barbazan, and Aurélien Latouche
- Subjects
0303 health sciences ,Chemotherapy ,medicine.medical_treatment ,Cell ,Chromosome ,Aneuploidy ,Biology ,medicine.disease ,medicine.disease_cause ,03 medical and health sciences ,0302 clinical medicine ,medicine.anatomical_structure ,Centrosome ,Tumor progression ,030220 oncology & carcinogenesis ,medicine ,Cancer research ,Carcinogenesis ,Ovarian cancer ,030304 developmental biology - Abstract
Centrosome amplification has been described as a common feature of human cancers and it is known to promote tumorigenesis when induced in animals. However, little is known about the real status of centrosome numbers in human cancers and whether numerical alterations are solely associated with poor prognosis. To address this question, we have analyzed a large cohort of human epithelial ovarian cancers (EOCs) from 100 patients using state-of-the-art microscopy to determine the Centrosome-Nucleus Index (CNI) of each tumor. We found that EOCs are highly heterogeneous, with infrequent but strong centrosome amplifications leading to higher CNI than in healthy tissues. Strikingly, while a correlation between CNI and genomic alterations, such as aneuploidy or chromosome rearrangements could not be established, we found that high CNI correlates with increased patient survival and sensitivity to chemotherapy. Using ovarian cancer cellular models to manipulate centrosome numbers and Patient-Derived Xenografts (PDXs), we found that higher CNIs can positively impact the response to chemotherapy and inhibit cell dissemination. Our findings highlight a novel paradigm linking centrosome amplification to the inhibition of tumor progression.
- Published
- 2019
22. Cell Cycle Asynchrony Generates DNA Damage at Mitotic Entry in Polyploid Cells
- Author
-
Maddalena Nano, Renata Basto, Carole Pennetier, Véronique Marthiens, Anthony Simon, and Vincent Fraisier
- Subjects
0303 health sciences ,DNA damage ,030302 biochemistry & molecular biology ,Biology ,Cell cycle ,Genome ,Cell biology ,03 medical and health sciences ,Polyploid ,Neuroblast ,Chromosome instability ,Mitosis ,Cytokinesis ,030304 developmental biology - Abstract
Polyploidy arises from the gain of complete chromosomes sets [1] and is known to promote cancer genome evolution. Recent evidence suggests that a large proportion of human tumours experience whole genome duplications (WGDs), which might favour the generation of highly abnormal karyotypes within a short time frame, rather than in a stepwise manner [2–6]. However, the molecular mechanisms linking whole genome duplication to genetic instability remain poorly understood. Further, possible mechanisms responsible for rapid genome reshuffling have not been described yet. Using repeated cytokinesis failure to induce polyploidization ofDrosophilaneural stem cells (NSCs, also called neuroblasts - NBs), we investigated the consequences of polyploidyin vivo. Here, we show that polyploid NSCs accumulate high levels of chromosome instability. Surprisingly, we found that DNA damage is generated in a subset of nuclei of polyploid NBs during mitosis, in an asymmetric manner. Importantly, our observations in flies were confirmed in mouse NSCs (mNSCs) after acute cytokinesis inhibition. Interestingly, DNA damage occurs in nuclei that were not ready to enter mitosis but were forced to do so when exposed to the mitotic environment of neighbouring nuclei within the same cell. Additionally, we found that polyploid cells are cell cycle asynchronous and forcing cell cycle synchronization is sufficient to lower the levels of DNA damage generated during mitosis. Overall, this work supports a model in which DNA damage at mitotic entry can generate a mutated genetic landscape that contributes to the onset of genetic instability.
- Published
- 2019
23. Chromosomes function as a barrier to mitotic spindle bipolarity in polyploid cells
- Author
-
Delphine Gogendeau, Maddalena Nano, Alix Goupil, Carole Pennetier, Gaëlle Letort, and Renata Basto
- Subjects
Centrosome ,Microtubule ,Chromosome instability ,Chromosome ,Biology ,Mitosis ,Cytokinesis ,Spindle apparatus ,Chromatin ,Cell biology - Abstract
Whole genome duplications (WGDs) are found in a variety of tumors and are associated with chromosomal instability (CIN) and poor prognosis [1,2]. When induced experimentally, through cytokinesis failure, polyploid cells generate tumors [3]. Cytokinesis failure results in the accumulation of double DNA content, but also of cytoplasmic organelles, such as centrosomes, which are the major microtubule (MT) organizing centers of animal cells. Importantly, even if there is a correlation between polyploidy and CIN [4], the underlying mechanisms generating error-prone mitosis in cells with extra DNA and extra centrosomes are not known. When considering polyploid mitosis, it is essential to take into account the increase in MT nucleation due to the presence of extra centrosomes and extra DNA. The presence of supernumerary centrosomes in a cell, centrosome amplification [5], is associated with mitotic spindle multipolarity and CIN [6–9]. Importantly, additional MTs can be nucleated from the chromatin (chromatin mediated pathway-CMP) or from pre-existing MTs-through the Augmin pathway. We hypothesized that the increase in DNA and centrosome content in a cell could lead to an increased MT mass, which might account for abnormal mitosis described in polyploid cells [4, 10, 11, 12]. Using genetics, live imaging and modeling approaches, we investigated the mechanisms establishing multipolarity in vivo in polyploid cells. We found that MT nucleation from the centrosomes is the major contributor to multipolarity, while other pathways seem to play minor roles. Unexpectedly, we found that even if Ncd/HSET, plays an essential role in promoting centrosome clustering in early mitosis, the increase in chromosome mass associated with cytokinesis failure functions as a barrier to centrosome clustering into two main poles. Our work provides a mechanistic link between polyploidy and the generation of CIN.
- Published
- 2019
- Full Text
- View/download PDF
24. Un dysfonctionnement du centromère compromet l'intégrité du pôle mitotique
- Author
-
Geneviève Almouzni, Karen Oegema, Raphaël Rodriguez, Marie Dumont, Anthony Simon, Solène Hervé, Daniele Fachinetti, Carole Pennetier, Franz Meitinger, Renata Basto, Simon Gemble, Dynamique du noyau [Institut Curie], Institut Curie [Paris]-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS), Centre de recherche de l'Institut Curie [Paris], Institut Curie [Paris], Lipides - Nutrition - Cancer (U866) (LNC), Université de Bourgogne (UB)-Institut National de la Santé et de la Recherche Médicale (INSERM)-AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Ecole Nationale Supérieure de Biologie Appliquée à la Nutrition et à l'Alimentation de Dijon (ENSBANA), Physico-Chimie-Curie (PCC), Centre National de la Recherche Scientifique (CNRS)-Institut Curie [Paris]-Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut de Chimie du CNRS (INC), Compartimentation et dynamique cellulaires (CDC), Centre National de la Recherche Scientifique (CNRS)-Institut Curie [Paris]-Université Pierre et Marie Curie - Paris 6 (UPMC), Centre national de recherches météorologiques (CNRM), Institut national des sciences de l'Univers (INSU - CNRS)-Météo France-Centre National de la Recherche Scientifique (CNRS), Ludwig Institute for Cancer Research, Institut de Chimie des Substances Naturelles (ICSN), Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie du CNRS (INC), Dynamique nucléaire et plasticité du génome (DNPG), and Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut Curie [Paris]-Centre National de la Recherche Scientifique (CNRS)
- Subjects
0301 basic medicine ,Centriole ,Chromosomal Proteins, Non-Histone ,[SDV]Life Sciences [q-bio] ,Kinetochore assembly ,Centromere ,Mitosis ,[SDV.BC]Life Sciences [q-bio]/Cellular Biology ,Spindle Apparatus ,Biology ,Microtubules ,General Biochemistry, Genetics and Molecular Biology ,Spindle pole body ,Cell Line ,Histones ,03 medical and health sciences ,0302 clinical medicine ,mitotic spindle pole integrity ,Centromere Protein A ,Chromosome Segregation ,[SDV.BC.IC]Life Sciences [q-bio]/Cellular Biology/Cell Behavior [q-bio.CB] ,chromosome mis-segregation ,Humans ,Spindle Poles ,centrosomes ,Kinetochores ,ComputingMilieux_MISCELLANEOUS ,Centrioles ,Centrosome ,centromeres ,microtubule dynamics ,Cell biology ,Spindle apparatus ,030104 developmental biology ,Mitotic spindle pole ,General Agricultural and Biological Sciences ,030217 neurology & neurosurgery - Abstract
International audience; Centromeres and centrosomes are crucial mitotic players. Centromeres are unique chromosomal sites characterized by the presence of the histone H3-variant centromere protein A (CENP-A) [1]. CENP-A recruits the majority of centromere components, collectively named the constitutive centromere associated network (CCAN) [2]. The CCAN is necessary for kinetochore assembly, a multiprotein complex that attaches spindle microtubules (MTs) and is required for chromosome segregation [3]. In most animal cells, the dominant site for MT nucleation in mitosis are the centrosomes, which are composed of two centrioles, surrounded by a protein-rich matrix of electron-dense pericentriolar material (PCM) [4]. The PCM is the site of MT nucleation during mitosis [5]. Even if centromeres and centrosomes are connected via MTs in mitosis, it is not known whether defects in either one of the two structures have an impact on the function of the other. Here, using high-resolution microscopy combined with rapid removal of CENP-A in human cells, we found that perturbation of centromere function impacts mitotic spindle pole integrity. This includes release of MT minus-ends from the centrosome, leading to PCM dispersion and centriole mis-positioning at the spindle poles. Mechanistically, we show that these defects result from abnormal spindle MT dynamics due to defective kinetochore-MT attachments. Importantly, restoring mitotic spindle pole integrity following centromere inactivation lead to a decrease in the frequency of chromosome mis-segregation. Overall, our work identifies an unexpected relationship between centromeres and maintenance of the mitotic pole integrity necessary to ensure mitotic accuracy and thus to maintain genetic stability.; Les centromères et les centrosomes sont des acteurs mitotiques essentiels. Les centromères sont des sites chromosomiques uniques caractérisés par la présence de l'histone H3-variante de la protéine centromère A (CENP-A) [1]. La CENP-A recrute la majorité des composants des centromères, collectivement appelés le réseau constitutif associé aux centromères (CCAN) [2]. Le CCAN est nécessaire à l'assemblage des cinétochores, un complexe multiprotéique qui fixe les microtubules du fuseau (MT) et est requis pour la ségrégation des chromosomes [3]. Dans la plupart des cellules animales, le site dominant pour la nucléation des MT dans la mitose sont les centrosomes, qui sont composés de deux centrioles, entourés par une matrice riche en protéines de matériel péricentriolaire dense en électrons (PCM) [4]. Le PCM est le site de nucléation de la MT pendant la mitose [5]. Même si les centromères et les centrosomes sont reliés par des MT lors de la mitose, on ne sait pas si des défauts dans l'une des deux structures ont un impact sur la fonction de l'autre. Ici, en utilisant la microscopie à haute résolution combinée à l'élimination rapide du CENP-A dans les cellules humaines, nous avons découvert que la perturbation de la fonction des centromères a un impact sur l'intégrité des pôles du fuseau mitotique. Cela inclut la libération des extrémités négatives de la MT du centrosome, ce qui entraîne une dispersion de la PCM et un mauvais positionnement du centriole au niveau des pôles du fuseau. Sur le plan mécanique, nous montrons que ces défauts résultent d'une dynamique anormale de la MT du fuseau due à des fixations cinétochores-MT défectueuses. Il est important de noter que la restauration de l'intégrité des pôles du fuseau mitotique après l'inactivation des centromères entraîne une diminution de la fréquence de la mauvaise ségrégation des chromosomes. Dans l'ensemble, notre travail identifie une relation inattendue entre les centromères et le maintien de l'intégrité du pôle mitotique nécessaire pour assurer la précision mitotique et donc pour maintenir la stabilité génétique.
- Published
- 2018
25. Differences in Mitotic Spindle Architecture in Mammalian Neural Stem Cells Influence Mitotic Accuracy during Brain Development
- Author
-
Nathalie Da Silva, Alexandre D Baffet, Carole Pennetier, Jean-Baptiste Brault, Tristan Piolot, Véronique Marthiens, Ludovic Leconte, Renata Basto, Diana Vargas-Hurtado, CHU Rothschild [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU), Compartimentation et dynamique cellulaires (CDC), Institut Curie [Paris]-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS), Muséfrem, Centre d'Histoire 'Espaces et Cultures' (CHEC), Université Blaise Pascal - Clermont-Ferrand 2 (UBP)-Université Blaise Pascal - Clermont-Ferrand 2 (UBP), Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut Curie [Paris]-Centre National de la Recherche Scientifique (CNRS), Virology Laboratory, Hôpital Rothchild, and Centre National de la Recherche Scientifique (CNRS)-Institut Curie [Paris]-Université Pierre et Marie Curie - Paris 6 (UPMC)
- Subjects
0301 basic medicine ,Male ,Cell type ,[SDV]Life Sciences [q-bio] ,Neurogenesis ,Mitosis ,Cell Cycle Proteins ,Spindle Apparatus ,Biology ,Microtubules ,General Biochemistry, Genetics and Molecular Biology ,Chromosome segregation ,03 medical and health sciences ,Mice ,0302 clinical medicine ,Neural Stem Cells ,Microtubule ,Pregnancy ,Chromosome Segregation ,Animals ,Kinetochores ,ComputingMilieux_MISCELLANEOUS ,Mammals ,Embryogenesis ,Brain ,Embryonic stem cell ,Neural stem cell ,Cell biology ,Spindle apparatus ,Mice, Inbred C57BL ,030104 developmental biology ,Female ,General Agricultural and Biological Sciences ,Microtubule-Associated Proteins ,030217 neurology & neurosurgery - Abstract
Summary A functional bipolar spindle is essential to segregate chromosomes correctly during mitosis. Across organisms and cell types, spindle architecture should be optimized to promote error-free divisions. However, it remains to be investigated whether mitotic spindle morphology adapts to changes in tissue properties, typical of embryonic development, in order to ensure different tasks, such as spindle positioning and chromosome segregation. We have characterized mitotic spindles in neural stem cells (NSCs) of the embryonic developing mouse neocortex. Surprisingly, we found a switch in spindle morphology from early to late neurogenic stages, which relies on an increase in inner spindle microtubule density and stability. Mechanistically, we identified the microtubule-associated protein TPX2 as one determinant of spindle shape, contributing not only to its robustness but also to correct chromosome segregation upon mitotic challenge. Our findings highlight a possible causal relationship between spindle architecture and mitotic accuracy with likely implications in brain size regulation.
- Published
- 2018
26. Chromosome structural anomalies due to aberrant spindle forces exerted at gene editing sites in meiosis
- Author
-
Marie-Hélène Verlhac, Joanne Kanaan, Marion Manil-Ségalen, Małgorzata Łuksza, Simon I. R. Lane, Marie-Emilie Terret, Keith T. Jones, Renata Basto, Véronique Marthiens, Centre interdisciplinaire de recherche en biologie (CIRB), Labex MemoLife, École normale supérieure - Paris (ENS Paris), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Collège de France (CdF (institution))-Ecole Superieure de Physique et de Chimie Industrielles de la Ville de Paris (ESPCI Paris), Université Paris sciences et lettres (PSL)-École normale supérieure - Paris (ENS Paris), Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Biologie Cellulaire et Cancer, Institut Curie [Paris]-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS), University of Southampton, École normale supérieure - Paris (ENS-PSL), Université Paris sciences et lettres (PSL)-École normale supérieure - Paris (ENS-PSL), Basto, Renata, Institut de Biologie Intégrative de la Cellule (I2BC), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS), Institut de biologie de l'ENS Paris (UMR 8197/1024) (IBENS), Département de Biologie - ENS Paris, Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-École normale supérieure - Paris (ENS Paris), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Collège de France (CdF (institution))-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Compartimentation et dynamique cellulaires (CDC), Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut Curie [Paris]-Centre National de la Recherche Scientifique (CNRS), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-École normale supérieure - Paris (ENS Paris), and Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)
- Subjects
0301 basic medicine ,PLK4 ,[SDV]Life Sciences [q-bio] ,Mice, Transgenic ,Spindle Apparatus ,[SDV.BC]Life Sciences [q-bio]/Cellular Biology ,Biology ,Article ,Spindle pole body ,Chromosome segregation ,Mice ,03 medical and health sciences ,Meiosis ,Microtubule ,Animals ,Research Articles ,ComputingMilieux_MISCELLANEOUS ,Microtubule nucleation ,Gene Editing ,Chromosome ,Cell Biology ,Chromosomes, Mammalian ,Cell biology ,[SDV] Life Sciences [q-bio] ,030104 developmental biology ,Oocytes ,Female ,Chromosome breakage ,Microtubule-Organizing Center - Abstract
Acentrosomal spindle assembly in mouse oocytes depends on chromosomes and acentriolar microtubule-organizing centers (aMTOCs). Manil-Ségalen et al. observe that Plk4-induced perturbation of aMTOCs coupled to Cre-mediated gene editing generates fragile chromosomes that break when subjected to forces exerted by altered meiosis I spindles., Mouse female meiotic spindles assemble from acentriolar microtubule-organizing centers (aMTOCs) that fragment into discrete foci. These are further sorted and clustered to form spindle poles, thus providing balanced forces for faithful chromosome segregation. To assess the impact of aMTOC biogenesis on spindle assembly, we genetically induced their precocious fragmentation in mouse oocytes using conditional overexpression of Plk4, a master microtubule-organizing center regulator. Excessive microtubule nucleation from these fragmented aMTOCs accelerated spindle assembly dynamics. Prematurely formed spindles promoted the breakage of three different fragilized bivalents, generated by the presence of recombined Lox P sites. Reducing the density of microtubules significantly diminished the extent of chromosome breakage. Thus, improper spindle forces can lead to widely described yet unexplained chromosomal structural anomalies with disruptive consequences on the ability of the gamete to transmit an uncorrupted genome.
- Published
- 2018
27. CHRONOCRISIS: When Cell Cycle Asynchrony Generates DNA Damage in Polyploid Cells
- Author
-
Simon Gemble, Renata Basto, Compartimentation et dynamique cellulaires (CDC), and Centre National de la Recherche Scientifique (CNRS)-Institut Curie [Paris]-Université Pierre et Marie Curie - Paris 6 (UPMC)
- Subjects
DNA damage ,[SDV]Life Sciences [q-bio] ,Mitosis ,Polyploid Cells ,Biology ,Genome ,Chromosomes ,General Biochemistry, Genetics and Molecular Biology ,Cell size ,Polyploidy ,03 medical and health sciences ,0302 clinical medicine ,Humans ,Polyploid cell ,ComputingMilieux_MISCELLANEOUS ,030304 developmental biology ,0303 health sciences ,Cell Cycle ,fungi ,food and beverages ,Cell cycle ,Asynchrony (computer programming) ,Cell biology ,030217 neurology & neurosurgery ,DNA Damage - Abstract
Polyploid cells contain multiple copies of all chromosomes. Polyploidization can be developmentally programmed to sustain tissue barrier function or to increase metabolic potential and cell size. Programmed polyploidy is normally associated with terminal differentiation and poor proliferation capacity. Conversely, non-programmed polyploidy can give rise to cells that retain the ability to proliferate. This can fuel rapid genome rearrangements and lead to diseases like cancer. Here, the mechanisms that generate polyploidy are reviewed and the possible challenges upon polyploid cell division are discussed. The discussion is framed around a recent study showing that asynchronous cell cycle progression (an event that is named "chronocrisis") of different nuclei from a polyploid cell can generate DNA damage at mitotic entry. The potential mechanisms explaining how mitosis in non-programmed polyploid cells can generate abnormal karyotypes and genetic instability are highlighted.
- Published
- 2020
28. Abstract A43: Centrosome amplification favors survival and impairs ovarian cancer progression
- Author
-
Aurélie Herbette, Bassirou Mboup, Anne Vincent-Salomon, Aurélien Latouche, Xavier Sastre-Garau, Oumou Goundiam, André Nicolas, Camille Cosson, Tatiana Popova, Sergio Roman-Roman, Jorge Barbazan, Didier Meseure, Véronique Becette, Didier Decaudin, Guillaume Bataillon, Marc-Henri Stern, Fatima Mechta-Grigoriou, Pierre Gestraud, Fariba Nemati, Renata Basto, Jean-Philippe Morretton, Odette Mariani, Claire Bonneau, and Roman Rouzier
- Subjects
Cancer Research ,Oncology ,Centrosome ,medicine ,Cancer research ,Biology ,Ovarian cancer ,medicine.disease - Abstract
Epithelial ovarian cancer (EOC) is the most lethal gynecologic malignancy. The most common subtype of EOC is high-grade serous (HGSOC), which responds at least initially to chemotherapy but has a worse overall prognosis. Genomic, transcriptomic, and proteogenomic profiling of HGSOC suggested a whole spectrum of molecular diversity, including homologous recombination pathway deficiencies (HRD). The centrosome is the main microtubule (MT)-organizing center of animal cells. It facilitates the accuracy of chromosome segregation during mitosis and influences cell polarity and migration. The presence of more than two centrosomes in a cell, centrosome amplification, has long been associated with tumorigenesis. However, little is known about the real status of centrosome numbers in human cancers and whether numerical alterations are solely associated with poor prognosis. We screened 100 samples of primary EOCs including 88 HGSOC, using immunofluorescence and state-of-the-art microscopy, to determine the centrosome-nucleus index (CNI). We integrated these data with genomic alterations, HRD status, and patient outcome. We found that EOCs are highly heterogeneous, with infrequent but strong centrosome amplifications leading to higher CNI than in healthy tissues. Strikingly, while a correlation between CNI and genomic alterations, such as aneuploidy or chromosome rearrangements, could not be established, we found that high CNI correlates with increased patient survival and sensitivity to chemotherapy, independently of HRD status. Using ovarian cancer cellular models to manipulate centrosome numbers and patient-derived xenografts (PDXs), we found that higher CNIs can positively impact the response to chemotherapy and inhibit peritoneal cell dissemination. Citation Format: Jean-Philippe Morretton, Aurelie Herbette, Camille Cosson, Bassirou Mboup, Aurelien Latouche, Pierre Gestraud, Tatiana Popova, Marc-Henri Stern, Fariba Nemati, Didier Decaudin, Guillaume Bataillon, Veronique Becette, Didier Meseure, Andre Nicolas, Odette Mariani, Claire Bonneau, Jorge Barbazan, Anne Vincent-Salomon, Fatima Mechta-Grigoriou, Sergio Roman-Roman, Roman Rouzier, Xavier Sastre-Garau, Oumou Goundiam, Renata Basto. Centrosome amplification favors survival and impairs ovarian cancer progression [abstract]. In: Proceedings of the AACR Special Conference on Advances in Ovarian Cancer Research; 2019 Sep 13-16, 2019; Atlanta, GA. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(13_Suppl):Abstract nr A43.
- Published
- 2020
29. Centrosome amplification disrupts renal development and causes cystogenesis
- Author
-
Lai Kuan Dionne, Sanjay Jain, Amanda Knoten, Kyuhwan Shim, Renata Basto, Véronique Marthiens, Moe R. Mahjoub, Jinzhi Wang, Tao Cheng, Masato Hoshi, Washington University School of Medicine in St. Louis, Washington University in Saint Louis (WUSTL), Department of Mathematics (St Louis), Institut Curie [Paris], University College of London [London] (UCL), Compartimentation et dynamique cellulaires (CDC), Centre National de la Recherche Scientifique (CNRS)-Institut Curie [Paris]-Université Pierre et Marie Curie - Paris 6 (UPMC), Department of Computer Science - Singapore, and National University of Singapore (NUS)
- Subjects
0301 basic medicine ,[SDV]Life Sciences [q-bio] ,Mitosis ,Cellular homeostasis ,Kidney development ,Spindle Apparatus ,Biology ,Kidney ,Article ,Mice ,03 medical and health sciences ,Cystic kidney disease ,0302 clinical medicine ,Ciliogenesis ,Morphogenesis ,medicine ,Animals ,Homeostasis ,Humans ,Research Articles ,ComputingMilieux_MISCELLANEOUS ,Cell Proliferation ,Centrosome ,Cystic kidney ,Cell Differentiation ,Epithelial Cells ,Cell Biology ,medicine.disease ,Renal hypoplasia ,3. Good health ,Spindle apparatus ,Cell biology ,030104 developmental biology ,030220 oncology & carcinogenesis - Abstract
Supernumerary centrosomes are commonly observed in cystic kidneys, but whether they are a cause or consequence of cystogenesis is unknown. Dionne et al. demonstrate that centrosome amplification disrupts renal development and is sufficient to induce cystogenesis in vivo., Centrosome number is tightly controlled to ensure proper ciliogenesis, mitotic spindle assembly, and cellular homeostasis. Centrosome amplification (the formation of excess centrosomes) has been noted in renal cells of patients and animal models of various types of cystic kidney disease. Whether this defect plays a causal role in cystogenesis remains unknown. Here, we investigate the consequences of centrosome amplification during kidney development, homeostasis, and after injury. Increasing centrosome number in vivo perturbed proliferation and differentiation of renal progenitors, resulting in defective branching morphogenesis and renal hypoplasia. Centrosome amplification disrupted mitotic spindle morphology, ciliary assembly, and signaling pathways essential for the function of renal progenitors, highlighting the mechanisms underlying the developmental defects. Importantly, centrosome amplification was sufficient to induce rapid cystogenesis shortly after birth. Finally, we discovered that centrosome amplification sensitized kidneys in adult mice, causing cystogenesis after ischemic renal injury. Our study defines a new mechanism underlying the pathogenesis of renal cystogenesis, and identifies a potentially new cellular target for therapy.
- Published
- 2018
30. Gene editing can generate fragile bivalents in mouse oocytes
- Author
-
Keith T. Jones, Marie-Emilie Terret, Véronique Marthiens, Marie-Hélène Verlhac, Marion Manil-Ségalen, Małgorzata Łuksza, Renata Basto, J Kannaan, and Simon I. R. Lane
- Subjects
PLK4 ,0303 health sciences ,Microtubule organizing center ,Biology ,Spindle pole body ,Cell biology ,Chromosome segregation ,03 medical and health sciences ,0302 clinical medicine ,Meiosis ,Microtubule ,Chromosome breakage ,030217 neurology & neurosurgery ,030304 developmental biology ,Microtubule nucleation - Abstract
Mouse female meiotic spindles assemble from acentriolar MTOCs (aMTOCs) that fragment into discrete foci. These are further sorted and clustered to form spindle poles, thus providing balanced forces for faithful chromosome segregation. To assess the impact of aMTOCs biogenesis on spindle assembly, we genetically induced their precocious fragmentation in mouse oocytes using conditional overexpression of Plk4, a master MTOC regulator. Excessive microtubule nucleation from these fragmented aMTOCs accelerated spindle assembly dynamics. Prematurely formed spindles promoted the breakage of three different fragilized bivalents, generated by the presence of recombined Lox P sites. Reducing the density of microtubules diminished the extent of chromosome breakage. Thus, improper spindle forces can lead to widely described yet unexplained chromosomal structural anomalies with disruptive consequences on the ability of the gamete to transmit an uncorrupted genome.
- Published
- 2018
- Full Text
- View/download PDF
31. When E-cadherin is away, centrosomes can play
- Author
-
Renata Basto, Diana Vargas-Hurtado, Centre de recherche de l'Institut Curie [Paris], Institut Curie [Paris], Biologie Cellulaire et Cancer, Institut Curie [Paris]-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS), Compartimentation et dynamique cellulaires (CDC), and Centre National de la Recherche Scientifique (CNRS)-Institut Curie [Paris]-Université Pierre et Marie Curie - Paris 6 (UPMC)
- Subjects
rho GTP-Binding Proteins ,0301 basic medicine ,[SDV]Life Sciences [q-bio] ,Cell ,Spindle Apparatus ,Biology ,Contractility ,03 medical and health sciences ,Discoidin Domain Receptor 1 ,medicine ,Guanine Nucleotide Exchange Factors ,Humans ,Spotlight ,Cluster analysis ,ComputingMilieux_MISCELLANEOUS ,Centrosome ,DDR1 ,Cadherin ,Cell Biology ,Cadherins ,Cell biology ,030104 developmental biology ,medicine.anatomical_structure ,Cancer cell ,Commentary ,rhoA GTP-Binding Protein - Abstract
Vargas-Hurtado and Basto highlight recent work from Rhys et al. revealing how E-cadherin affects clustering of extra centrosomes., Centrosome clustering is a process frequently used by cancer cells with extra centrosomes to avoid multipolar divisions. How cell-intrinsic properties influence clustering is not entirely known. In this issue, Rhys et al. (2017. J. Cell Biol. https://doi.org/10.1083/jcb.201704102) report an unexpected link between clustering capacity and cortical contractility through E-cadherin and DDR1 proteins.
- Published
- 2017
32. Consequences of Centrosome Dysfunction During Brain Development
- Author
-
Maddalena, Nano and Renata, Basto
- Subjects
Centrosome ,Disease Models, Animal ,Species Specificity ,Microcephaly ,Morphogenesis ,Animals ,Brain ,Gene Expression Regulation, Developmental ,Humans ,Mitosis ,Biological Evolution - Abstract
Development requires cell proliferation, differentiation and spatial organization of daughter cells to occur in a highly controlled manner. The mode of cell division, the extent of proliferation and the spatial distribution of mitosis allow the formation of tissues of the right size and with the correct structural organization. All these aspects depend on cell cycle duration, correct chromosome segregation and spindle orientation. The centrosome, which is the main microtubule-organizing centre (MTOC) of animal cells, contributes to all these processes. As one of the most structurally complex organs in our body, the brain is particularly susceptible to centrosome dysfunction. Autosomal recessive primary microcephaly (MCPH), primordial dwarfism disease Seckel syndrome (SCKS) and microcephalic osteodysplastic primordial dwarfism type II (MOPD-II) are often connected to mutations in centrosomal genes. In this chapter, we discuss the consequences of centrosome dysfunction during development and how they can contribute to the etiology of human diseases.
- Published
- 2017
33. Centrosome amplification and cancer: A question of sufficiency
- Author
-
Renata Basto, Jordan W. Raff, Centre de recherche de l'Institut Curie [Paris], Institut Curie [Paris], and University of Oxford, Oxford, United Kingdom
- Subjects
0301 basic medicine ,[SDV]Life Sciences [q-bio] ,Computational biology ,Biology ,medicine.disease_cause ,General Biochemistry, Genetics and Molecular Biology ,Article ,03 medical and health sciences ,Neoplasms ,medicine ,Animals ,Molecular Biology ,book ,ComputingMilieux_MISCELLANEOUS ,Genetics ,Centrosome ,Developmental cell ,Cancer ,Cell Biology ,medicine.disease ,030104 developmental biology ,Cell Transformation, Neoplastic ,Feature (computer vision) ,book.journal ,Carcinogenesis ,Developmental Biology - Abstract
Centrosome amplification is a common feature of human tumors, but whether this is a cause or a consequence of cancer remains unclear. Here, we test the consequence of centrosome amplification by creating mice in which centrosome number can be chronically increased in the absence of additional genetic defects. We show that increasing centrosome number elevated tumor initiation in a mouse model of intestinal neoplasia. Most importantly, we demonstrate that supernumerary centrosomes are sufficient to drive aneuploidy and the development of spontaneous tumors in multiple tissues. Tumors arising from centrosome amplification exhibit frequent mitotic errors and possess complex karyotypes, recapitulating a common feature of human cancer. Together, our data support a direct causal relationship between centrosome amplification, genomic instability and tumor development.
- Published
- 2017
34. Consequences of Centrosome Dysfunction During Brain Development
- Author
-
Maddalena Nano, Renata Basto, Centre de recherche de l'Institut Curie [Paris], and Institut Curie [Paris]
- Subjects
0301 basic medicine ,Microcephaly ,Cell division ,[SDV]Life Sciences [q-bio] ,Microtubule organizing center ,Cell cycle ,Biology ,medicine.disease ,Cell biology ,03 medical and health sciences ,030104 developmental biology ,Seckel syndrome ,Centrosome ,medicine ,Primordial dwarfism ,Mitosis ,ComputingMilieux_MISCELLANEOUS - Abstract
Development requires cell proliferation, differentiation and spatial organization of daughter cells to occur in a highly controlled manner. The mode of cell division, the extent of proliferation and the spatial distribution of mitosis allow the formation of tissues of the right size and with the correct structural organization. All these aspects depend on cell cycle duration, correct chromosome segregation and spindle orientation. The centrosome, which is the main microtubule-organizing centre (MTOC) of animal cells, contributes to all these processes. As one of the most structurally complex organs in our body, the brain is particularly susceptible to centrosome dysfunction. Autosomal recessive primary microcephaly (MCPH), primordial dwarfism disease Seckel syndrome (SCKS) and microcephalic osteodysplastic primordial dwarfism type II (MOPD-II) are often connected to mutations in centrosomal genes. In this chapter, we discuss the consequences of centrosome dysfunction during development and how they can contribute to the etiology of human diseases.
- Published
- 2017
35. Centrosome and Centriole
- Author
-
Renata Basto, Karen Oegema, Renata Basto, and Karen Oegema
- Subjects
- Cell division, Centrosomes, Cytology
- Abstract
This new volume of Methods in Cell Biology looks at methods for analyzing centrosomes and centrioles. Chapters cover such topics as methods to analyze centrosomes, centriole biogenesis and function in multi-ciliated cells, laser manipulation of centrosomes or CLEM, analysis of centrosomes in human cancers and tissues, proximity interaction techniques to study centrosomes, and genome engineering for creating conditional alleles in human cells. Covers sections on model systems and functional studies, imaging-based approaches and emerging studies Chapters are written by experts in the field Cutting-edge material
- Published
- 2015
36. Methods in Cilia and Flagella
- Author
-
Renata Basto, Wallace F. Marshall, Renata Basto, and Wallace F. Marshall
- Subjects
- Cilia and ciliary motion--Laboratory manuals, Flagella (Microbiology)--Laboratory manuals
- Abstract
The goal of this book is to collect methods and protocols for studying cilia in a wide range of different cell types, so that researchers from many fields of biology can start exploring the role of cilia in their own system. Chapters are written by experts in the field Cutting-edge material
- Published
- 2015
37. Overexpressing centriole-replication proteins in vivo induces centriole overduplication and de novo formation
- Author
-
Renata Basto, Jordan W. Raff, Naomi R. Stevens, Nina Peel, University of Cambridge [UK] (CAM), Centre de recherche de l'Institut Curie [Paris], and Institut Curie [Paris]
- Subjects
Male ,PLK4 ,Embryo, Nonmammalian ,Centriole ,[SDV]Life Sciences [q-bio] ,Recombinant Fusion Proteins ,Green Fluorescent Proteins ,CELLCYCLE ,Protein Serine-Threonine Kinases ,Biology ,Article ,General Biochemistry, Genetics and Molecular Biology ,Centriole elongation ,03 medical and health sciences ,Oogenesis ,0302 clinical medicine ,Spermatocytes ,Animals ,Drosophila Proteins ,Basal body ,ComputingMilieux_MISCELLANEOUS ,Centrioles ,Ovum ,030304 developmental biology ,Genetics ,0303 health sciences ,Agricultural and Biological Sciences(all) ,Biochemistry, Genetics and Molecular Biology(all) ,Brain ,Cell biology ,Centrosome ,[SDU]Sciences of the Universe [physics] ,Larva ,CELLBIO ,Drosophila ,CEP135 ,Centriole replication ,General Agricultural and Biological Sciences ,Microtubule-Associated Proteins ,030217 neurology & neurosurgery ,Centriole assembly - Abstract
International audience; Background: Centrosomes have important roles in many aspects of cell organization, and aberrations in their number and function are associated with various diseases, including cancer. Centrosomes consist of a pair of centrioles surrounded by a pericentriolar matrix (PCM), and their replication is tightly regulated. Here, we investigate the effects of overexpressing the three proteins known to be required for centriole replication in Drosophila-DSas-6, DSas-4, and Sak. Results: By directly observing centriole replication in living Drosophila embryos, we show that the overexpression of GFP-DSas-6 can drive extra rounds of centriole replication within a single cell cycle. Extra centriole-like structures also accumulate in brain cells that overexpress either GFP-DSas-6 or GFP-Sak, but not DSas-4-GFP. No extra centrioles accumulate in spermatocytes that overexpress any of these three proteins. Most remarkably, the overexpression of any one of these three proteins results in the rapid de novo formation of many hundreds of centriole-like structures in unfertilized eggs, which normally do not contain centrioles. Conclusions: Our data suggest that the levels of centriolar DSas-6 determine the number of daughter centrioles formed during centriole replication. Overexpression of either DSas-6 or Sak can induce the formation of extra centrioles in some tissues but not others, suggesting that centriole replication is regulated differently in different tissues. The finding that the overexpression of DSas-4, DSas-6, or Sak can rapidly induce the de novo formation of centriole-like structures in Drosophila eggs suggests that this process results from the stabilization of centriole-precursors that are normally present in the egg.
- Published
- 2016
38. The Janus soul of centrosomes: a paradoxical role in disease?
- Author
-
Maddalena Nano, Renata Basto, Centre de recherche de l'Institut Curie [Paris], and Institut Curie [Paris]
- Subjects
0301 basic medicine ,Centrosome ,Cell division ,[SDV]Life Sciences [q-bio] ,Microtubule organizing center ,Centrosome cycle ,Cell fate determination ,Biology ,Aneuploidy ,Cell biology ,03 medical and health sciences ,030104 developmental biology ,Chromosome instability ,Ciliogenesis ,Chromosomal Instability ,Neoplasms ,Genetics ,Animals ,Chromosomes, Human ,Humans ,Mitosis ,ComputingMilieux_MISCELLANEOUS - Abstract
The centrosome is the main microtubule organizing center of animal cells. It contributes to spindle assembly and orientation during mitosis and to ciliogenesis in interphase. Numerical and structural defects in this organelle are known to be associated with developmental disorders such as dwarfism and microcephaly, but only recently, the molecular mechanisms linking centrosome aberrations to altered physiology are being elucidated. Defects in centrosome number or structure have also been described in cancer. These opposite clinical outcomes--arising from reduced proliferation and overproliferation respectively--can be explained in light of the tissue- and developmental-specific requirements for centrosome functions. The pathological outcomes of centrosome deficiencies have become clearer when considering its consequences. Among them, there are genetic instability (mainly aneuploidy, a defect in chromosome number), defects in the symmetry of cell division (important for cell fate specification and tissue architecture) and impaired ciliogenesis. In this review, we discuss the origins and the consequences of centrosome flaws, with particular attention on how they contribute to developmental diseases.
- Published
- 2016
39. Consequences of Numerical Centrosome Defects in Development and Disease
- Author
-
Davide Gambarotto and Renata Basto
- Subjects
0301 basic medicine ,Genome instability ,Genetics ,Disease ,Biology ,medicine.disease ,Spindle pole body ,03 medical and health sciences ,030104 developmental biology ,0302 clinical medicine ,Centrosome ,Autosomal Recessive Primary Microcephaly ,medicine ,Primordial dwarfism ,Gene ,030217 neurology & neurosurgery ,Organism - Abstract
Defects in centrosome number or structure can have considerable consequences for the physiology of an organism. Aberrant centrosome number has been proposed for a century to contribute to genome instability and tumour formation. However, in the last decade, mutations in centrosome genes have been described in diseases characterised by defective growth. Centrosome dysfunction can therefore have opposite effects on the homeostasis of the organism. Here we discuss how deregulation of centrosome number during embryonic development might contribute to growth defective syndromes such as autosomal recessive primary microcephaly (MCPH) and primordial dwarfism. We further discuss how the same defects might play a role in cancer when present in adult tissues.
- Published
- 2016
40. Aneuploidy causes premature differentiation of neural and intestinal stem cells
- Author
-
Delphine Gogendeau, Carole Pennetier, Davide Gambarotto, Renata Basto, Allison J. Bardin, Katarzyna Siudeja, Centre de recherche de l'Institut Curie [Paris], and Institut Curie [Paris]
- Subjects
Microcephaly ,Cellular differentiation ,[SDV]Life Sciences [q-bio] ,General Physics and Astronomy ,Aneuploidy ,Biology ,General Biochemistry, Genetics and Molecular Biology ,Article ,Neural Stem Cells ,medicine ,Animals ,Wings, Animal ,ComputingMilieux_MISCELLANEOUS ,Cell Proliferation ,Genetics ,Multidisciplinary ,Cell growth ,Stem Cells ,G1 Phase ,Brain ,Cell Differentiation ,General Chemistry ,Organ Size ,Cell cycle ,medicine.disease ,Immunohistochemistry ,Neural stem cell ,Cell biology ,Intestines ,Apoptosis ,Drosophila ,Stem cell - Abstract
Aneuploidy is associated with a variety of diseases such as cancer and microcephaly. Although many studies have addressed the consequences of a non-euploid genome in cells, little is known about their overall consequences in tissue and organism development. Here we use two different mutant conditions to address the consequences of aneuploidy during tissue development and homeostasis in Drosophila. We show that aneuploidy causes brain size reduction due to a decrease in the number of proliferative neural stem cells (NSCs), but not through apoptosis. Instead, aneuploid NSCs present an extended G1 phase, which leads to cell cycle exit and premature differentiation. Moreover, we show that this response to aneuploidy is also present in adult intestinal stem cells but not in the wing disc. Our work highlights a neural and intestine stem cell-specific response to aneuploidy, which prevents their proliferation and expansion., It is unclear why certain tissues are more susceptible to the consequences of aneuploidy. Here, in Drosophila, Gogendeau et al. identify aneuploidy as the cause of lengthened G1 and premature differentiation in both neural and adult intestinal stem cells, which prevents cells with abnormal genomes from cycling.
- Published
- 2015
41. Sas-4 proteins are required during basal body duplication inParamecium
- Author
-
Graça Raposo, Delphine Gogendeau, Renata Basto, Jean Cohen, Ilse Hurbain, Centre de recherche de l'Institut Curie [Paris], Institut Curie [Paris], and Curie Institute
- Subjects
Paramecium ,Centriole ,Recombinant Fusion Proteins ,[SDV]Life Sciences [q-bio] ,sports ,Protozoan Proteins ,03 medical and health sciences ,Procentriole ,0302 clinical medicine ,Tubulin ,Microtubule ,Gene duplication ,Animals ,Humans ,Basal body ,Cilia ,Gene Silencing ,Molecular Biology ,Cytoskeleton ,Centrioles ,030304 developmental biology ,Genetics ,0303 health sciences ,biology ,Cilium ,Articles ,Cell Biology ,biology.organism_classification ,Cell biology ,sports.league ,Paramecium tetraurelia ,030217 neurology & neurosurgery - Abstract
This study investigated the role of Sas-4 in basal body duplication in Paramecium and found that Sas-4 proteins are required to assemble and stabilize the germinative disk and cartwheel, which suggests that Sas-4 plays an essential role in basal body duplication., Centrioles and basal bodies are structurally related organelles composed of nine microtubule (MT) triplets. Studies performed in Caenorhabditis elegans embryos have shown that centriole duplication takes place in sequential way, in which different proteins are recruited in a specific order to assemble a procentriole. ZYG-1 initiates centriole duplication by triggering the recruitment of a complex of SAS-5 and SAS-6, which then recruits the final player, SAS-4, to allow the incorporation of MT singlets. It is thought that a similar mechanism (that also involves additional proteins) is present in other animal cells, but it remains to be investigated whether the same players and their ascribed functions are conserved during basal body duplication in cells that exclusively contain basal bodies. To investigate this question, we have used the multiciliated protist Paramecium tetraurelia. Here we show that in the absence of PtSas4, two types of defects in basal body duplication can be identified. In the majority of cases, the germinative disk and cartwheel, the first structures assembled during duplication, are not detected. In addition, if daughter basal bodies were formed, they invariably had defects in MT recruitment. Our results suggest that PtSas4 has a broader function than its animal orthologues.
- Published
- 2011
42. Centrosome Amplification Can Initiate Tumorigenesis in Flies
- Author
-
Jordan W. Raff, Renata Basto, Nina Peel, Anna Franz, Alexey Khodjakov, Tatiana Vinadogrova, Kathrin Brunk, Centre de recherche de l'Institut Curie [Paris], Institut Curie [Paris], and University of Cambridge [UK] (CAM)
- Subjects
PLK4 ,[SDV]Life Sciences [q-bio] ,Green Fluorescent Proteins ,Kinesins ,Mitosis ,Centrosome cycle ,CELLCYCLE ,Spindle Apparatus ,Protein Serine-Threonine Kinases ,Biology ,General Biochemistry, Genetics and Molecular Biology ,Animals, Genetically Modified ,03 medical and health sciences ,0302 clinical medicine ,Animals ,Drosophila Proteins ,ComputingMilieux_MISCELLANEOUS ,030304 developmental biology ,Centrosome ,0303 health sciences ,Biochemistry, Genetics and Molecular Biology(all) ,fungi ,Spindle apparatus ,Cell biology ,Spindle checkpoint ,Drosophila melanogaster ,Larva ,030220 oncology & carcinogenesis ,CELLBIO ,Multipolar spindles ,Centriole assembly - Abstract
International audience; Centrosome amplification is a common feature of many cancer cells, and it has been previously proposed that centrosome amplification can drive genetic instability and so tumorigenesis. To test this hypothesis, we generated Drosophila lines that have extra centrosomes in 60% of their somatic cells. Many cells with extra centrosomes initially form multipolar spindles, but these spindles ultimately become bipolar. This requires a delay in mitosis that is mediated by the spindle assembly checkpoint (SAC). As a result of this delay, there is no dramatic increase in genetic instability in flies with extra centrosomes, and these flies maintain a stable diploid genome over many generations. The asymmetric division of the larval neural stem cells, however, is compromised in the presence of extra centrosomes, and larval brain cells with extra centrosomes can generate metastatic tumors when transplanted into the abdomens of wild-type hosts. Thus, centrosome amplification can initiate tumorigenesis in flies.
- Published
- 2008
43. From Stem Cell to Embryo without Centrioles
- Author
-
Renata Basto, Alexandre A.S.F. Raposo, Naomi R. Stevens, Daniel St Johnston, Jordan W. Raff, and University of Cambridge [UK] (CAM)
- Subjects
Centriole ,[SDV]Life Sciences [q-bio] ,Embryonic Development ,Centrosome cycle ,Biology ,Microtubules ,General Biochemistry, Genetics and Molecular Biology ,03 medical and health sciences ,0302 clinical medicine ,Oogenesis ,Microtubule ,Report ,medicine ,Animals ,Drosophila Proteins ,RNA, Messenger ,Mitosis ,ComputingMilieux_MISCELLANEOUS ,030304 developmental biology ,Centrioles ,0303 health sciences ,Agricultural and Biological Sciences(all) ,Biochemistry, Genetics and Molecular Biology(all) ,fungi ,DNA ,Oocyte ,Cell biology ,medicine.anatomical_structure ,Centrosome ,Oocytes ,CELLBIO ,Drosophila ,Female ,Centriole replication ,Stem cell ,General Agricultural and Biological Sciences ,Microtubule-Associated Proteins ,Totipotent Stem Cells ,030217 neurology & neurosurgery - Abstract
International audience; Centrosome asymmetry plays a key role in ensuring the asymmetric division of Drosophila neural stem cells (neuroblasts [NBs]) and male germline stem cells (GSCs) [1-3]. In both cases, one centrosome is anchored close to a specific cortical region during interphase, thus defining the orientation of the spindle during the ensuing mitosis. To test whether asymmetric centrosome behavior is a general feature of stem cells, we have studied female GSCs, which divide asymmetrically, producing another GSC and a cystoblast. The cystoblast then divides and matures into an oocyte, a process in which centrosomes exhibit a series of complex behaviors proposed to play a crucial role in oogenesis [4-6]. We show that the interphase centrosome does not define spindle orientation in female GSCs and that DSas-4 mutant GSCs [7], lacking centrioles and centrosomes, invariably divide asymmetrically to produce cystoblasts that proceed normally through oogenesis-remarkably, oocyte specification, microtubule organization, and mRNA localization are all unperturbed. Mature oocytes can be fertilized, but embryos that cannot support centriole replication arrest very early in development. Thus, centrosomes are dispensable for oogenesis but essential for early embryogenesis. These results reveal that asymmetric centrosome behavior is not an essential feature of stem cell divisions. Results and Discussion
- Published
- 2007
- Full Text
- View/download PDF
44. New insights into centrosome imaging in Drosophila and mouse neuroepithelial tissues
- Author
-
Maria A, Rujano, Renata, Basto, and Véronique, Marthiens
- Subjects
Central Nervous System ,Centrosome ,Male ,Tissue Culture Techniques ,Mice ,Neuroepithelial Cells ,Animals ,Drosophila ,Female - Abstract
The centrosome is the main microtubule-organizing center in animal cells. It participates in the assembly of a bipolar spindle that ensures accurate segregation of chromosomes during mitosis. Recently, mutations in centrosome genes have been identified in patients affected by neurodevelopmental disorders. In fact, the etiology of several neurodevelopmental pathologies seems to be linked to defects in the assembly of the mitotic spindle in the neural stem cell compartment during neurogenesis. Therefore, getting better insights into the structure and function/dysfunction of the mitotic spindle apparatus in an intact tissue environment is of utmost importance. However, imaging nanometer-scale structures like centrosomes and microtubule bundles within the depth of a tissue is still challenging. Here we describe two procedures to acquire high-resolution images on fixed tissues and to perform live imaging of microtubule-based structures in the neuroepithelia of the Drosophila brain and of the mouse neocortex. We take advantage of the accumulation of centrosomes and mitotic figures at the apical surface of these polarized tissues to improve the quality of staining and imaging. Both Drosophila and mouse models with centrosome dysfunction showed abnormalities in the neuroepithelium reminiscent of the ones described in brains of human patients. These observations have highlighted their value as model organisms to study the etiology of human neurodevelopmental pathologies.
- Published
- 2015
45. Transient PLK4 overexpression accelerates tumorigenesis in p53-deficient epidermis
- Author
-
Andrea E. Karambelas, Véronique Marthiens, Virginie Moers, Özdemirhan Serçin, Marie Le Mercier, Bram Boeckx, Jean-Christophe Larsimont, Renata Basto, Diether Lambrechts, Cédric Blanpain, Département de mathématiques Université Libre de Bruxelles, and Université libre de Bruxelles (ULB)
- Subjects
0301 basic medicine ,PLK4 ,Skin Neoplasms ,Transgene ,[SDV]Life Sciences [q-bio] ,Aneuploidy ,Apoptosis ,Biology ,Protein Serine-Threonine Kinases ,medicine.disease_cause ,03 medical and health sciences ,Mice ,medicine ,Animals ,Progenitor cell ,ComputingMilieux_MISCELLANEOUS ,Centrosome ,integumentary system ,Epidermis (botany) ,Cell Biology ,medicine.disease ,Cell biology ,030104 developmental biology ,Cell Transformation, Neoplastic ,Cancer research ,Epidermis ,Tumor Suppressor Protein p53 ,Carcinogenesis - Abstract
Aneuploidy is found in most solid tumours, but it remains unclear whether it is the cause or the consequence of tumorigenesis. Using Plk4 overexpression (PLK4OE) during epidermal development, we assess the impact of centrosome amplification and aneuploidy on skin development and tumorigenesis. PLK4OE in the developing epidermis induced centrosome amplification and multipolar divisions, leading to p53 stabilization and apoptosis of epidermal progenitors. The resulting delayed epidermal stratification led to skin barrier defects. Plk4 transgene expression was shut down postnatally in the surviving mice and PLK4OE mice never developed skin tumours. Concomitant PLK4OE and p53 deletion (PLK4OE/p53cKO) rescued the differentiation defects, but did not prevent the apoptosis of PLK4OE cells. Remarkably, the short-term presence of cells with supernumerary centrosomes in PLK4OE/p53cKO mice was sufficient to generate aneuploidy in the adult epidermis and triggered spontaneous skin cancers with complete penetrance. These results reveal that aneuploidy induced by transient centrosome amplification can accelerate tumorigenesis in p53-deficient cells.
- Published
- 2015
46. Moesin Is a Major Regulator of Centrosome Behavior in Epithelial Cells with Extra Centrosomes
- Author
-
Renata Basto, Davide Gambarotto, Delphine Gogendeau, Florent Dingli, Dora Sabino, Carole Pennetier, Guillaume Arras, Damarys Loew, Maddalena Nano, Centre de recherche de l'Institut Curie [Paris], and Institut Curie [Paris]
- Subjects
Programmed cell death ,Moesin ,[SDV]Life Sciences [q-bio] ,Centrosome cycle ,Spindle Apparatus ,Biology ,General Biochemistry, Genetics and Molecular Biology ,Article ,03 medical and health sciences ,0302 clinical medicine ,medicine ,Animals ,Mitosis ,ComputingMilieux_MISCELLANEOUS ,030304 developmental biology ,Centrosome ,0303 health sciences ,FERM domain ,Cell Death ,Agricultural and Biological Sciences(all) ,Biochemistry, Genetics and Molecular Biology(all) ,Microfilament Proteins ,Epithelial Cells ,Aneuploidy ,Epithelium ,Cell biology ,Spindle apparatus ,Up-Regulation ,medicine.anatomical_structure ,030220 oncology & carcinogenesis ,Drosophila ,General Agricultural and Biological Sciences - Abstract
Summary Centrosome amplification has severe consequences during development and is thought to contribute to a variety of diseases such as cancer and microcephaly. However, the adverse effects of centrosome amplification in epithelia are still not known. Here, we investigate the consequences of centrosome amplification in the Drosophila wing disc epithelium. We found that epithelial cells exhibit mechanisms of clustering but also inactivation of extra centrosomes. Importantly, these mechanisms are not fully efficient, and both aneuploidy and cell death can be detected. Epithelial cells with extra centrosomes generate tumors when transplanted into WT hosts and inhibition of cell death results in tissue over-growth and disorganization. Using SILAC-fly, we found that Moesin, a FERM domain protein, is specifically upregulated in wing discs with extra centrosomes. Moesin localizes to the centrosomes and mitotic spindle during mitosis, and we show that Moesin upregulation influences extra-centrosome behavior and robust bipolar spindle formation. This study provides a mechanistic explanation for the increased aneuploidy and transformation potential primed by centrosome amplification in epithelial tissues., Graphical Abstract, Highlights • Consequences of centrosome amplification in epithelia are discussed • Centrosome clustering or inactivation is not fully efficient • High levels of Moesin contribute to defects in bipolar spindle assembly • Centrosome amplification generates aneuploidy and epithelial transformation, Sabino et al. study the consequences of centrosome amplification in an epithelium. They find that mechanisms of extra-centrosome clustering and inactivation are not fully efficient. High levels of Moesin, an ERM protein, contribute to a failure in bipolar spindle assembly leading to aneuploidy and priming tumor-initiating events.
- Published
- 2015
47. Quantitative analysis of flagellar proteins in Drosophila sperm tails
- Author
-
Teresa, Mendes Maia, Perrine, Paul-Gilloteaux, Renata, Basto, Compartimentation et dynamique cellulaires (CDC), Centre National de la Recherche Scientifique (CNRS)-Institut Curie [Paris]-Université Pierre et Marie Curie - Paris 6 (UPMC), and Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut Curie [Paris]-Centre National de la Recherche Scientifique (CNRS)
- Subjects
Male ,Proteome ,Optical Imaging ,fluorescence quantification ,Cell Differentiation ,[SDV.BC]Life Sciences [q-bio]/Cellular Biology ,motile sperm ,tubulin post-translation modifications ,Sperm individualization ,Drosophila melanogaster ,Tubulin ,Flagella ,Sperm Tail ,[INFO.INFO-TI]Computer Science [cs]/Image Processing [eess.IV] ,Image Processing, Computer-Assisted ,Animals ,Drosophila Proteins ,Drosophila ,Protein Processing, Post-Translational - Abstract
International audience; The cilium has a well-defined structure, which can still accommodate some morphological and molecular composition diversity to suit the functional requirements of different cell types. The sperm flagellum of the fruit fly Drosophila melanogaster appears as a good model to study the genetic regulation of axoneme assembly and motility, due to the wealth of genetic tools publically available for this organism. In addition, the fruit fly's sperm flagellum displays quite a long axoneme (∼1.8mm), which may facilitate both histological and biochemical analyses. Here, we present a protocol for imaging and quantitatively analyze proteins, which associate with the fly differentiating, and mature sperm flagella. We will use as an example the quantification of tubulin polyglycylation in wild-type testes and in Bug22 mutant testes, which present defects in the deposition of this posttranslational modification. During sperm biogenesis, flagella appear tightly bundled, which makes it more challenging to get accurate measurements of protein levels from immunostained specimens. The method we present is based on the use of a novel semiautomated, macro installed in the image processing software ImageJ. It allows to measure fluorescence levels in closely associated sperm tails, through an exact distinction between positive and background signals, and provides background-corrected pixel intensity values that can directly be used for data analysis.
- Published
- 2015
48. Preface
- Author
-
Karen Oegema, Renata Basto, Centre de recherche de l'Institut Curie [Paris], and Institut Curie [Paris]
- Subjects
0303 health sciences ,03 medical and health sciences ,0302 clinical medicine ,Centriole ,Cytological Techniques ,Centrosome ,030220 oncology & carcinogenesis ,[SDV]Life Sciences [q-bio] ,Biology ,ComputingMilieux_MISCELLANEOUS ,030304 developmental biology ,Cell biology - Abstract
International audience
- Published
- 2015
49. Quantitative analysis of flagellar proteins in Drosophila sperm tails
- Author
-
Teresa Mendes Maia, Perrine Paul-Gilloteaux, Renata Basto, Centre de recherche de l'Institut Curie [Paris], and Institut Curie [Paris]
- Subjects
Axoneme ,0303 health sciences ,Sperm flagellum ,biology ,Sperm individualization ,[SDV]Life Sciences [q-bio] ,Flagellum ,biology.organism_classification ,Sperm ,Axoneme assembly ,Cell biology ,03 medical and health sciences ,0302 clinical medicine ,Polyglycylation ,Drosophila melanogaster ,030217 neurology & neurosurgery ,ComputingMilieux_MISCELLANEOUS ,030304 developmental biology - Abstract
The cilium has a well-defined structure, which can still accommodate some morphological and molecular composition diversity to suit the functional requirements of different cell types. The sperm flagellum of the fruit fly Drosophila melanogaster appears as a good model to study the genetic regulation of axoneme assembly and motility, due to the wealth of genetic tools publically available for this organism. In addition, the fruit fly's sperm flagellum displays quite a long axoneme (∼1.8mm), which may facilitate both histological and biochemical analyses. Here, we present a protocol for imaging and quantitatively analyze proteins, which associate with the fly differentiating, and mature sperm flagella. We will use as an example the quantification of tubulin polyglycylation in wild-type testes and in Bug22 mutant testes, which present defects in the deposition of this posttranslational modification. During sperm biogenesis, flagella appear tightly bundled, which makes it more challenging to get accurate measurements of protein levels from immunostained specimens. The method we present is based on the use of a novel semiautomated, macro installed in the image processing software ImageJ. It allows to measure fluorescence levels in closely associated sperm tails, through an exact distinction between positive and background signals, and provides background-corrected pixel intensity values that can directly be used for data analysis.
- Published
- 2015
50. Preface
- Author
-
Wallace F. Marshall, Renata Basto, Centre de recherche de l'Institut Curie [Paris], and Institut Curie [Paris]
- Subjects
Cilium ,[SDV]Life Sciences [q-bio] ,Biology ,Flagellum ,Neuroscience ,ComputingMilieux_MISCELLANEOUS ,Introductory Journal Article - Abstract
International audience
- Published
- 2015
Catalog
Discovery Service for Jio Institute Digital Library
For full access to our library's resources, please sign in.