Search

Your search keyword '"Henri-François Renard"' showing total 21 results
Start Over Author "Henri-François Renard"
21 results on '"Henri-François Renard"'

1. Endophilin-A3 and Galectin-8 control the clathrin-independent endocytosis of CD166

Author

Henri-François Renard, François Tyckaert, Cristina Lo Giudice, Thibault Hirsch, Cesar Augusto Valades-Cruz, Camille Lemaigre, Massiullah Shafaq-Zadah, Christian Wunder, Ruddy Wattiez, Ludger Johannes, Pierre van der Bruggen, David Alsteens, and Pierre Morsomme

Subjects
Science
Abstract

How and which cell surface molecules are taken up by clathrin-independent endocytosis is an ongoing area of research. Here, the authors show that the tumor marker CD166 is a clathrin-independent cargo that is taken up by endophilin-A3 and galectin-8, which regulates cancer cell migration.

Published
2020
Full Text
View/download PDF

2. Yeast α-arrestin Art2 is the key regulator of ubiquitylation-dependent endocytosis of plasma membrane vitamin B1 transporters.

Author

Jérôme Savocco, Sylvain Nootens, Wilhelmine Afokpa, Mathilde Bausart, Xiaoqian Chen, Jennifer Villers, Henri-François Renard, Martine Prévost, Ruddy Wattiez, and Pierre Morsomme

Subjects
Biology (General), QH301-705.5
Abstract

Endocytosis of membrane proteins in yeast requires α-arrestin-mediated ubiquitylation by the ubiquitin ligase Rsp5. Yet, the diversity of α-arrestin targets studied is restricted to a small subset of plasma membrane (PM) proteins. Here, we performed quantitative proteomics to identify new targets of 12 α-arrestins and gained insight into the diversity of pathways affected by α-arrestins, including the cell wall integrity pathway and PM-endoplasmic reticulum contact sites. We found that Art2 is the main regulator of substrate- and stress-induced ubiquitylation and endocytosis of the thiamine (vitamin B1) transporters: Thi7, nicotinamide riboside transporter 1 (Nrt1), and Thi72. Genetic screening allowed for the isolation of transport-defective Thi7 mutants, which impaired thiamine-induced endocytosis. Coexpression of inactive mutants with wild-type Thi7 revealed that both transporter conformation and transport activity are important to induce endocytosis. Finally, we provide evidence that Art2 mediated Thi7 endocytosis is regulated by the target of rapamycin complex 1 (TORC1) and requires the Sit4 phosphatase but is not inhibited by the Npr1 kinase.

Published
2019
Full Text
View/download PDF

3. N‐BAR and F‐BAR proteins—endophilin‐A3 and PSTPIP1—control clathrin‐independent endocytosis of L1CAM

Author

Camille Lemaigre, Apolline Ceuppens, Cesar Augusto Valades‐Cruz, Benjamin Ledoux, Bastien Vanbeneden, Mujtaba Hassan, Fredrik R. Zetterberg, Ulf J. Nilsson, Ludger Johannes, Christian Wunder, Henri‐François Renard, Pierre Morsomme, and UCL - SST/LIBST - Louvain Institute of Biomolecular Science and Technology

Subjects
Structural Biology, Genetics, Cell Biology, Molecular Biology, Biochemistry
Abstract

Recent advances in the field demonstrate the high diversity and complexity of endocytic pathways. In the current study, we focus on the endocytosis of L1CAM. This glycoprotein plays a major role in the development of the nervous system, and is involved in cancer development and is associated with metastases and poor prognosis. Two L1CAM isoforms are subject to endocytosis: isoform 1, described as a clathrinmediated cargo; isoform 2, whose endocytosis has never been studied. Deciphering the molecular machinery of isoform 2 internalisation should contribute to a better understanding of its pathophysiological role. First, we demonstrated in our cellular context that both isoforms of L1CAM are mainly a clathrin-independent cargo, which was not expected for isoform 1. Second, the mechanism of L1CAM endocytosis is specifically mediated by the N-BAR domain protein endophilin-A3. Third, we discovered PSTPIP1, an F-BAR domain protein, as a novel actor in this endocytic process. Finally, we identified galectins as endocytic partners and negative regulators of L1CAM endocytosis. In summary, the interplay of the BAR proteins endophilin-A3 and PSTPIP1, and galectins fine tune the clathrin-independent endocytosis of L1CAM.

Published
2023

4. Host cell egress of Brucella abortus requires <scp>BNIP3L</scp> ‐mediated mitophagy

Author

Jérémy Verbeke, Youri Fayt, Lisa Martin, Oya Yilmaz, Jaroslaw Sedzicki, Angéline Reboul, Michel Jadot, Patricia Renard, Christoph Dehio, Henri‐François Renard, Jean‐Jacques Letesson, Xavier De Bolle, and Thierry Arnould

Subjects
BNIP3L, iron, mitophagy, General Immunology and Microbiology, intracellular trafficking, General Neuroscience, Brucella, Molecular Biology, General Biochemistry, Genetics and Molecular Biology
Abstract

The facultative intracellular pathogen Brucella abortus interacts with several organelles of the host cell to reach its replicative niche inside the endoplasmic reticulum. However, little is known about the interplay between the intracellular bacteria and the host cell mitochondria. Here, we showed that B. abortus triggers substantive mitochondrial network fragmentation, accompanied by mitophagy and the formation of mitochondrial Brucella-containing vacuoles during the late steps of cellular infection. Brucella-induced expression of the mitophagy receptor BNIP3L is essential for these events and relies on the iron-dependent stabilisation of the hypoxia-inducible factor 1α. Functionally, BNIP3L-mediated mitophagy appears to be advantageous for bacterial exit from the host cell as BNIP3L depletion drastically reduces the number of reinfection events. Altogether, these findings highlight the intricate link between Brucella trafficking and the mitochondria during host cell infection.

Published
2023

5. Unconventional endocytic mechanisms

Author

Henri-François Renard and Emmanuel Boucrot

Subjects
0303 health sciences, Pinocytosis, Endocytic cycle, Cell Biology, Receptor-mediated endocytosis, Biology, Endocytosis, Clathrin, Transmembrane protein, Bulk endocytosis, Cell biology, 03 medical and health sciences, Cytosol, 0302 clinical medicine, biology.protein, 030217 neurology & neurosurgery, 030304 developmental biology
Abstract

Endocytosis mediates the uptake of extracellular proteins, micronutrients and transmembrane cell surface proteins. Importantly, many viruses, toxins and bacteria hijack endocytosis to infect cells. The canonical pathway is clathrin-mediated endocytosis (CME) and is active in all eukaryotic cells to support critical house-keeping functions. Unconventional mechanisms of endocytosis exit in parallel of CME, to internalize specific cargoes and support various cellular functions. These clathrin-independent endocytic (CIE) routes use three distinct mechanisms: acute signaling-induced membrane remodeling drives macropinocytosis, activity-dependent bulk endocytosis (ADBE), massive endocytosis (MEND) and EGFR non-clathrin endocytosis (EGFR-NCE). Cargo capture and local membrane deformation by cytosolic proteins is used by fast endophilin-mediated endocytosis (FEME), IL-2Rβ endocytosis and ultrafast endocytosis at synapses. Finally, the formation of endocytic pits by clustering of extracellular lipids or cargoes according to the Glycolipid-Lectin (GL-Lect) hypothesis mediates the uptake of SV40 virus, Shiga and cholera toxins, and galectin-clustered receptors by the CLIC/GEEC and the endophilin-A3-mediated CIE.

Published
2021

6. BNIP3L-mediated mitophagy triggered by Brucella in host cells is required for bacterial egress

Author

Jérémy Verbeke, Youri Fayt, Lisa Martin, Oya Yilmaz, Jaroslaw Sedzicki, Angeline Reboul, Michel Jadot, Patricia Renard, Christoph Dehio, Henri-François Renard, Jean-Jacques Letesson, Xavier De Bolle, and Thierry Arnould

Abstract

The facultative intracellular pathogen Brucella abortus interacts with several organelles of the host cell to reach its replicative niche inside the endoplasmic reticulum. However, little is known about the interplay between the bacteria and the host cell mitochondria. Here, we showed that B. abortus triggers a strong mitochondrial network fragmentation accompanied by mitophagy and the formation of mitochondrial Brucella-containing vacuoles in the late steps of cellular infection. The expression of the mitophagy receptor BNIP3L induced by B. abortus is essential for these events and relies on the iron-dependent stabilization of the hypoxia-inducible factor 1 alpha. Functionally, BNIP3L-mediated mitophagy appears to be advantageous for bacterial exit of the host cell as BNIP3L depletion drastically reduced the number of reinfection events. Altogether, these findings highlight the intricate link between Brucella trafficking and the mitochondria during host cell infection.

Published
2022

8. Rac1, actin cytoskeleton and microtubules are key players in clathrin-independent endophilin-A3-mediated endocytosis

Author

François Tyckaert, Natacha Zanin, Pierre Morsomme, Henri-François Renard, and UCL - SST/LIBST - Louvain Institute of Biomolecular Science and Technology

Subjects
rac1 GTP-Binding Protein, Actin Cytoskeleton, CD166, endocytosis, actin, microtubule, endophilin, Humans, Microtubule, Cell Biology, Endophilin, Microtubules, Actins, Clathrin, Actin, Endocytosis
Abstract

Endocytic mechanisms actively regulate plasma membrane composition and sustain fundamental cellular functions. Recently, we identified a clathrin-independent endocytic (CIE) modality mediated by the BAR domain protein endophilin-A3 (endoA3, encoded by SH3GL3), which controls the cell surface homeostasis of the tumor marker CD166 (also known as ALCAM). Deciphering the molecular machinery of endoA3-dependent CIE should therefore contribute to a better understanding of its pathophysiological role, which remains so far unknown. Here, we investigate the role of actin, Rho GTPases and microtubules, which are major players in CIE processes, in this mechanism. We show that the actin cytoskeleton is dynamically associated with endoA3- and CD166-positive endocytic carriers, and that its perturbation strongly inhibits the process of CD166 uptake. We also reveal that the Rho GTPase Rac1, but not Cdc42, is a master regulator of this endocytic route. Finally, we provide evidence that microtubules and kinesin molecular motors are required to potentiate endoA3-dependent endocytosis. Of note, our study also highlights potential compensation phenomena between endoA3-dependent CIE and macropinocytosis. Altogether, our data deepen our understanding of this CIE modality and further differentiate it from other unconventional endocytic mechanisms. This article has an associated First Person interview with the first author of the paper.

Published
2022

10. Endophilin-A3 and Galectin-8 control the clathrin-independent endocytosis of CD166

Author

Pierre van der Bruggen, Camille Lemaigre, Cesar Augusto Valades-Cruz, Henri-François Renard, François Tyckaert, Ludger Johannes, Ruddy Wattiez, Cristina Lo Giudice, Pierre Morsomme, David Alsteens, Massiullah Shafaq-Zadah, Christian Wunder, Thibault Hirsch, UCL - SST/LIBST - Louvain Institute of Biomolecular Science and Technology, Bodescot, Myriam, Assimilation de Données et Microscopie à Feuille de Lumière Structurée pour la Modélisation des Voies d'Endocytose et d'Exocytose en Cellule Unique - - DALLISH2016 - ANR-16-CE23-0005 - AAPG2016 - VALID, Développment d'une infrastructure française distribuée coordonnée - - France-BioImaging2010 - ANR-10-INBS-0004 - INBS - VALID, Initiative d'excellence - Paris Sciences et Lettres - - PSL2010 - ANR-10-IDEX-0001 - IDEX - VALID, Endocytic Membrane Compartmentalization by Galectins - GALECTCOMPART - - EC:FP7:ERC2014-04-01 - 2019-03-31 - 340485 - VALID, Université Catholique de Louvain = Catholic University of Louvain (UCL), Space-timE RePresentation, Imaging and cellular dynamics of molecular COmplexes (SERPICO), Inria Rennes – Bretagne Atlantique, Institut National de Recherche en Informatique et en Automatique (Inria)-Institut National de Recherche en Informatique et en Automatique (Inria), Biologie Cellulaire et Cancer, Institut Curie [Paris]-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS), Chimie biologique des membranes et ciblage thérapeutique (CBMCT - UMR 3666 / U1143), Institut Curie [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Université de Mons (UMons), Walloon Excellence in Life sciences and BIOtechnology [Liège] (WELBIO), This work was supported by grants from the 'Fonds National de la Recherche Scientifique' (FNRS, CDR-J.0119.19) and the 'Communauté française de Belgique–Actions de Recherches Concertées' (17/22-085). This work was also supported by the French National Research Agency (DALLISH–ANR-16-CE23-0005), and by Inria in the frame of NAVISCOPE-IPL (Inria Project Lab). The bioprofiling platform used for the proteomic analysis was supported by the FNRS, the European Regional Development Fund, and the Walloon Region, Belgium. The LLSM was financed by PIA France-Bioimaging (ANR-10-INBS-04_01), LabEx DCBiol, LabEx CelTisPhyBio ANR-11-LABX-0038, Agence Nationale de la Recherche (ANR-16-CE23-0005-02, ANR-19-CE13-0001-01), HFSP (RGP0029/2014), and European Research Council (ERC project 340485). We greatly acknowledge the Cell and Tissue Imaging Facility (PICT-IBiSA) and Nikon Imaging Centre, Institut Curie, member of the French National Research Infrastructure France-BioImaging (ANR-10-INBS-04). H.-F.R. is a FNRS postdoctoral research fellow (Belgium). F.T., T.H. and C.L. are supported by PhD fellowships from FRIA/FNRS (Belgium). C.L.G. is an EMBO Long-term postdoctoral fellow. P.V.D.B. and D.A. are supported by WELBIO (Fédération Wallonie-Bruxelles, Belgium). This work was supported by the European Research Council under the European Union’s Horizon 2020 research and innovation program (grant agreement no. 758224). D.A. is a research associate of the FNRS (Belgium)., ANR-16-CE23-0005,DALLISH,Assimilation de Données et Microscopie à Feuille de Lumière Structurée pour la Modélisation des Voies d'Endocytose et d'Exocytose en Cellule Unique(2016), ANR-10-INBS-0004,France-BioImaging,Développment d'une infrastructure française distribuée coordonnée(2010), ANR-10-IDEX-0001,PSL,Paris Sciences et Lettres(2010), European Project: 340485,EC:FP7:ERC,ERC-2013-ADG,GALECTCOMPART(2014), UCL - SSS/DDUV - Institut de Duve, and Centre National de la Recherche Scientifique (CNRS)-Institut Curie [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut de Chimie du CNRS (INC)

Subjects
Fetal Proteins, 0301 basic medicine, Intravital Microscopy, Endocytic cycle, General Physics and Astronomy, Mice, 0302 clinical medicine, Cell Movement, Neoplasms, Chlorocebus aethiops, RNA, Small Interfering, lcsh:Science, Multidisciplinary, Cell adhesion molecule, Chemistry, Activated-Leukocyte Cell Adhesion Molecule, Signal transducing adaptor protein, Endocytosis, Recombinant Proteins, 3. Good health, Cell biology, Galectin-8, Gene Knockdown Techniques, 030220 oncology & carcinogenesis, Science, Cell Adhesion Molecules, Neuronal, Galectins, [SDV.CAN]Life Sciences [q-bio]/Cancer, Genetics and Molecular Biology, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, Article, General Biochemistry, Genetics and Molecular Biology, 03 medical and health sciences, [SDV.CAN] Life Sciences [q-bio]/Cancer, Antigens, CD, Cell Line, Tumor, Cell Adhesion, Animals, Humans, Cell adhesion, [SDV.BC] Life Sciences [q-bio]/Cellular Biology, ALCAM, Adaptor Proteins, Signal Transducing, Cell Membrane, General Chemistry, Fibroblasts, Clathrin, 030104 developmental biology, General Biochemistry, lcsh:Q
Abstract

While several clathrin-independent endocytic processes have been described so far, their biological relevance often remains elusive, especially in pathophysiological contexts such as cancer. In this study, we find that the tumor marker CD166/ALCAM (Activated Leukocyte Cell Adhesion Molecule) is a clathrin-independent cargo. We show that endophilin-A3—but neither A1 nor A2 isoforms—functionally associates with CD166-containing early endocytic carriers and physically interacts with the cargo. Our data further demonstrates that the three endophilin-A isoforms control the uptake of distinct subsets of cargoes. In addition, we provide strong evidence that the construction of endocytic sites from which CD166 is taken up in an endophilin-A3-dependent manner is driven by extracellular galectin-8. Taken together, our data reveal the existence of a previously uncharacterized clathrin-independent endocytic modality, that modulates the abundance of CD166 at the cell surface, and regulates adhesive and migratory properties of cancer cells., How and which cell surface molecules are taken up by clathrin-independent endocytosis is an ongoing area of research. Here, the authors show that the tumor marker CD166 is a clathrin-independent cargo that is taken up by endophilin-A3 and galectin-8, which regulates cancer cell migration.

Published
2020

11. Functional dissection of the retrograde Shiga toxin trafficking inhibitor Retro-2

Author

Raphaël Sierocki, Julien Barbier, Adam D. Linstedt, Ludger Johannes, Alison Forrester, Stefan J. Rathjen, Mathilde Munier, Sylvain Pichard, Daniel Gillet, Henri-François Renard, Christophe Lamaze, Damarys Loew, Maria Daniela Garcia-Castillo, Jennifer Martinez, Livia Tepshi, Jean-Christophe Cintrat, Collin Bachert, Audrey Couhert, Florent Dingli, Cesar Augusto Valades-Cruz, Chimie biologique des membranes et ciblage thérapeutique (CBMCT - UMR 3666 / U1143), Institut Curie [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Service d'Ingénierie Moléculaire pour la Santé (ex SIMOPRO) (SIMoS), Médicaments et Technologies pour la Santé (MTS), Université Paris-Saclay-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-Université Paris-Saclay-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), ANR-19-CE13-0001,GlycoTopoSwitch,La topologie des glycans comme interrupteur moléculaire pour l'activité des intérgines(2019), Carnegie Mellon University [Pittsburgh] (CMU), Service de Chimie Bio-Organique et de Marquage (SCBM), Centre de recherche de l'Institut Curie [Paris], Institut Curie [Paris], Human Frontier Science Program RGP0029-2014, Swedish Research Council European Commission K2015-99X-22877-01-6, Joint Ministerial Program of R&D against CBRNE Risks, Ile de France Région DIM Malinf initiative 140101, Région Ile-de-France, Fondation pour la Recherche Médicale, French Atomic Energy Commission, ANR-11-BSV2-0018,RETROscreen,Génétique chimique de la voie du transport rétrograde(2011), ANR-14-CE16-0004,AntiHUS,Preuve de concept in vivo de molécules contre le syndrome hémolytique et urémique(2014), European Project: 340485,EC:FP7:ERC,ERC-2013-ADG,GALECTCOMPART(2014), Centre National de la Recherche Scientifique (CNRS)-Institut Curie [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut de Chimie du CNRS (INC), Serva, Camille, BLANC - Génétique chimique de la voie du transport rétrograde - - RETROscreen2011 - ANR-11-BSV2-0018 - BLANC - VALID, Appel à projets générique - Preuve de concept in vivo de molécules contre le syndrome hémolytique et urémique - - AntiHUS2014 - ANR-14-CE16-0004 - Appel à projets générique - VALID, La topologie des glycans comme interrupteur moléculaire pour l'activité des intérgines - - GlycoTopoSwitch2019 - ANR-19-CE13-0001 - AAPG2019 - VALID, and Endocytic Membrane Compartmentalization by Galectins - GALECTCOMPART - - EC:FP7:ERC2014-04-01 - 2019-03-31 - 340485 - VALID

Subjects
Retrograde transport, [SDV]Life Sciences [q-bio], Vesicular Transport Proteins, Golgi Apparatus, Endoplasmic Reticulum, Shiga Toxins, chemistry.chemical_compound, anterograde trafficking, COPII, mass spectrometry, 0303 health sciences, biology, Qa-SNARE Proteins, Chemistry, trans-Golgi network, 030302 biochemistry & molecular biology, Shiga-like toxin, Shiga toxin, Transport protein, Cell biology, Protein Transport, chemical genetics, Ricin, SNARE, Benzamides, click chemistry, symbols, Retro-2, GPP130, biological phenomena, cell phenomena, and immunity, retention using selective hooks, syntaxin-5, endocrine system, Sec16A, Endosome, small molecule, chemical biology, Endosomes, Thiophenes, STxB, 03 medical and health sciences, symbols.namesake, [SDV.BBM] Life Sciences [q-bio]/Biochemistry, Molecular Biology, Humans, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, proximity ligation assay, Molecular Biology, 030304 developmental biology, Endoplasmic reticulum, Biological Transport, Cell Biology, Golgi apparatus, nervous system, Chaperone (protein), Axoplasmic transport, biology.protein, HeLa Cells
Abstract

International audience; The retrograde transport inhibitor Retro-2 has a protective effect on cells and in mice against Shiga-like toxins and ricin. Retro-2 causes toxin accumulation in early endosomes and relocalization of the Golgi SNARE protein syntaxin-5 to the endoplasmic reticulum. The molecular mechanisms by which this is achieved remain unknown. Here, we show that Retro-2 targets the endoplasmic reticulum exit site component Sec16A, affecting anterograde transport of syntaxin-5 from the endoplasmic reticulum to the Golgi. The formation of canonical SNARE complexes involving syntaxin-5 is not affected in Retro-2-treated cells. By contrast, the interaction of syntaxin-5 with a newly discovered binding partner, the retrograde trafficking chaperone GPP130, is abolished, and we show that GPP130 must indeed bind to syntaxin-5 to drive Shiga toxin transport from the endosomes to the Golgi. We therefore identify Sec16A as a druggable target and provide evidence for a non-SNARE function for syntaxin-5 in interaction with GPP130.

Published
2020

12. Yeast α-arrestin Art2 is the key regulator of ubiquitylation-dependent endocytosis of plasma membrane vitamin B1 transporters

Author

Henri-François Renard, Xiaoqian Chen, Jérôme Savocco, Ruddy Wattiez, Jennifer Villers, Sylvain Nootens, Mathilde Bausart, Martine Prévost, Wilhelmine Afokpa, Pierre Morsomme, and UCL - SST/LIBST - Louvain Institute of Biomolecular Science and Technology

Subjects
Models, Molecular, Proteomics, B Vitamins, 0301 basic medicine, Arrestins, Cell Membranes, Plasma protein binding, Endoplasmic Reticulum, Biochemistry, Protein Structure, Secondary, chemistry.chemical_compound, 0302 clinical medicine, Ubiquitin, Cell Wall, Gene Expression Regulation, Fungal, Immunologie, Protein Phosphatase 2, Thiamine, Post-Translational Modification, Phosphorylation, Biology (General), Secretory Pathway, biology, Organic Compounds, General Neuroscience, Ubiquitin-Protein Ligase Complexes, Vitamins, Sciences bio-médicales et agricoles, Endocytosis, Enzymes, Ubiquitin ligase, Cell biology, Chemistry, Phenotypes, Cell Processes, Physical Sciences, Cellular Structures and Organelles, Signal transduction, General Agricultural and Biological Sciences, Biologie, Protein Binding, Signal Transduction, Research Article, Saccharomyces cerevisiae Proteins, QH301-705.5, Nucleoside Transport Proteins, Saccharomyces cerevisiae, Green Fluorescent Protein, General Biochemistry, Genetics and Molecular Biology, 03 medical and health sciences, Genetics, Arrestin, Endosomal Sorting Complexes Required for Transport, General Immunology and Microbiology, Cell Membrane, Organic Chemistry, Ubiquitination, Chemical Compounds, Phosphatases, Neurosciences cognitives, Membrane Transport Proteins, Biology and Life Sciences, Proteins, Membrane Proteins, Cell Biology, Luminescent Proteins, 030104 developmental biology, Membrane protein, chemistry, Mutation, Nicotinamide riboside, Enzymology, biology.protein, Microbiologie et protistologie [bacteriol.virolog.mycolog.], Protein Kinases, Protein Processing, Post-Translational, 030217 neurology & neurosurgery, Transcription Factors
Abstract

Endocytosis of membrane proteins in yeast requires α-arrestin-mediated ubiquitylation by the ubiquitin ligase Rsp5. Yet, the diversity of α-arrestin targets studied is restricted to a small subset of plasma membrane (PM) proteins. Here, we performed quantitative proteomics to identify new targets of 12 α-arrestins and gained insight into the diversity of pathways affected by α-arrestins, including the cell wall integrity pathway and PM–endoplasmic reticulum contact sites. We found that Art2 is the main regulator of substrate- and stress-induced ubiquitylation and endocytosis of the thiamine (vitamin B1) transporters: Thi7, nicotinamide riboside transporter 1 (Nrt1), and Thi72. Genetic screening allowed for the isolation of transport-defective Thi7 mutants, which impaired thiamine-induced endocytosis. Coexpression of inactive mutants with wild-type Thi7 revealed that both transporter conformation and transport activity are important to induce endocytosis. Finally, we provide evidence that Art2 mediated Thi7 endocytosis is regulated by the target of rapamycin complex 1 (TORC1) and requires the Sit4 phosphatase but is not inhibited by the Npr1 kinase., SCOPUS: ar.j, info:eu-repo/semantics/published

Published
2019

13. Increasing Diversity of Biological Membrane Fission Mechanisms

Author

Henri-François Renard, Ludger Johannes, Pierre Morsomme, and UCL - SST/ISV - Institut des sciences de la vie

Subjects
0301 basic medicine, Dynamins, Fission, Biology, Endocytosis, Models, Biological, Cell membrane, 03 medical and health sciences, 0302 clinical medicine, Membrane fission, medicine, Animals, Humans, fission, Dynamin, Cytokinesis, scission, Endosomal Sorting Complexes Required for Transport, Mechanism (biology), Cell Membrane, Biological membrane, Cell Biology, Cell biology, 030104 developmental biology, medicine.anatomical_structure, membranes, Cell Nucleus Division, 030217 neurology & neurosurgery
Abstract

Membrane fission is essential to life. It is required for many fundamental cellular processes, as diverse as cyto- and karyokinesis, organelle division, membrane repair, and membrane trafficking and endocytosis. While membrane fission was originally seen as resulting from the action of mechanoenzymes such as dynamin, it is clear that the reality is more complex. In this review, we propose an updated overview of fission mechanisms, and try to extract essential requirements for each. We also present examples of cellular processes that involve these fission mechanisms. Finally, we list pending questions, whether they are specific to a peculiar fission mechanism or more general to the field.

Published
2018

14. Endophilin-A2 functions in membrane scission in clathrin-independent endocytosis

Author

Anne K. Kenworthy, Mijo Simunovic, Anne A. Schmidt, Valérie Chambon, Patricia Bassereau, Henri-François Renard, Christian Wunder, Cécile Sykes, Joël Lemière, Emmanuel Boucrot, Christophe Lamaze, Harvey T. McMahon, Maria Daniela Garcia-Castillo, Senthil Arumugam, Ludger Johannes, Bondidier, Martine, Chimie biologique des membranes et ciblage thérapeutique ( CBMCT - UMR 3666 / U1143 ), Université Paris Descartes - Paris 5 ( UPD5 ) -Institut Curie-Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS ), Physico-Chimie-Curie ( PCC ), Centre National de la Recherche Scientifique ( CNRS ) -INSTITUT CURIE-Université Pierre et Marie Curie - Paris 6 ( UPMC ), Department of Chemistry, University of Chicago, Université Paris Diderot - Paris 7 ( UPD7 ), Institute of Structural and Molecular Biology, Birkbeck College, Department of Molecular Physiology and Biophysics, Vanderbilt University School of Medicine. Nashville, Vanderbilt University School of Medicine. Nashville-Vanderbilt University School of Medicine. Nashville, Institut Jacques Monod ( IJM ), Université Paris Diderot - Paris 7 ( UPD7 ) -Centre National de la Recherche Scientifique ( CNRS ), Laboratory of Molecular Biology, Division of Cell Biology, Medical Research Council Laboratory of Molecular Biology, Cambridge CB2 0QH, UK, Agence Nationale pour la Recherche : (ANR-09-BLAN-283, ANR-10-LBX-0038, ANR-11 BSV2 014 03, ANR-12-BSV5-0014), Indo-French Centre for the Promotion of Advanced Science (project no. 3803), Marie Curie Actions — Networks for Initial Training (FP7-PEOPLE-2010-ITN), Marie Curie International Reintegration Grant (FP7-RG-277078), European Research Council advanced grant (project 340485), Royal Society (RG120481), Fondation ARC pour la Recherche sur le Cancer (DEQ20120323737), National Institutes of Health (RO1 GM106720), Ligue contre le Cancer, Comité de Paris (RS08/75-89), Fondation ARC pour la Recherche sur le Cancer, AXA Research Funds, Biological Sciences Research Council, Chateaubriand fellowship, France and Chicago Collaborating in the Sciences grant, Chimie biologique des membranes et ciblage thérapeutique (CBMCT - UMR 3666 / U1143), Université Paris Descartes - Paris 5 (UPD5)-Institut Curie [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Physico-Chimie-Curie (PCC), Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut Curie [Paris]-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Université Paris Diderot - Paris 7 (UPD7), Vanderbilt University School of Medicine [Nashville], Institut Jacques Monod (IJM (UMR_7592)), Université Paris Diderot - Paris 7 (UPD7)-Centre National de la Recherche Scientifique (CNRS), Université Paris Descartes - Paris 5 (UPD5)-Institut Curie [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie du CNRS (INC), and Centre National de la Recherche Scientifique (CNRS)-Institut Curie [Paris]-Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut de Chimie du CNRS (INC)

Subjects
MESH : Cell Line, MESH: Rats, MESH : Endocytosis, MESH : Cell Membrane, MESH: Shiga Toxin, Endocytic cycle, MESH : Actins, MESH : Dynamins, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, macromolecular substances, MESH: Acyltransferases, Biology, MESH: Actins, Endocytosis, environment and public health, Clathrin, Cell membrane, 03 medical and health sciences, 0302 clinical medicine, medicine, BAR domain, MESH: Animals, MESH: Clathrin, Endophilin-A2, [SDV.BC] Life Sciences [q-bio]/Cellular Biology, MESH: Cholera Toxin, 030304 developmental biology, Dynamin, 0303 health sciences, MESH: Humans, MESH : Clathrin, Multidisciplinary, MESH : Rats, MESH : Cholera Toxin, [ SDV.BC ] Life Sciences [q-bio]/Cellular Biology, MESH : Humans, MESH: Cell Line, Cell biology, MESH: Dynamins, medicine.anatomical_structure, MESH : Shiga Toxin, MESH: Endocytosis, Amphiphysin, biology.protein, MESH : Animals, MESH : Acyltransferases, 030217 neurology & neurosurgery, MESH: Cell Membrane
Abstract

International audience; During endocytosis, energy is invested to narrow the necks of cargo-containing plasma membrane invaginations to radii at which the opposing segments spontaneously coalesce, thereby leading to the detachment by scission of endocytic uptake carriers. In the clathrin pathway, dynamin uses mechanical energy from GTP hydrolysis to this effect, assisted by the BIN/amphiphysin/Rvs (BAR) domain-containing protein endophilin. Clathrin-independent endocytic events are often less reliant on dynamin, and whether in these cases BAR domain proteins such as endophilin contribute to scission has remained unexplored. Here we show, in human and other mammalian cell lines, that endophilin-A2 (endoA2) specifically and functionally associates with very early uptake structures that are induced by the bacterial Shiga and cholera toxins, which are both clathrin-independent endocytic cargoes. In controlled in vitro systems, endoA2 reshapes membranes before scission. Furthermore, we demonstrate that endoA2, dynamin and actin contribute in parallel to the scission of Shiga-toxin-induced tubules. Our results establish a novel function of endoA2 in clathrin-independent endocytosis. They document that distinct scission factors operate in an additive manner, and predict that specificity within a given uptake process arises from defined combinations of universal modules. Our findings highlight a previously unnoticed link between membrane scaffolding by endoA2 and pulling-force-driven dynamic scission.

Published
2014

15. Friction Mediates Scission of Tubular Membranes Scaffolded by BAR Proteins

Author

Andrew Callan-Jones, Patricia Bassereau, Jacques Prost, Krishnan Raghunathan, Dhiraj Bhatia, Jean-Baptiste Manneville, Henri-François Renard, Gregory A. Voth, Ludger Johannes, Emma Evergren, Mijo Simunovic, Harvey T. McMahon, Anne K. Kenworthy, James Franck Institute, University of Chicago, 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), Chimie biologique des membranes et ciblage thérapeutique (CBMCT - UMR 3666 / U1143), Université Paris Descartes - Paris 5 (UPD5)-Institut Curie [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie du CNRS (INC), Centre for Cancer Research and Cell Biology, Queen's University [Belfast] (QUB), Medical Research Council Laboratory of Molecular Biology, Cambridge, Vanderbilt University School of Medicine [Nashville], National University of Singapore (NUS), Matière et Systèmes Complexes (MSC (UMR_7057)), Centre National de la Recherche Scientifique (CNRS)-Université Paris Diderot - Paris 7 (UPD7), Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut Curie [Paris]-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut Curie [Paris]-Centre National de la Recherche Scientifique (CNRS), Université Paris Descartes - Paris 5 (UPD5)-Institut Curie [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Matière et Systèmes Complexes (MSC), Université Paris Diderot - Paris 7 (UPD7)-Centre National de la Recherche Scientifique (CNRS), ANR-16-CE23-0005,DALLISH,Assimilation de Données et Microscopie à Feuille de Lumière Structurée pour la Modélisation des Voies d'Endocytose et d'Exocytose en Cellule Unique(2016), Physico-Chimie-Curie ( PCC ), Centre National de la Recherche Scientifique ( CNRS ) -INSTITUT CURIE-Université Pierre et Marie Curie - Paris 6 ( UPMC ), Compartimentation et dynamique cellulaires ( CDC ), Chimie biologique des membranes et ciblage thérapeutique ( CBMCT - UMR 3666 / U1143 ), Université Paris Descartes - Paris 5 ( UPD5 ) -Institut Curie-Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS ), Queen's University [Belfast] ( QUB ), National University of Singapore ( NUS ), Matière et Systèmes Complexes ( MSC ), Université Paris Diderot - Paris 7 ( UPD7 ) -Centre National de la Recherche Scientifique ( CNRS ), and HAL UPMC, Gestionnaire

Subjects
0301 basic medicine, Scaffold protein, Lysis, Friction, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, Biology, Endocytosis, General Biochemistry, Genetics and Molecular Biology, 03 medical and health sciences, Protein Domains, Molecular motor, BAR domain, Animals, Humans, [SDV.BC] Life Sciences [q-bio]/Cellular Biology, Bond cleavage, [ SDV.BC ] Life Sciences [q-bio]/Cellular Biology, Membrane Proteins, Lipid Metabolism, Biomechanical Phenomena, Rats, 030104 developmental biology, Membrane, Biochemistry, Biophysics, Elongation, Acyltransferases
Abstract

International audience; Membrane scission is essential for intracellular trafficking. While BAR domain proteins such as endophilin have been reported in dynamin-independent scission of tubular membrane necks, the cutting mechanism has yet to be deciphered. Here, we combine a theoretical model, in vitro, and in vivo experiments revealing how protein scaffolds may cut tubular membranes. We demonstrate that the protein scaffold bound to the underlying tube creates a frictional barrier for lipid diffusion; tube elongation thus builds local membrane tension until the membrane undergoes scission through lysis. We call this mechanism friction-driven scission (FDS). In cells, motors pull tubes, particularly during endocytosis. Through reconstitution, we show that motors not only can pull out and extend protein-scaffolded tubes but also can cut them by FDS. FDS is generic, operating even in the absence of amphipathic helices in the BAR domain, and could in principle apply to any high-friction protein and membrane assembly.

Published
2017

16. Rab12 Localizes to Shiga Toxin-Induced Plasma Membrane Invaginations and Controls Toxin Transport

Author

Winfried Römer, Ludger Johannes, Henri-François Renard, Gustaf E. Rydell, Florent Dingli, Maria Daniela Garcia-Castillo, Damarys Loew, and Christophe Lamaze

Subjects
biology, Toxin, media_common.quotation_subject, Toxin transport, Shiga toxin, Cell Biology, Transfection, medicine.disease_cause, Biochemistry, Clathrin, Cell biology, Structural Biology, Stable isotope labeling by amino acids in cell culture, Genetics, biology.protein, medicine, Internalization, Receptor, Molecular Biology, media_common
Abstract

Several exogenous and endogenous cargo proteins are internalized independently of clathrin, including the bacterial Shiga toxin. The mechanisms underlying early steps of clathrin-independent uptake remain largely unknown. In this study, we have designed a protocol to obtain gradient fractions containing Shiga toxin internalization intermediates. Using stable isotope labeling with amino acids in cell culture (SILAC) and quantitative mass spectrometry, Rab12 was found in association with these very early uptake carriers. The localization of the GTPase on Shiga toxin-induced plasma membrane invaginations was shown by fluorescence microscopy in cells transfected with GFP-Rab12. Furthermore, using a quantitative biochemical assay, it was found that the amount of receptor-binding B-subunit of Shiga toxin reaching the trans-Golgi/TGN membranes was decreased in Rab12-depleted cells, and that cells were partially protected against intoxication by Shiga-like toxin 1 under these conditions. These findings demonstrate the functional importance of Rab12 for retrograde toxin trafficking. Among several other intracellular transport pathways, only the steady-state localizations of TGN46 and cation-independent mannose-6-phosphate receptor were affected. These data thus strongly suggest that Rab12 functions in the retrograde transport route.

Published
2014

17. How curvature-generating proteins build scaffolds on membrane nanotubes

Author

Patricia Bassereau, Gregory A. Voth, Mijo Simunovic, Ivan Golushko, Harvey T. McMahon, Vladimir Lorman, Henri-François Renard, Ludger Johannes, Emma Evergren, Coline Prévost, 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), Laboratoire Charles Coulomb (L2C), Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS), UCL - SST/ISV - Institut des sciences de la vie, and Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut Curie [Paris]-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)

Subjects
0301 basic medicine, Scaffold, Materials science, Surface Properties, Bar (music), [PHYS.PHYS.PHYS-BIO-PH]Physics [physics]/Physics [physics]/Biological Physics [physics.bio-ph], Nanotechnology, Molecular Dynamics Simulation, Endocytosis, Fluorescence, Protein Structure, Secondary, 03 medical and health sciences, Molecular dynamics, 0302 clinical medicine, Protein Domains, Fluorescence microscope, Computer Simulation, Adaptor Proteins, Signal Transducing, Binding Sites, Nanotubes, Multidisciplinary, X-Rays, Cell Membrane, Membrane Proteins, Membrane nanotube, Biological Sciences, Lipids, 030104 developmental biology, Membrane, Structural Homology, Protein, Calibration, Amphiphysin, Biophysics, 030217 neurology & neurosurgery
Abstract

International audience; Bin/Amphiphysin/Rvs (BAR) domain proteins control the curvature of lipid membranes in endocytosis, trafficking, cell motility, the formation of complex subcellular structures, and many other cellular phenomena. They form 3D assemblies that act as molecular scaffolds to reshape the membrane and alter its mechanical properties. It is unknown, however, how a protein scaffold forms and how BAR domains interact in these assemblies at protein densities relevant for a cell. In this work, we use various experimental, theoretical, and simulation approaches to explore how BAR proteins organize to form a scaffold on a membrane nanotube. By combining quantitative microscopy with analytical modeling, we demonstrate that a highly curving BAR protein endophilin nucleates its scaffolds at the ends of a membrane tube, contrary to a weaker curving protein centaurin, which binds evenly along the tube's length. Our work implies that the nature of local protein-membrane interactions can affect the specific localization of proteins on membrane-remodeling sites. Furthermore, we show that amphipathic helices are dispensable in forming protein scaffolds. Finally, we explore a possible molecular structure of a BAR-domain scaffold using coarse-grained molecular dynamics simulations. Together with fluorescence microscopy, the simulations show that proteins need only to cover 30-40% of a tube's surface to form a rigid assembly. Our work provides mechanical and structural insights into the way BAR proteins may sculpt the membrane as a high-order cooperative assembly in important biological processes.

Published
2016

18. Efficient ER Exit and Vacuole Targeting of Yeast Sna2p Require Two Tyrosine-Based Sorting Motifs

Author

Henri-François Renard, Pierre Morsomme, Didier Demaegd, and Bérengère Guerriat

Subjects
Saccharomyces cerevisiae Proteins, Vesicle-Associated Membrane Protein 2, Amino Acid Motifs, Saccharomyces cerevisiae, Vacuole, Biology, Endoplasmic Reticulum, Biochemistry, Fungal Proteins, Protein structure, Structural Biology, Yeasts, Genetics, Tyrosine, Molecular Biology, Secretory pathway, Membrane Proteins, Biological Transport, Cell Biology, biology.organism_classification, Protein Structure, Tertiary, Transport protein, Cell biology, Transcription Factor AP-1, Protein Transport, Directed mutagenesis, Membrane protein, Vacuoles
Abstract

SNA (Sensitive to Na(+)) proteins form a membrane protein family, which, in the yeast Saccharomyces cerevisiae, is composed of four members: Sna1p/Pmp3p, Sna2p, Sna3p and Sna4p. In this study, we focused on the 79 residue Sna2p protein. We found that Sna2p is localized in the vacuolar membrane. Directed mutagenesis showed that two functional tyrosine motifs YXXØ are present in the C-terminal region. Each of these is involved in a different Golgi-to-vacuole targeting pathway: the tyrosine 65 motif is involved in adaptor protein (AP-1)-dependent targeting, whereas the tyrosine 75 motif is involved in AP-3-dependent targeting. Moreover, our data suggest that these motifs also play a crucial role in the exit of Sna2p from the endoplasmic reticulum (ER). Directed mutagenesis of these tyrosines led to a partial redirection of Sna2p to lipid bodies, probably because of a decrease in ER exit efficiency. Sna2p is the first yeast protein in which two YXXØ motifs have been identified and both were shown to be functional at two different steps of the secretory pathway, ER exit and Golgi-to-vacuole transport.

Published
2010

19. Retrograde transport is not required for cytosolic translocation of the B-subunit of Shiga toxin

Author

Estelle Dransart, Eric Tartour, Henri-François Renard, Stefan J. Rathjen, Bahne Stechmann, Thi Tran, Mike Lord, Ludger Johannes, Jean-Christophe Cintrat, Jost Enninga, Alexandre Bobard, Maria Daniela Garcia-Castillo, Christophe Lamaze, Chimie biologique des membranes et ciblage thérapeutique (CBMCT - UMR 3666 / U1143), Université Paris Descartes - Paris 5 (UPD5)-Institut Curie [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie du CNRS (INC), Institut Curie [Paris], Paris-Centre de Recherche Cardiovasculaire (PARCC - UMR-S U970), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Hôpital Européen Georges Pompidou [APHP] (HEGP), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO), Hôpital Européen Georges Pompidou [APHP] (HEGP), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO), Université Paris Descartes - Paris 5 (UPD5), Université Sorbonne Paris Cité (USPC), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Dynamique des Interactions Hôte-Pathogène - Dynamics of Host-Pathogen Interactions, Institut Pasteur [Paris]-Centre National de la Recherche Scientifique (CNRS), University of Warwick [Coventry], Service de Chimie Bio-Organique et de Marquage (SCBM), Médicaments et Technologies pour la Santé (MTS), Université Paris-Saclay-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-Université Paris-Saclay-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), This work was supported by grants from the Institut National du Cancer [grant number PLBIO11-022-IDF-JOHANNES], Agence Nationale pour la Recherche [grant number ANR-11 BSV2 018 03], European Research Council (ERC) advanced grant (project 340485) to L.J., and by fellowships from AXA Research Funds and Association pour la Recherche sur le Cancer to M.D.G-C. The Johannes team is members of Labex CelTisPhyBio [grant number 11-LBX-0038] and of Idex Paris Sciences et Lettres [grant number ANR-10-IDEX-0001-02 PSL]. The Tartour team is members of Labex Immuno-Oncology and SIRIC-CARPEM and is labeled by the Ligue Nationale contre le Cancer. J.E. is supported by an ERC starting grant [Rupteffect, grant number 261166] and is member of the LabEx initiative IBEID. His group has been supported by the CARNOT-MIE program., We also thank the following people for their help with experiments and reagents, as well as scientific discussion: Bruno Goud, Stéphanie Miserey-Lenkei, Sebastian Amigorena and Daniel Gillet., ANR-11-IDEX-0005,USPC,Université Sorbonne Paris Cité(2011), ANR-10-IDEX-0001,PSL,Paris Sciences et Lettres(2010), ANR-10-LABX-0062,IBEID,Integrative Biology of Emerging Infectious Diseases(2010), European Project: 340485,EC:FP7:ERC,ERC-2013-ADG,GALECTCOMPART(2014), European Project: 261166,EC:FP7:ERC,ERC-2010-StG_20091118,RUPTEFFECTS(2011), Langlais, Laurence, Université Sorbonne Paris Cité - - USPC2011 - ANR-11-IDEX-0005 - IDEX - VALID, Initiative d'excellence - Paris Sciences et Lettres - - PSL2010 - ANR-10-IDEX-0001 - IDEX - VALID, Integrative Biology of Emerging Infectious Diseases - - IBEID2010 - ANR-10-LABX-0062 - LABX - VALID, Endocytic Membrane Compartmentalization by Galectins - GALECTCOMPART - - EC:FP7:ERC2014-04-01 - 2019-03-31 - 340485 - VALID, Revealing the mechanism of host membrane rupture by invasive pathogens and its role in triggering the immune response - RUPTEFFECTS - - EC:FP7:ERC2011-03-01 - 2016-02-29 - 261166 - VALID, Université Paris Descartes - Paris 5 (UPD5)-Institut Curie [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS), and UCL - SST/ISV - Institut des sciences de la vie

Subjects
Lactamase, MESH: Fluorescence Resonance Energy Transfer, Retro compound, Cytomegalovirus, Chromosomal translocation, CD8-Positive T-Lymphocytes, [SDV.BC.IC] Life Sciences [q-bio]/Cellular Biology/Cell Behavior [q-bio.CB], Sec22B, Epitopes, chemistry.chemical_compound, MESH: Cell Compartmentation, Cytosol, 0302 clinical medicine, Rab5, MESH: Cytosol, Rab6, Rab7, Cytotoxic T-lymphocyte, [SDV.BC.IC]Life Sciences [q-bio]/Cellular Biology/Cell Behavior [q-bio.CB], Fluorescence Resonance Energy Transfer, Golgi, Endosome, PPMP, Microdomain, [SDV.BBM.BC] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], Cancer, 0303 health sciences, Infectious disease, biology, Brefeldin A, Endosomal escape, MESH: CD8-Positive T-Lymphocytes, Endocytosis, Cell biology, Methyl-beta-cyclodextrin, Cholesterol, Biochemistry, Nanodomain, MESH: Protein Biosynthesis, MESH: Endocytosis, Ribosome Inactivating Proteins, Type 1, symbols, Immunotherapy, Bafilomycin A1, Dendritic cell, MESH: Cytomegalovirus, MESH: Epitopes, MESH: Biological Transport, MESH: Shiga Toxin, Endosomes, MESH: Ribosome Inactivating Proteins, Type 1, Major histocompatibility complex, Shiga Toxin, 03 medical and health sciences, symbols.namesake, Antigen, Humans, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], Raft, 030304 developmental biology, MESH: Humans, Endoplasmic reticulum, rab7 GTP-Binding Proteins, Biological Transport, Cell Biology, Golgi apparatus, Shiga toxin, Saporins, Cell Compartmentation, MESH: rab GTP-Binding Proteins, Antigen cross-presentation, chemistry, rab GTP-Binding Proteins, MESH: Endosomes, Protein Biosynthesis, MESH: HeLa Cells, biology.protein, Brefeldin-A, 030217 neurology & neurosurgery, HeLa Cells
Abstract

Antigen-presenting cells have the remarkable capacity to transfer exogenous antigens to the cytosol for processing by proteasomes and subsequent presentation on major histocompatibility complex class-I (MHC-I) molecules, a process termed cross-presentation. This is the target of biomedical approaches that aim to trigger a therapeutic immune response. The receptor-binding B-subunit of Shiga toxin (STxB) has been developed as an antigen delivery tool for such immunotherapy applications. In this study, we have analyzed pathways and trafficking factors that are involved in this process. A covalent conjugate between STxB and saporin was generated to quantitatively sample the membrane translocation step to the cytosol in differentiated monocyte-derived THP-1 cells. We have found that retrograde trafficking to the Golgi complex was not required for STxB–saporin translocation to the cytosol or for STxB-dependent antigen cross-presentation. Depletion of endosomal Rab7 inhibited, and lowering membrane cholesterol levels favored STxB–saporin translocation. Interestingly, experiments with reducible and non-reducible linker-arm–STxB conjugates led to the conclusion that after translocation, STxB remains associated with the cytosolic membrane leaflet. In summary, we report new facets of the endosomal escape process bearing relevance to antigen cross-presentation.

Published
2015

20. Shiga toxin stimulates clathrin-independent endocytosis of VAMP2/3/8 SNARE proteins

Author

Christophe Lamaze, Maria Daniela Garcia-Castillo, Valérie Chambon, Henri-François Renard, and Ludger Johannes

Subjects
VAMP3, Endocytic cycle, biology.protein, Shiga toxin, Cell Biology, Receptor-mediated endocytosis, Biology, Endocytosis, Clathrin, Transmembrane protein, Exocytosis, Cell biology
Abstract

Endocytosis is an essential cellular process that is often hijacked by pathogens and pathogenic products. Endocytic processes can be classified into two broad categories, those that are dependent on clathrin and those that are not. The SNARE proteins VAMP2, VAMP3 and VAMP8 are internalized in a clathrin-dependent manner. However, the full scope of their endocytic behavior has not yet been elucidated. Here, we found that VAMP2, VAMP3 and VAMP8 are localized on plasma membrane invaginations and very early uptake structures that are induced by the bacterial Shiga toxin, which enters cells by clathrin-independent endocytosis. We show that toxin trafficking into cells and cell intoxication rely on these SNARE proteins. Of note, the cellular uptake of VAMP3 is increased in the presence of Shiga toxin, even when clathrin-dependent endocytosis is blocked. We therefore conclude that VAMP2, VAMP3 and VAMP8 are removed from the plasma membrane by non-clathrin-mediated pathways, in addition to by clathrin-dependent uptake. Moreover, our study identifies these SNARE proteins as the first transmembrane trafficking factors that functionally associate at the plasma membrane with the toxin-driven clathrin-independent invaginations during the uptake process.

Published
2015

21. Rab12 localizes to Shiga toxin-induced plasma membrane invaginations and controls toxin transport

Author

Gustaf E, Rydell, Henri-François, Renard, Maria-Daniela, Garcia-Castillo, Florent, Dingli, Damarys, Loew, Christophe, Lamaze, Winfried, Römer, and Ludger, Johannes

Subjects
Protein Transport, rab GTP-Binding Proteins, Cell Membrane, Humans, Endocytosis, HeLa Cells, Shiga Toxin
Abstract

Several exogenous and endogenous cargo proteins are internalized independently of clathrin, including the bacterial Shiga toxin. The mechanisms underlying early steps of clathrin-independent uptake remain largely unknown. In this study, we have designed a protocol to obtain gradient fractions containing Shiga toxin internalization intermediates. Using stable isotope labeling with amino acids in cell culture (SILAC) and quantitative mass spectrometry, Rab12 was found in association with these very early uptake carriers. The localization of the GTPase on Shiga toxin-induced plasma membrane invaginations was shown by fluorescence microscopy in cells transfected with GFP-Rab12. Furthermore, using a quantitative biochemical assay, it was found that the amount of receptor-binding B-subunit of Shiga toxin reaching the trans-Golgi/TGN membranes was decreased in Rab12-depleted cells, and that cells were partially protected against intoxication by Shiga-like toxin 1 under these conditions. These findings demonstrate the functional importance of Rab12 for retrograde toxin trafficking. Among several other intracellular transport pathways, only the steady-state localizations of TGN46 and cation-independent mannose-6-phosphate receptor were affected. These data thus strongly suggest that Rab12 functions in the retrograde transport route.

Published
2013

Catalog

Books, media, physical & digital resources
See catalog results