35 results on '"Sergé, Arnauld"'
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2. Recirculating Foxp3+ regulatory T cells are restimulated in the thymus under Aire control
- Author
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Charaix, Jonathan, Borelli, Alexia, Santamaria, Jérémy C., Chasson, Lionel, Giraud, Matthieu, Sergé, Arnauld, and Irla, Magali
- Published
- 2022
- Full Text
- View/download PDF
3. Intrathymic adeno-associated virus gene transfer rapidly restores thymic function and long-term persistence of gene-corrected T cells
- Author
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Pouzolles, Marie, Machado, Alice, Guilbaud, Mickaël, Irla, Magali, Gailhac, Sarah, Barennes, Pierre, Cesana, Daniela, Calabria, Andrea, Benedicenti, Fabrizio, Sergé, Arnauld, Raman, Indu, Li, Quan-Zhen, Montini, Eugenio, Klatzmann, David, Adjali, Oumeya, Taylor, Naomi, and Zimmermann, Valérie S.
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- 2020
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4. Transforming Growth Factor-beta signaling in αβ thymocytes promotes negative selection
- Author
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McCarron, Mark J., Irla, Magali, Sergé, Arnauld, Soudja, Saidi M’Homa, and Marie, Julien C.
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- 2019
- Full Text
- View/download PDF
5. Recirculating Foxp3+ regulatory T cells are restimulated in the thymus under Aire control.
- Author
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Charaix, Jonathan, Borelli, Alexia, Santamaria, Jérémy C., Chasson, Lionel, Giraud, Matthieu, Sergé, Arnauld, and Irla, Magali
- Abstract
Thymically-derived Foxp3
+ regulatory T cells (Treg ) critically control immunological tolerance. These cells are generated in the medulla through high affinity interactions with medullary thymic epithelial cells (mTEC) expressing the Autoimmune regulator (Aire). Recent advances have revealed that thymic Treg contain not only developing but also recirculating cells from the periphery. Although Aire is implicated in the generation of Foxp3+ Treg , its role in the biology of recirculating Treg remains elusive. Here, we show that Aire regulates the suppressive signature of recirculating Treg independently of the remodeling of the medullary 3D organization throughout life where Treg reside. Accordingly, the adoptive transfer of peripheral Foxp3+ Treg in AireKO recipients led to an impaired suppressive signature upon their entry into the thymus. Furthermore, recirculating Treg from AireKO mice failed to attenuate the severity of multiorgan autoimmunity, demonstrating that their suppressive function is altered. Using bone marrow chimeras, we reveal that mTEC-specific expression of Aire controls the suppressive signature of recirculating Treg . Finally, mature mTEC lacking Aire were inefficient in stimulating peripheral Treg both in polyclonal and antigen-specific co-culture assays. Overall, this study demonstrates that Aire confers to mTEC the ability to restimulate recirculating Treg , unravelling a novel function for this master regulator in Treg biology. [ABSTRACT FROM AUTHOR]- Published
- 2022
- Full Text
- View/download PDF
6. Nidogen-1 Contributes to the Interaction Network Involved in Pro-B Cell Retention in the Peri-sinusoidal Hematopoietic Stem Cell Niche
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Balzano, Marielle, De Grandis, Maria, Vu Manh, Thien-Phong, Chasson, Lionel, Bardin, Florence, Farina, Anne, Sergé, Arnauld, Bidaut, Ghislain, Charbord, Pierre, Hérault, Léonard, Bailly, Anne-Laure, Cartier-Michaud, Amandine, Boned, Annie, Dalod, Marc, Duprez, Estelle, Genever, Paul, Coles, Mark, Bajenoff, Marc, Xerri, Luc, Aurrand-Lions, Michel, Schiff, Claudine, Mancini, Stéphane, Mancini, Stéphane J.C., Centre de Recherche en Cancérologie de Marseille (CRCM), Aix Marseille Université (AMU)-Institut Paoli-Calmettes, Fédération nationale des Centres de lutte contre le Cancer (FNCLCC)-Fédération nationale des Centres de lutte contre le Cancer (FNCLCC)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Centre d'Immunologie de Marseille - Luminy (CIML), Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Aix Marseille Université (AMU), Génétique, immunothérapie, chimie et cancer (GICC), UMR 6239 CNRS [2008-2011] (GICC UMR 6239 CNRS), Université de Tours-Centre National de la Recherche Scientifique (CNRS), Informatique, Biologie Intégrative et Systèmes Complexes (IBISC), Université d'Évry-Val-d'Essonne (UEVE), University of York [York, UK], Institut de pharmacologie moléculaire et cellulaire (IPMC), Université Nice Sophia Antipolis (... - 2019) (UNS), Université Côte d'Azur (UCA)-Université Côte d'Azur (UCA)-Centre National de la Recherche Scientifique (CNRS), Département de Biopathologie, Institut Paoli-Calmettes, Fédération nationale des Centres de lutte contre le Cancer (FNCLCC)-Fédération nationale des Centres de lutte contre le Cancer (FNCLCC), Laboratoire des images et des signaux (LIS), Centre National de la Recherche Scientifique (CNRS)-Institut National Polytechnique de Grenoble (INPG)-Université Joseph Fourier - Grenoble 1 (UJF), Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Paoli-Calmettes, Fédération nationale des Centres de lutte contre le Cancer (FNCLCC)-Fédération nationale des Centres de lutte contre le Cancer (FNCLCC)-Aix Marseille Université (AMU), Fédération nationale des Centres de lutte contre le Cancer (FNCLCC), Mi-mAbs (C/O CIML), Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Aix Marseille Université (AMU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Aix Marseille Université (AMU), Laboratoire de Biologie du Développement [Paris] (LBD), Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS)-Institut de Biologie Paris Seine (IBPS), Sorbonne Université (SU)-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), Institut de Biologie Paris Seine (IBPS), Sorbonne Université (SU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Aix Marseille Université (AMU), Groupe innovation et ciblage cellulaire (GICC), EA 7501 [2018-...] (GICC EA 7501), Université de Tours, University of Oxford [Oxford], Aix Marseille Université (AMU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Aix Marseille Université (AMU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Aix Marseille Université (AMU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Laboratoire de Biologie du Développement [IBPS] (LBD), Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)-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), Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS), CCSD, Accord Elsevier, and ANR-17-CE14-0019,OSTEOVALYMPH,Autocrinie et paracrinie de l'axe de signalisation CXCL12/CXCR4-CXCR7 dans la niche ostéo-vasculaire: impact sur la spécification et l'engagement lymphoïde des cellules souches hématopoïétiques(2017)
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[SDV.MHEP.HEM] Life Sciences [q-bio]/Human health and pathology/Hematology ,bone marrow ,interaction network ,[SDV]Life Sciences [q-bio] ,[SDV.CAN]Life Sciences [q-bio]/Cancer ,Mice ,[SDV.CAN] Life Sciences [q-bio]/Cancer ,[SDV.BC.IC]Life Sciences [q-bio]/Cellular Biology/Cell Behavior [q-bio.CB] ,B cell development ,Animals ,Stem Cell Niche ,lcsh:QH301-705.5 ,ComputingMilieux_MISCELLANEOUS ,Mice, Knockout ,[SDV.MHEP] Life Sciences [q-bio]/Human health and pathology ,Membrane Glycoproteins ,Interleukin-7 ,Precursor Cells, B-Lymphoid ,stromal cell niche ,[SDV.MHEP.HEM]Life Sciences [q-bio]/Human health and pathology/Hematology ,Hematopoietic Stem Cells ,[SDV] Life Sciences [q-bio] ,[SDV.IMM.IA]Life Sciences [q-bio]/Immunology/Adaptive immunology ,lcsh:Biology (General) ,hematopoietic stem cell ,Stromal Cells ,[SDV.MHEP]Life Sciences [q-bio]/Human health and pathology - Abstract
Summary: In the bone marrow, CXCL12 and IL-7 are essential for B cell differentiation, whereas hematopoietic stem cell (HSC) maintenance requires SCF and CXCL12. Peri-sinusoidal stromal (PSS) cells are the main source of IL-7, but their characterization as a pro-B cell niche remains limited. Here, we characterize pro-B cell supporting stromal cells and decipher the interaction network allowing pro-B cell retention. Preferential contacts are found between pro-B cells and PSS cells, which homogeneously express HSC and B cell niche genes. Furthermore, pro-B cells are frequently located in the vicinity of HSCs in the same niche. Using an interactome bioinformatics pipeline, we identify Nidogen-1 as essential for pro-B cell retention in the peri-sinusoidal niche as confirmed in Nidogen-1−/− mice. Finally, human pro-B cells and hematopoietic progenitors are observed close to similar IL-7+ stromal cells. Thus, a multispecific niche exists in mouse and human supporting both early progenitors and committed hematopoietic lineages. : Balzano et al. show that bone marrow peri-sinusoidal stromal cells, which form the hematopoietic stem cell niche, also express B cell niche genes including IL-7 and Nidogen-1. Loss of Nidogen-1 expression specifically affects access of pro-B cells to IL-7, resulting in impaired expansion and differentiation of early B cells. Keywords: B cell development, hematopoietic stem cell, stromal cell niche, interaction network, bone marrow
- Published
- 2019
7. Genetic, structural, and chemical insights into the dual function of GRASP55 in germ cell Golgi remodeling and JAM-C polarized localization during spermatogenesis
- Author
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Cartier-Michaud, Amandine, Bailly, Anne-Laure, Betzi, Stéphane, Shi, Xiaoli, Lissitzky, Jean-Claude, Zarubica, Ana, Sergé, Arnauld, ROCHE, Philippe, Lugari, Adrien, HAMON, Véronique, Bardin, Florence, Derviaux, Carine, Lembo, Frédérique, Audebert, Stéphane, Marchetto, Sylvie, Durand, Bénédicte, Borg, Jean-Paul, Shi, Ning, Morelli, Xavier, Aurrand-Lions, Michel, Institut Paoli-Calmettes, Fédération nationale des Centres de lutte contre le Cancer (FNCLCC), Centre de Recherche en Cancérologie de Marseille (CRCM), Aix Marseille Université (AMU)-Institut Paoli-Calmettes, Fédération nationale des Centres de lutte contre le Cancer (FNCLCC)-Fédération nationale des Centres de lutte contre le Cancer (FNCLCC)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Centre d'Immunologie de Marseille - Luminy (CIML), Aix Marseille Université (AMU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), ANR-11-IDEX-0001,Amidex,INITIATIVE D'EXCELLENCE AIX MARSEILLE UNIVERSITE(2011), and Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Aix Marseille Université (AMU)
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Male ,Physiology ,[SDV]Life Sciences [q-bio] ,Golgi Apparatus ,Mice ,Animal Cells ,Reproductive Physiology ,Spermatocytes ,Medicine and Health Sciences ,Cell Cycle and Cell Division ,Testes ,Cells, Cultured ,Staining ,Chromosome Biology ,Intracellular Signaling Peptides and Proteins ,Specimen preparation and treatment ,Spermatids ,humanities ,Protein Transport ,Meiosis ,Seminiferous tubules ,Cell Processes ,cardiovascular system ,Cellular Types ,Anatomy ,Cellular Structures and Organelles ,Genital Anatomy ,Protein Binding ,Research Article ,endocrine system ,lcsh:QH426-470 ,education ,Immunoglobulins ,[SDV.BC]Life Sciences [q-bio]/Cellular Biology ,Germ cells ,Acrosomes ,Animals ,Spermatogenesis ,Binding Sites ,fungi ,Reproductive System ,DAPI staining ,Membrane Proteins ,Biology and Life Sciences ,Cell Biology ,Spermatogonia ,Sperm ,Mice, Inbred C57BL ,Research and analysis methods ,lcsh:Genetics ,Nuclear staining ,Carrier Proteins ,Cell Adhesion Molecules - Abstract
Spermatogenesis is a dynamic process that is regulated by adhesive interactions between germ and Sertoli cells. Germ cells express the Junctional Adhesion Molecule-C (JAM-C, encoded by Jam3), which localizes to germ/Sertoli cell contacts. JAM-C is involved in germ cell polarity and acrosome formation. Using a proteomic approach, we demonstrated that JAM-C interacted with the Golgi reassembly stacking protein of 55 kDa (GRASP55, encoded by Gorasp2) in developing germ cells. Generation and study of Gorasp2-/- mice revealed that knock-out mice suffered from spermatogenesis defects. Acrosome formation and polarized localization of JAM-C in spermatids were altered in Gorasp2-/- mice. In addition, Golgi morphology of spermatocytes was disturbed in Gorasp2-/- mice. Crystal structures of GRASP55 in complex with JAM-C or JAM-B revealed that GRASP55 interacted via PDZ-mediated interactions with JAMs and induced a conformational change in GRASP55 with respect of its free conformation. An in silico pharmacophore approach identified a chemical compound called Graspin that inhibited PDZ-mediated interactions of GRASP55 with JAMs. Treatment of mice with Graspin hampered the polarized localization of JAM-C in spermatids, induced the premature release of spermatids and affected the Golgi morphology of meiotic spermatocytes., Author summary Spermatogenesis defects are a common cause of male sterility. Spermatogenesis occurs in the seminiferous tubules of the testes and involves adhesive interactions between developing germ cells and Sertoli cells. Knock-out mouse models identified several adhesion molecules that are critically involved in spermatogenesis. We previously demonstrated that the Junctional Adhesion Molecule-C (JAM-C) plays a crucial role in establishing spermatids polarity. The latter involves rearrangements of the Golgi apparatus in spermatids which contribute to acrosome formation. The present study demonstrated that the C-terminal cytosolic region of JAM-C interacted with the Golgi reassembly stacking protein of 55 kDa (GRASP55) encoded by Gorasp2 and that spermatogenesis was impaired in Gorasp2-deficient mice. We developed an inhibitor of GRASP55 interaction with JAM-C and demonstrated that treatment of wild-type mice with the inhibitory compound induced germ cell loss. Therefore, the male infertility-associated pathway identified in this study is important not only from a genetic point of view, but also as a potential target for male contraception.
- Published
- 2017
8. Mouvements cellulaires et moléculaires
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Sergé, Arnauld, Irla, Magali, Sergé, Arnauld, Aix Marseille Université (AMU), Adhésion et Inflammation (LAI), Aix Marseille Université (AMU)-Assistance Publique - Hôpitaux de Marseille (APHM)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Centre d'Immunologie de Marseille - Luminy (CIML), and Aix Marseille Université (AMU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)
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[SDV] Life Sciences [q-bio] ,[SDV]Life Sciences [q-bio] ,ComputingMilieux_MISCELLANEOUS - Abstract
International audience
- Published
- 2013
9. Mutation of FOP/FGFR1OP in mice recapitulates human short rib-polydactyly ciliopathy.
- Author
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Cabaud, Olivier, Roubin, Régine, Comte, Audrey, Bascunana, Virginie, Sergé, Arnauld, Sedjaï, Fatima, Birnbaum, Daniel, Rosnet, Olivier, and Acquaviva, Claire
- Published
- 2018
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10. CD154 Costimulation Shifts the Local T-Cell Receptor Repertoire Not Only During Thymic Selection but Also During Peripheral T-Dependent Humoral Immune Responses.
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Fähnrich, Anke, Klein, Sebastian, Sergé, Arnauld, Nyhoegen, Christin, Kombrink, Sabrina, Möller, Steffen, Keller, Karsten, Westermann, Jürgen, and Kalies, Kathrin
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CD antigens ,T cell receptors ,HUMORAL immunity - Abstract
CD154 is a transmembrane cytokine expressed transiently on activated CD4 T cells upon T-cell receptor (TCR) stimulation that interacts with CD40 on antigen-presenting cells. The signaling via CD154:CD40 is essential for B-cell maturation and germinal center formation and also for the final differentiation of CD4 T cells during T-dependent humoral immune responses. Recent data demonstrate that CD154 is critically involved in the selection of T-cell clones during the negative selection process in the thymus. Whether CD154 signaling influences the TCR repertoire during peripheral T-dependent humoral immune responses has not yet been elucidated. To find out, we used CD154- deficient mice and assessed the global TCRβ repertoire in T-cell zones (TCZ) of spleens by high-throughput sequencing after induction of a Th2 response to the multiepitopic antigen sheep red blood cells. Qualitative and quantitative comparison of the splenic TCZ-specific TCRß repertoires revealed that CD154 deficiency shifts the distribution of Vβ-Jβ genes after antigen exposure. This data led to the conclusion that costimulation via CD154:CD40 during the interaction of T cells with CD40-matured B cells contributes to the recruitment of T-cell clones into the immune response and thereby shapes the peripheral TCR repertoire. [ABSTRACT FROM AUTHOR]
- Published
- 2018
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11. Lymphotoxin α fine-tunes T cell clonal deletion by regulating thymic entry of antigen-presenting cells.
- Author
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Lopes, Noëlla, Charaix, Jonathan, Cédile, Oriane, Sergé, Arnauld, and Irla, Magali
- Abstract
Medullary thymic epithelial cells (mTEC) purge the T cell repertoire of autoreactive thymocytes. Although dendritic cells (DC) reinforce this process by transporting innocuous peripheral self-antigens, the mechanisms that control their thymic entry remain unclear. Here we show that mTEC-CD4
+ thymocyte crosstalk regulates the thymus homing of SHPS-1+ conventional DCs (cDC), plasmacytoid DCs (pDC) and macrophages. This homing process is controlled by lymphotoxin α (LTα), which negatively regulates CCL2, CCL8 and CCL12 chemokines in mTECs. Consequently, Ltα-deficient mice have increased expression of these chemokines that correlates with augmented classical NF-κB subunits and increased thymic recruitment of cDCs, pDCs and macrophages. This enhanced migration depends mainly on the chemokine receptor CCR2, and increases thymic clonal deletion. Altogether, this study identifies a fine-tuning mechanism of T cell repertoire selection and paves the way for therapeutic interventions to treat autoimmune disorders. [ABSTRACT FROM AUTHOR]- Published
- 2018
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12. Multiple-target tracing (MTT) algorithm probes molecular dynamics at cell surface
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Bertaux Nicolas, Marguet Didier, Sergé Arnauld, Rigneault Hervé, and Aix Marseille Université (AMU)
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Surface (mathematics) ,Chemistry ,[SDV]Life Sciences [q-bio] ,Cell ,Tracing ,Multiple target ,Cell biology ,Molecular dynamics ,medicine.anatomical_structure ,medicine ,General Earth and Planetary Sciences ,Biological system ,ComputingMilieux_MISCELLANEOUS ,General Environmental Science - Abstract
International audience
- Published
- 2008
13. For3D: Full organ reconstruction in 3D, an automatized tool for deciphering the complexity of lymphoid organs.
- Author
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Sergé, Arnauld, Bailly, Anne-Laure, Aurrand-Lions, Michel, Imhof, Beat A., and Irla, Magali
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LYMPHOID tissue , *IMMUNOLOGY , *IMMUNOHISTOCHEMISTRY , *IMMUNOFLUORESCENCE , *LYMPH nodes , *TUMORS - Abstract
To decipher the complex topology of lymphoid structures, we developed an automated process called Full Organ Reconstruction in 3D (For3D). A dedicated image-processing pipeline is applied to entire collections of immunolabeled serial sections, acquired with a slide-scanning microscope. This method is automated, flexible and readily applicable in two days to frozen or paraffin-embedded organs stained by fluorescence or brightfield immunohistochemistry. 3D-reconstructed organs can be visualized, rotated and analyzed to quantify substructures of interest. Usefulness of For3D is exemplified here through topological analysis of several mouse lymphoid organs exhibiting a complex organization: (i) the thymus, composed of two compartments, a medulla intricately imbricated into a surrounding cortex, (ii) lymph nodes, also highly compartmentalized into cortex, paracortex and medulla and (iii) the vascularization of an EG7 primary thymoma. This open-source algorithm, based on ImageJ and Matlab scripts, offers a user-friendly interface and is widely applicable to any organ or tissue, hence readily adaptable to a broad range of biomedical samples. [ABSTRACT FROM AUTHOR]
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- 2015
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14. Thymic crosstalk coordinates medulla organization and T-cell tolerance induction.
- Author
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Lopes, Noëlla, Sergé, Arnauld, Ferrier, Pierre, and Irla, Magali
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T cells ,EPITHELIAL cells ,DENDRITIC cells ,CELL differentiation ,T cell receptors - Abstract
The thymus ensures the generation of a functional and highly diverse T-cell repertoire. The thymic medulla, which is mainly composed of medullary thymic epithelial cells (mTECs) and dendritic cells (DCs), provides a specialized microenvironment dedicated to the establishment of T-cell tolerance. mTECs play a privileged role in this pivotal process by their unique capacity to express a broad range of peripheral self-antigens that are presented to developing T cells. Reciprocally, developing T cells control mTEC differentiation and organization. These bidirectional interactions are commonly referred to as thymic crosstalk. This review focuses on the relative contributions of mTEC and DC subsets to the deletion of autoreactive T cells and the generation of natural regulatory T cells. We also summarize current knowledge regarding how hematopoietic cells conversely control the composition and complex three-dimensional organization of the thymic medulla. [ABSTRACT FROM AUTHOR]
- Published
- 2015
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15. Barcoding T Cell Calcium Response Diversity with Methods for Automated and Accurate Analysis of Cell Signals (MAAACS).
- Author
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Salles, Audrey, Billaudeau, Cyrille, Sergé, Arnauld, Bernard, Anne-Marie, Phélipot, Marie-Claire, Bertaux, Nicolas, Fallet, Mathieu, Grenot, Pierre, Marguet, Didier, He, Hai-Tao, and Hamon, Yannick
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BAR codes ,T cells ,CELL-mediated lympholysis ,CALCIUM metabolism ,CELL proliferation ,PHYSIOLOGY - Abstract
We introduce a series of experimental procedures enabling sensitive calcium monitoring in T cell populations by confocal video-microscopy. Tracking and post-acquisition analysis was performed using Methods for Automated and Accurate Analysis of Cell Signals (MAAACS), a fully customized program that associates a high throughput tracking algorithm, an intuitive reconnection routine and a statistical platform to provide, at a glance, the calcium barcode of a population of individual T-cells. Combined with a sensitive calcium probe, this method allowed us to unravel the heterogeneity in shape and intensity of the calcium response in T cell populations and especially in naive T cells, which display intracellular calcium oscillations upon stimulation by antigen presenting cells. [ABSTRACT FROM AUTHOR]
- Published
- 2013
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16. Antigen Recognition by Autoreactive CD4+ Thymocytes Drives Homeostasis of the Thymic Medulla.
- Author
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Irla, Magali, Guerri, Lucia, Guenot, Jeanne, Sergé, Arnauld, Lantz, Olivier, Liston, Adrian, Imhof, Beat A., Palmer, Ed, and Reith, Walter
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IMMUNOGLOBULINS ,HOMEOSTASIS ,T cell receptors ,EPITHELIAL cells ,THYMOCYTES ,MEDICAL research - Abstract
The thymic medulla is dedicated for purging the T-cell receptor (TCR) repertoire of self-reactive specificities. Medullary thymic epithelial cells (mTECs) play a pivotal role in this process because they express numerous peripheral tissue-restricted self-antigens. Although it is well known that medulla formation depends on the development of single-positive (SP) thymocytes, the mechanisms underlying this requirement are incompletely understood. We demonstrate here that conventional SP CD4
+ thymocytes bearing autoreactive TCRs drive a homeostatic process that fine-tunes medullary plasticity in adult mice by governing the expansion and patterning of the medulla. This process exhibits strict dependence on TCR-reactivity with self-antigens expressed by mTECs, as well as engagement of the CD28-CD80/CD86 costimulatory axis. These interactions induce the expression of lymphotoxin a in autoreactive CD4+ thymocytes and RANK in mTECs. Lymphotoxin in turn drives mTEC development in synergy with RANKL and CD40L. Our results show that Ag-dependent interactions between autoreactive CD4+ thymocytes and mTECs fine-tune homeostasis of the medulla by completing the signaling axes implicated in mTEC expansion and medullary organization. [ABSTRACT FROM AUTHOR]- Published
- 2012
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17. Quantification of GPCR internalization by single-molecule microscopy in living cells.
- Author
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Sergé, Arnauld, de Keijzer, Sandra, Van Hemert, Freek, Hickman, Mark R., Hereld, Dale, Spaink, Herman P., Schmidt, Thomas, and Snaar-Jagalska, B. Ewa
- Published
- 2011
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18. Dynamic multiple-target tracing to probe spatiotemporal cartography of cell membranes.
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Sergé, Arnauld, Bertaux, Nicolas, Rigneault, Hervé, and Marguet, Didier
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CELL membranes , *BIOLOGICAL membranes , *PARTICLES , *QUANTUM dots , *MOLECULAR dynamics - Abstract
Although the highly dynamic and mosaic organization of the plasma membrane is well-recognized, depicting a resolved, global view of this organization remains challenging. We present an analytical single-particle tracking (SPT) method and tool, multiple-target tracing (MTT), that takes advantage of the high spatial resolution provided by single-fluorophore sensitivity. MTT can be used to generate dynamic maps at high densities of tracked particles, thereby providing global representation of molecular dynamics in cell membranes. Deflation by subtracting detected peaks allows detection of lower-intensity peaks. We exhaustively detected particles using MTT, with performance reaching theoretical limits, and then reconnected trajectories integrating the statistical information from past trajectories. We demonstrate the potential of this method by applying it to the epidermal growth factor receptor (EGFR) labeled with quantum dots (Qdots), in the plasma membrane of live cells. We anticipate the use of MTT to explore molecular dynamics and interactions at the cell membrane. [ABSTRACT FROM AUTHOR]
- Published
- 2008
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19. Active surface transport of metabotropic glutamate receptors through binding to microtubules and actin flow.
- Author
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Sergé, Arnauld, Fourgeaud, Lawrence, Hémar, Agnés, and Choquet, Daniel
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NEUROTRANSMITTERS , *ACTIN , *MICROTUBULES , *SYNAPSES , *CYTOSKELETAL proteins , *CELL membranes - Abstract
Receptors for neurotransmitters are concentrated and stabilized at given sites such as synapses through interactions with scaffolding proteins and cytoskeletal elements. The transport of receptors first involves directed vesicular trafficking of intracellularly stored receptors followed by their targeting to the plasma membrane. Once expressed at the cell surface, receptors are thought to reach their final location by random Brownian diffusion in the plasma membrane plane. Here, we investigate whether the metabotropic glutamate receptor mGluR5 can also be transported actively on the cell surface. We used single particle tracking to follow mGluR5 movement in real time at the surface of neuronal growth cones or fibroblast lamellipodia, both of which bear a particularly active cytoskeleton. We found that after a certain lag time mGluR5 undergoes directed rearward transport, which depends on actin flow. On actin depolymerization, directed movement was suppressed, but receptors still bound to a rigid structure. By contrast, receptor transport and immobilization was fully suppressed by microtubule depolymerization but favored by microtubule stabilization. Furthermore, mGluR5 could be immunoprecipitated with tubulin from rat brains, confirming the ability of mGluR5 to bind to microtubules. We propose that mGluR5 can be transported on the cell surface through actin-mediated retrograde transport of microtubules. This process may play a role in receptor targeting and organization during synapse formation or during glutamate-mediated growth cone chemotaxis. [ABSTRACT FROM AUTHOR]
- Published
- 2003
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20. Analyzing normal and disrupted leukemic stem cell adhesion to bone marrow stromal cells by single-molecule tracking nanoscopy.
- Author
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Gorshkova, Oksana, Cappaï, Jessica, Maillot, Loriane, and Sergé, Arnauld
- Subjects
MESENCHYMAL stem cells ,STEM cells ,SINGLE molecules ,STROMAL cells ,CELL adhesion ,IMMUNOGLOBULINS - Abstract
Leukemic stem cells (LSCs) adhere to bone niches through adhesion molecules. These interactions, which are deeply reorganized in tumors, contribute to LSC resistance to chemotherapy and leukemia relapse. However, LSC adhesion mechanisms and potential therapeutic disruption using blocking antibodies remain largely unknown. Junctional adhesion molecule C (JAM-C, also known as JAM3) overexpression by LSCs correlates with increased leukemia severity, and thus constitutes a putative therapeutic target. Here, we took advantage of the ability of nanoscopy to detect single molecules with nanometric accuracy to characterize junctional adhesion molecule (JAM) dynamics at leuko-stromal contacts. Videonanoscopy trajectories were reconstructed using our dedicated multi-target tracing algorithm, pipelined with dual-color analyses (MTT2col). JAM-C expressed by LSCs engaged in transient interactions with JAM-B (also known as JAM2) expressed by stromal cells. JAM recruitment and colocalization at cell contacts were proportional to JAM-C level and reduced by a blocking anti-JAM-C antibody. MTT2col revealed, at single-molecule resolution, the ability of blocking antibodies to destabilize LSC binding to their niches, opening opportunities for disrupting LSC resistance mechanisms. [ABSTRACT FROM AUTHOR]
- Published
- 2021
- Full Text
- View/download PDF
21. Signalling complexes and clusters: functional advantages and methodological hurdles.
- Author
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Cebecauer, Marek, Spitaler, Martin, Sergé, Arnauld, and Magee, Anthony I.
- Subjects
CELLULAR signal transduction ,MACROMOLECULES ,CELL migration ,IMAGING systems ,MICROMETERS - Abstract
Signalling molecules integrate, codify and transport information in cells. Organisation of these molecules in complexes and clusters improves the efficiency, fidelity and robustness of cellular signalling. Here, we summarise current views on how signalling molecules assemble into macromolecular complexes and clusters and how they use their physical properties to transduce environmental information into a variety of cellular processes. In addition, we discuss recent innovations in live-cell imaging at the sub-micrometer scale and the challenges of object (particle) tracking, both of which help us to observe signalling complexes and clusters and to examine their dynamic character. [ABSTRACT FROM AUTHOR]
- Published
- 2010
- Full Text
- View/download PDF
22. Cancer-associated fibroblast-derived annexin A6+ extracellular vesicles support pancreatic cancer aggressiveness.
- Author
-
Leca, Julie, Martinez, Sébastien, Lac, Sophie, Nigri, Jérémy, Secq, Véronique, Rubis, Marion, Bressy, Christian, Sergé, Arnauld, Lavaut, Marie-Noelle, Dusetti, Nelson, Loncle, Céline, Roques, Julie, Pietrasz, Daniel, Bousquet, Corinne, Garcia, Stéphane, Granjeaud, Samuel, Ouaissi, Mehdi, Bachet, Jean Baptiste, Brun, Christine, and Iovanna, Juan L.
- Subjects
- *
STROMAL cells , *ADENOCARCINOMA , *DUCTAL carcinoma , *CANCER cells , *TUMOR markers , *METASTASIS - Abstract
The intratumoral microenvironment, or stroma, is of major importance in the pathobiology of pancreatic ductal adenocarcinoma (PDA), and specific conditions in the stroma may promote increased cancer aggressiveness. We hypothesized that this heterogeneous and evolving compartment drastically influences tumor cell abilities, which in turn influences PDA aggressiveness through crosstalk that is mediated by extracellular vesicles (EVs). Here, we have analyzed the PDA proteomic stromal signature and identified a contribution of the annexin A6/LDL receptor-related protein 1/thrombospondin 1 (ANXA6/LRP1/TSP1) complex in tumor cell crosstalk. Formation of the ANXA6/LRP1/TSP1 complex was restricted to cancer-associated fibroblasts (CAFs) and required physiopathologic culture conditions that improved tumor cell survival and migration. Increased PDA aggressiveness was dependent on tumor cell-mediated uptake of CAF-derived ANXA6+ EVs carrying the ANXA6/LRP1/TSP1 complex. Depletion of ANXA6 in CAFs impaired complex formation and subsequently impaired PDA and metastasis occurrence, while injection of CAF-derived ANXA6+ EVs enhanced tumorigenesis. We found that the presence of ANXA6+ EVs in serum was restricted to PDA patients and represents a potential biomarker for PDA grade. These findings suggest that CAF-tumor cell crosstalk supported by ANXA6+ EVs is predictive of PDA aggressiveness, highlighting a therapeutic target and potential biomarker for PDA. [ABSTRACT FROM AUTHOR]
- Published
- 2016
- Full Text
- View/download PDF
23. PRICKLE1 Contributes to Cancer Cell Dissemination through Its Interaction with mTORC2.
- Author
-
Daulat, Avais M., Bertucci, François, Audebert, Stéphane, Sergé, Arnauld, Finetti, Pascal, Josselin, Emmanuelle, Castellano, Rémy, Birnbaum, Daniel, Angers, Stéphane, and Borg, Jean-Paul
- Subjects
- *
CANCER cells , *CELL communication , *CANCER invasiveness , *PRICKLE1 gene , *MTOR protein , *PROGRESSION-free survival - Abstract
Summary Components of the evolutionarily conserved developmental planar cell polarity (PCP) pathway were recently described to play a prominent role in cancer cell dissemination. However, the molecular mechanisms by which PCP molecules drive the spread of cancer cells remain largely unknown. PRICKLE1 encodes a PCP protein bound to the promigratory serine/threonine kinase MINK1. We identify RICTOR, a member of the mTORC2 complex, as a PRICKLE1-binding partner and show that the integrity of the PRICKLE1-MINK1-RICTOR complex is required for activation of AKT, regulation of focal adhesions, and cancer cell migration. Disruption of the PRICKLE1-RICTOR interaction results in a strong impairment of breast cancer cell dissemination in xenograft assays. Finally, we show that upregulation of PRICKLE1 in basal breast cancers, a subtype characterized by high metastatic potential, is associated with poor metastasis-free survival. [ABSTRACT FROM AUTHOR]
- Published
- 2016
- Full Text
- View/download PDF
24. Three-Dimensional Visualization of the Mouse Thymus Organization in Health and Immunodeficiency.
- Author
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Irla, Magali, Guenot, Jeanne, Sealy, Gregg, Reith, Walter, Imhof, Beat A., and Sergé, Arnauld
- Subjects
- *
LABORATORY mice , *IMMUNODEFICIENCY , *LYMPHOID tissue , *THYMUS , *THREE-dimensional imaging in biology , *HEALTH - Abstract
Lymphoid organs exhibit complex structures tightly related to their function. Surprisingly, although the thymic medulla constitutes a specialized microenvironment dedicated to the induction of T cell tolerance, its three-dimensional topology remains largely elusive because it has been studied mainly in two dimensions using thymic sections. To overcome this limitation, we have developed an automated method for full organ reconstruction in three dimensions, allowing visualization of intact mouse lymphoid organs from a collection of immunolabeled slices. We validated full organ reconstruction in three dimensions by reconstructing the well-characterized structure of skin-draining lymph nodes, before revisiting the complex and poorly described corticomedullary organization of the thymus. Wild-type thymi contain ~200 small medullae that are connected to or separated from a major medullary compartment. In contrast, thymi of immunodeficient Rag2-/- mice exhibit only ~20 small, unconnected medullary islets. Upon total body irradiation, medullary complexity was partially reduced and then recovered upon bone marrow transplantation. This intricate topology presents fractal properties, resulting in a considerable corticomedullary area. This feature ensures short distances between cortex and medulla, hence efficient thymocyte migration, as assessed by mathematical models. Remarkably, this junction is enriched, particularly in neonates, in medullary thymic epithelial cells expressing the autoimmune regulator. The emergence of a major medullary compartment is induced by CD4+ thymocytes via CD80/86 and lymphotoxin-α signals. This comprehensive three-dimensional view of the medulla emphasizes a complex topology favoring efficient interactions between developing T cells and autoimmune regulator-positive medullary thymic epithelial cells, a key process for central tolerance induction. [ABSTRACT FROM AUTHOR]
- Published
- 2013
- Full Text
- View/download PDF
25. Differential Requirement for CCR4 and CCR7 during the Development of Innate and Adaptive αβT Cells in the Adult Thymus
- Author
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Graham Anderson, Arnauld Sergé, William E. Jenkinson, Andrea J. White, Jennifer E. Cowan, Magali Irla, Peter J. L. Lane, Sonia M. Parnell, Nicholas I. McCarthy, Andrea Bacon, Eric J. Jenkinson, Sergé, Arnauld, Medical Research Council Centre for Immune Regulation, Institute for Biomedical Research, University of Birmingham, Centre de Recherche en Cancérologie de Marseille (CRCM), Aix Marseille Université (AMU)-Institut Paoli-Calmettes, Fédération nationale des Centres de lutte contre le Cancer (FNCLCC)-Fédération nationale des Centres de lutte contre le Cancer (FNCLCC)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Centre d'Immunologie de Marseille - Luminy (CIML), and Aix Marseille Université (AMU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)
- Subjects
Receptors, CCR7 ,Receptors, CCR4 ,Regulatory T cell ,Cellular differentiation ,T cell ,Receptors, Antigen, T-Cell, alpha-beta ,[SDV]Life Sciences [q-bio] ,Immunology ,Thymus Gland ,Biology ,Adaptive Immunity ,T-Lymphocytes, Regulatory ,Clonal deletion ,Mice ,T-Lymphocyte Subsets ,medicine ,Immunology and Allergy ,Animals ,Cell Lineage ,ComputingMilieux_MISCELLANEOUS ,Mice, Knockout ,Immune System Development ,FOXP3 ,Cell Differentiation ,Epithelial Cells ,Natural killer T cell ,Immunity, Innate ,Cell biology ,Mice, Inbred C57BL ,[SDV] Life Sciences [q-bio] ,Thymocyte ,medicine.anatomical_structure ,Thymocyte migration ,Biomarkers - Abstract
αβT cell development depends upon serial migration of thymocyte precursors through cortical and medullary microenvironments, enabling specialized stromal cells to provide important signals at specific stages of their development. Although conventional αβT cells are subject to clonal deletion in the medulla, entry into the thymus medulla also fosters αβT cell differentiation. For example, during postnatal periods, the medulla is involved in the intrathymic generation of multiple αβT cell lineages, notably the induction of Foxp3+ regulatory T cell development and the completion of invariant NKT cell development. Although migration of conventional αβT cells to the medulla is mediated by the chemokine receptor CCR7, how other T cell subsets gain access to medullary areas during their normal development is not clear. In this study, we show that combining a panel of thymocyte maturation markers with cell surface analysis of CCR7 and CCR4 identifies distinct stages in the development of multiple αβT cell lineages in the thymus. Although Aire regulates expression of the CCR4 ligands CCL17 and CCL22, we show that CCR4 is dispensable for thymocyte migration and development in the adult thymus, demonstrating defective T cell development in Aire−/− mice is not because of a loss of CCR4-mediated migration. Moreover, we reveal that CCR7 controls the development of invariant NKT cells by enabling their access to IL-15 trans-presentation in the thymic medulla and influences the balance of early and late intrathymic stages of Foxp3+ regulatory T cell development. Collectively, our data identify novel roles for CCR7 during intrathymic T cell development, highlighting its importance in enabling multiple αβT cell lineages to access the thymic medulla.
- Published
- 2014
26. Single Molecule Tracking Nanoscopy Extended to Two Colors with MTT2col for the Analysis of Cell-Cell Interactions in Leukemia.
- Author
-
Maillot L, Irla M, and Sergé A
- Abstract
Single molecule tracking (SMT) is a powerful technique to study molecular dynamics, and is particularly adapted to monitor the motion and interactions of cell membrane components. Assessing interactions among two molecular populations is classically performed by several approaches, including dual-color videomicroscopy, which allows monitoring of interactions through colocalization events. Other techniques, such as fluorescence recovery after photobleaching (FRAP), Förster resonance energy transfer (FRET), and fluorescence correlation spectroscopy (FCS), are also utilized to measure molecular dynamics. We developed MTT2col, a set of algorithmic tools extending multi-target tracing (MTT) to dual-color acquisition (https://github.com/arnauldserge1/MTT2col). In this protocol, we used MTT2col to monitor adhesion molecules at the contact between leukemic stem cells and stromal cells, a process involved in cancer resistance to chemotherapy and in relapse. Our dual-color single molecule protocol includes the following steps: (i) labeling molecules of interest with fluorescent probes, (ii) video-acquisition, (iii) analyses using our MTT2col in-house software, to obtain positions and trajectories, followed by (iv) detailed analyses of colocalization, distribution, and dynamic motion modes, according to the issues addressed. MTT2col is a robust and efficient SMT algorithm. Both MTT and MTT2col are open-source software that can be adapted and further developed for specific analyses. Graphical abstract ., Competing Interests: Competing interestsWe declare no competing interest., (Copyright © 2022 The Authors; exclusive licensee Bio-protocol LLC.)
- Published
- 2022
- Full Text
- View/download PDF
27. EB1-dependent long survival of glioblastoma-grafted mice with the oral tubulin-binder BAL101553 is associated with inhibition of tumor angiogenesis.
- Author
-
Bergès R, Tchoghandjian A, Sergé A, Honoré S, Figarella-Branger D, Bachmann F, Lane HA, and Braguer D
- Abstract
Glioblastoma (GBM) are aggressive brain tumors with limited treatment options. Cancer stem-like cells (CSLCs) contribute to GBM invasiveness, representing promising targets. BAL101553, a prodrug of BAL27862, is a novel small molecule tubulin-binding agent, promoting tumor cell death through spindle assembly checkpoint activation, which is currently in Phase 1/2a in advanced solid tumor patients including GBM. This study aimed to evaluate long-term daily oral BAL101553 treatment of mice orthotopically grafted with GBM CSLCs (GBM6) according to EB1 expression-level, and to decipher its mechanism of action on GBM stem cells. Oral treatment with BAL101553 for 100 days provoked a large EB1 expression level-dependent survival benefit, together with a decrease in tumor growth and brain invasion. Formation of vascular structures by the fluorescent GBM6-GFP-sh0 cells, mimicking endothelial vascular networks, was observed in the brains of control grafted mice. Following BAL101553 treatment, vessels were no longer detectable, suggesting inhibition of the endothelial trans-differentiation of GBM stem cells. In vitro , BAL27862 treatment resulted in a switch to the endothelial-like phenotype of GBM6 towards an astrocytic phenotype. Moreover, the drug inhibited secretion of VEGF, thus preventing normal endothelial cell migration activated by CSLCs. The decrease in VEGF secretion was confirmed in a human GBM explant following drug treatment. Altogether, our data first confirm the potential of EB1 expression as a response-predictive biomarker of BAL101553 in GBM we previously published and add new insights in BAL101553 long-term action by counteracting CSLCs mediated tumor angiogenesis. Our results strongly support BAL101553 clinical studies in GBM patients., Competing Interests: CONFLICTS OF INTEREST HAL and FB are employed by Basilea Pharmaceutic International AG and own Basilea stocks/options. Basilea fundings partially contributed to experimental costs., (Copyright: © 2019 Bergès et al.)
- Published
- 2020
- Full Text
- View/download PDF
28. Nidogen-1 Contributes to the Interaction Network Involved in Pro-B Cell Retention in the Peri-sinusoidal Hematopoietic Stem Cell Niche.
- Author
-
Balzano M, De Grandis M, Vu Manh TP, Chasson L, Bardin F, Farina A, Sergé A, Bidaut G, Charbord P, Hérault L, Bailly AL, Cartier-Michaud A, Boned A, Dalod M, Duprez E, Genever P, Coles M, Bajenoff M, Xerri L, Aurrand-Lions M, Schiff C, and Mancini SJC
- Subjects
- Animals, Hematopoietic Stem Cells cytology, Interleukin-7 genetics, Interleukin-7 immunology, Membrane Glycoproteins genetics, Mice, Mice, Knockout, Precursor Cells, B-Lymphoid cytology, Stromal Cells cytology, Stromal Cells immunology, Hematopoietic Stem Cells immunology, Membrane Glycoproteins immunology, Precursor Cells, B-Lymphoid immunology, Stem Cell Niche immunology
- Abstract
In the bone marrow, CXCL12 and IL-7 are essential for B cell differentiation, whereas hematopoietic stem cell (HSC) maintenance requires SCF and CXCL12. Peri-sinusoidal stromal (PSS) cells are the main source of IL-7, but their characterization as a pro-B cell niche remains limited. Here, we characterize pro-B cell supporting stromal cells and decipher the interaction network allowing pro-B cell retention. Preferential contacts are found between pro-B cells and PSS cells, which homogeneously express HSC and B cell niche genes. Furthermore, pro-B cells are frequently located in the vicinity of HSCs in the same niche. Using an interactome bioinformatics pipeline, we identify Nidogen-1 as essential for pro-B cell retention in the peri-sinusoidal niche as confirmed in Nidogen-1
-/- mice. Finally, human pro-B cells and hematopoietic progenitors are observed close to similar IL-7+ stromal cells. Thus, a multispecific niche exists in mouse and human supporting both early progenitors and committed hematopoietic lineages., (Copyright © 2019 The Author(s). Published by Elsevier Inc. All rights reserved.)- Published
- 2019
- Full Text
- View/download PDF
29. JAM-C Identifies Src Family Kinase-Activated Leukemia-Initiating Cells and Predicts Poor Prognosis in Acute Myeloid Leukemia.
- Author
-
De Grandis M, Bardin F, Fauriat C, Zemmour C, El-Kaoutari A, Sergé A, Granjeaud S, Pouyet L, Montersino C, Chretien AS, Mozziconacci MJ, Castellano R, Bidaut G, Boher JM, Collette Y, Mancini SJC, Vey N, and Aurrand-Lions M
- Subjects
- ADP-ribosyl Cyclase 1 metabolism, Animals, Antigens, CD34 metabolism, Biomarkers, Tumor genetics, Cell Adhesion Molecules genetics, Cell Line, Tumor, Enzyme Activation, Female, Gene Expression Profiling, Humans, Interleukin-3 Receptor alpha Subunit metabolism, Membrane Glycoproteins metabolism, Mice, Mice, Inbred BALB C, Mice, Knockout, Neoplasm Transplantation, Neoplastic Stem Cells cytology, Transplantation, Heterologous, Biomarkers, Tumor metabolism, Cell Adhesion Molecules metabolism, Leukemia, Myeloid, Acute pathology, Neoplastic Stem Cells pathology, src-Family Kinases metabolism
- Abstract
Acute myeloid leukemia (AML) originates from hematopoietic stem and progenitor cells that acquire somatic mutations, leading to disease and clonogenic evolution. AML is characterized by accumulation of immature myeloid cells in the bone marrow and phenotypic cellular heterogeneity reflective of normal hematopoietic differentiation. Here, we show that JAM-C expression defines a subset of leukemic cells endowed with leukemia-initiating cell activity (LIC). Stratification of de novo AML patients at diagnosis based on JAM-C-expressing cells frequencies in the blood served as an independent prognostic marker for disease outcome. Using publicly available leukemic stem cell (LSC) gene expression profiles and gene expression data generated from JAM-C-expressing leukemic cells, we defined a single cell core gene expression signature correlated to JAM-C expression that reveals LSC heterogeneity. Finally, we demonstrated that JAM-C controls Src family kinase (SFK) activation in LSC and that LIC with exacerbated SFK activation was uniquely found within the JAM-C-expressing LSC compartment. Cancer Res; 77(23); 6627-40. ©2017 AACR ., (©2017 American Association for Cancer Research.)
- Published
- 2017
- Full Text
- View/download PDF
30. Genetic, structural, and chemical insights into the dual function of GRASP55 in germ cell Golgi remodeling and JAM-C polarized localization during spermatogenesis.
- Author
-
Cartier-Michaud A, Bailly AL, Betzi S, Shi X, Lissitzky JC, Zarubica A, Sergé A, Roche P, Lugari A, Hamon V, Bardin F, Derviaux C, Lembo F, Audebert S, Marchetto S, Durand B, Borg JP, Shi N, Morelli X, and Aurrand-Lions M
- Subjects
- Animals, Binding Sites, Carrier Proteins chemistry, Carrier Proteins genetics, Cells, Cultured, Golgi Apparatus ultrastructure, Intracellular Signaling Peptides and Proteins, Male, Membrane Proteins chemistry, Membrane Proteins genetics, Mice, Mice, Inbred C57BL, Protein Binding, Protein Transport, Spermatogonia cytology, Carrier Proteins metabolism, Cell Adhesion Molecules metabolism, Golgi Apparatus metabolism, Immunoglobulins metabolism, Membrane Proteins metabolism, Spermatogenesis, Spermatogonia metabolism
- Abstract
Spermatogenesis is a dynamic process that is regulated by adhesive interactions between germ and Sertoli cells. Germ cells express the Junctional Adhesion Molecule-C (JAM-C, encoded by Jam3), which localizes to germ/Sertoli cell contacts. JAM-C is involved in germ cell polarity and acrosome formation. Using a proteomic approach, we demonstrated that JAM-C interacted with the Golgi reassembly stacking protein of 55 kDa (GRASP55, encoded by Gorasp2) in developing germ cells. Generation and study of Gorasp2-/- mice revealed that knock-out mice suffered from spermatogenesis defects. Acrosome formation and polarized localization of JAM-C in spermatids were altered in Gorasp2-/- mice. In addition, Golgi morphology of spermatocytes was disturbed in Gorasp2-/- mice. Crystal structures of GRASP55 in complex with JAM-C or JAM-B revealed that GRASP55 interacted via PDZ-mediated interactions with JAMs and induced a conformational change in GRASP55 with respect of its free conformation. An in silico pharmacophore approach identified a chemical compound called Graspin that inhibited PDZ-mediated interactions of GRASP55 with JAMs. Treatment of mice with Graspin hampered the polarized localization of JAM-C in spermatids, induced the premature release of spermatids and affected the Golgi morphology of meiotic spermatocytes.
- Published
- 2017
- Full Text
- View/download PDF
31. The Molecular Architecture of Cell Adhesion: Dynamic Remodeling Revealed by Videonanoscopy.
- Author
-
Sergé A
- Abstract
The plasma membrane delimits the cell, which is the basic unit of living organisms, and is also a privileged site for cell communication with the environment. Cell adhesion can occur through cell-cell and cell-matrix contacts. Adhesion proteins such as integrins and cadherins also constitute receptors for inside-out and outside-in signaling within proteolipidic platforms. Adhesion molecule targeting and stabilization relies on specific features such as preferential segregation by the sub-membrane cytoskeleton meshwork and within membrane proteolipidic microdomains. This review presents an overview of the recent insights brought by the latest developments in microscopy, to unravel the molecular remodeling occurring at cell contacts. The dynamic aspect of cell adhesion was recently highlighted by super-resolution videomicroscopy, also named videonanoscopy. By circumventing the diffraction limit of light, nanoscopy has allowed the monitoring of molecular localization and behavior at the single-molecule level, on fixed and living cells. Accessing molecular-resolution details such as quantitatively monitoring components entering and leaving cell contacts by lateral diffusion and reversible association has revealed an unexpected plasticity. Adhesion structures can be highly specialized, such as focal adhesion in motile cells, as well as immune and neuronal synapses. Spatiotemporal reorganization of adhesion molecules, receptors, and adaptors directly relates to structure/function modulation. Assembly of these supramolecular complexes is continuously balanced by dynamic events, remodeling adhesions on various timescales, notably by molecular conformation switches, lateral diffusion within the membrane and endo/exocytosis. Pathological alterations in cell adhesion are involved in cancer evolution, through cancer stem cell interaction with stromal niches, growth, extravasation, and metastasis.
- Published
- 2016
- Full Text
- View/download PDF
32. Mapping molecular diffusion in the plasma membrane by Multiple-Target Tracing (MTT).
- Author
-
Rouger V, Bertaux N, Trombik T, Mailfert S, Billaudeau C, Marguet D, and Sergé A
- Subjects
- Algorithms, Animals, COS Cells, Cell Membrane metabolism, Chlorocebus aethiops, Diffusion, Microscopy, Video methods, Monte Carlo Method, Receptors, Cell Surface chemistry, Receptors, Cell Surface metabolism, Cell Membrane chemistry, Quantum Dots
- Abstract
Our goal is to obtain a comprehensive description of molecular processes occurring at cellular membranes in different biological functions. We aim at characterizing the complex organization and dynamics of the plasma membrane at single-molecule level, by developing analytic tools dedicated to Single-Particle Tracking (SPT) at high density: Multiple-Target Tracing (MTT). Single-molecule videomicroscopy, offering millisecond and nanometric resolution, allows a detailed representation of membrane organization by accurately mapping descriptors such as cell receptors localization, mobility, confinement or interactions. We revisited SPT, both experimentally and algorithmically. Experimental aspects included optimizing setup and cell labeling, with a particular emphasis on reaching the highest possible labeling density, in order to provide a dynamic snapshot of molecular dynamics as it occurs within the membrane. Algorithmic issues concerned each step used for rebuilding trajectories: peaks detection, estimation and reconnection, addressed by specific tools from image analysis. Implementing deflation after detection allows rescuing peaks initially hidden by neighboring, stronger peaks. Of note, improving detection directly impacts reconnection, by reducing gaps within trajectories. Performances have been evaluated using Monte-Carlo simulations for various labeling density and noise values, which typically represent the two major limitations for parallel measurements at high spatiotemporal resolution. The nanometric accuracy obtained for single molecules, using either successive on/off photoswitching or non-linear optics, can deliver exhaustive observations. This is the basis of nanoscopy methods such as STORM, PALM, RESOLFT or STED, which may often require imaging fixed samples. The central task is the detection and estimation of diffraction-limited peaks emanating from single-molecules. Hence, providing adequate assumptions such as handling a constant positional accuracy instead of Brownian motion, MTT is straightforwardly suited for nanoscopic analyses. Furthermore, MTT can fundamentally be used at any scale: not only for molecules, but also for cells or animals, for instance. Hence, MTT is a powerful tracking algorithm that finds applications at molecular and cellular scales.
- Published
- 2012
- Full Text
- View/download PDF
33. Antigen recognition by autoreactive CD4⁺ thymocytes drives homeostasis of the thymic medulla.
- Author
-
Irla M, Guerri L, Guenot J, Sergé A, Lantz O, Liston A, Imhof BA, Palmer E, and Reith W
- Subjects
- Animals, Body Patterning, CD40 Antigens metabolism, CD40 Ligand metabolism, Cell Proliferation, Epithelial Cells metabolism, Epithelial Cells physiology, Female, Gene Expression, Lymphotoxin beta Receptor metabolism, Lymphotoxin-alpha genetics, Lymphotoxin-alpha metabolism, Male, Mice, Mice, Inbred C57BL, Mice, Knockout, Receptor Activator of Nuclear Factor-kappa B metabolism, Signal Transduction, Thymocytes immunology, Thymocytes metabolism, Thymus Gland cytology, Thymus Gland immunology, Tissue Culture Techniques, Autoantigens immunology, CD4 Antigens metabolism, Homeostasis, Thymocytes physiology, Thymus Gland growth & development
- Abstract
The thymic medulla is dedicated for purging the T-cell receptor (TCR) repertoire of self-reactive specificities. Medullary thymic epithelial cells (mTECs) play a pivotal role in this process because they express numerous peripheral tissue-restricted self-antigens. Although it is well known that medulla formation depends on the development of single-positive (SP) thymocytes, the mechanisms underlying this requirement are incompletely understood. We demonstrate here that conventional SP CD4⁺ thymocytes bearing autoreactive TCRs drive a homeostatic process that fine-tunes medullary plasticity in adult mice by governing the expansion and patterning of the medulla. This process exhibits strict dependence on TCR-reactivity with self-antigens expressed by mTECs, as well as engagement of the CD28-CD80/CD86 costimulatory axis. These interactions induce the expression of lymphotoxin α in autoreactive CD4⁺ thymocytes and RANK in mTECs. Lymphotoxin in turn drives mTEC development in synergy with RANKL and CD40L. Our results show that Ag-dependent interactions between autoreactive CD4⁺ thymocytes and mTECs fine-tune homeostasis of the medulla by completing the signaling axes implicated in mTEC expansion and medullary organization.
- Published
- 2012
- Full Text
- View/download PDF
34. A spatially restricted increase in receptor mobility is involved in directional sensing during Dictyostelium discoideum chemotaxis.
- Author
-
de Keijzer S, Sergé A, van Hemert F, Lommerse PH, Lamers GE, Spaink HP, Schmidt T, and Snaar-Jagalska BE
- Subjects
- Animals, Dictyostelium cytology, Dimerization, GTP-Binding Protein alpha Subunits metabolism, GTP-Binding Protein beta Subunits metabolism, GTP-Binding Protein gamma Subunits metabolism, Recombinant Fusion Proteins metabolism, Chemotactic Factors metabolism, Chemotaxis physiology, Dictyostelium physiology, Protozoan Proteins metabolism, Receptors, Cyclic AMP metabolism
- Abstract
The directed cell migration towards a chemotactic source, chemotaxis, involves three complex and interrelated processes: directional sensing, cell polarization and motility. Directional sensing allows migrating eukaryotic cells to chemotax in extremely shallow gradients (<2% across the cell body) of the chemoattractant. Although directional sensing has been observed as spatially restricted responses along the plasma membrane, our understanding of the ;compass' of the cell that controls the gradient-induced translocation of proteins during chemotactic movements is still largely lacking. Until now, the dynamical behaviour and mobility of the chemoattractant-receptor molecule has been neglected in models describing the directional sensing mechanisms. Here, we show by single-molecule microscopy an agonist-induced increase in the mobile fraction of cAMP-receptor at the leading edge of chemotacting Dictyostelium discoideum cells. The onset of receptor mobility was correlated to the uncoupling and activation of the Galpha2-protein. A finite-element simulation showed that the increase in mobile fraction of the activated receptor enabled the amplified generation of activated Gbetagamma-dimers at the leading edge of the cell, faithfully representing a primary linear amplification step in directional sensing. We propose here that modulation of the receptor mobility is directly involved in directional sensing and provides a new mechanistic basis for the primary amplification step in current theoretical models that describe directional sensing.
- Published
- 2008
- Full Text
- View/download PDF
35. Receptor activation and homer differentially control the lateral mobility of metabotropic glutamate receptor 5 in the neuronal membrane.
- Author
-
Sergé A, Fourgeaud L, Hémar A, and Choquet D
- Subjects
- Animals, Carrier Proteins genetics, Cell Line, Cell Membrane drug effects, Cells, Cultured, Diffusion drug effects, Epithelial Cells cytology, Epithelial Cells drug effects, Epithelial Cells metabolism, Excitatory Amino Acid Agonists pharmacology, Green Fluorescent Proteins, Homer Scaffolding Proteins, Immunohistochemistry, Luminescent Proteins genetics, Microscopy, Video, Microspheres, Neurons cytology, Neurons drug effects, Neuropeptides genetics, Protein Binding physiology, Protein Isoforms genetics, Protein Isoforms metabolism, Protein Transport drug effects, Proto-Oncogene Proteins c-myc genetics, Rats, Receptor Aggregation drug effects, Receptor Aggregation physiology, Receptor, Metabotropic Glutamate 5, Receptors, Metabotropic Glutamate genetics, Recombinant Fusion Proteins genetics, Recombinant Fusion Proteins metabolism, Transfection, Carrier Proteins metabolism, Cell Membrane metabolism, Neurons metabolism, Neuropeptides metabolism, Protein Transport physiology, Receptors, Metabotropic Glutamate metabolism
- Abstract
Glutamate receptors are clustered at the membrane through interactions with intracellular scaffolding proteins and cytoskeletal elements but can also be found in intracellular compartments or dispersed in the membrane. This distribution results from an equilibrium between the different pools of receptors whose dynamic is poorly known. The group I metabotropic glutamate receptor 5 (mGluR5) is concentrated in an annulus around the postsynaptic density but also found in large amounts in the extrasynaptic membrane. To analyze the dynamic of stabilization of mGluR5, we used single-particle tracking, force measurements, and fluorescence recovery to measure the mobility of mGluR5. We found that receptor activation increases receptor diffusion, whereas the scaffolding protein Homer favors confinement of receptor movements within clusters of Homer-mGluR5. However, this stabilization is reversible, because even in the presence of Homer, receptors still enter and exit from clusters at fast rates. Furthermore, clusters themselves are highly dynamic both in their movements and in their composition, which can vary within tens of seconds. Thus, exchange of receptors between dispersed and clustered states is fast and regulated during physiological processes. These properties may explain certain fast changes in receptor composition observed at postsynaptic densities.
- Published
- 2002
- Full Text
- View/download PDF
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