Publications de l'équipe - Trafic endocytique et ciblage intracellulaire - Centre de Recherche Institut Curie
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Publications de l’équipe
Trafic endocytique et ciblage intracellulaire
Année de publication : 2021
Ivashenka# A, Wunder# C, Chambon# V, Sandhoff R, Jennemann R, Dransart# D, Podsypanina
K, Lombard B, Loew D, Lamaze C, Poirier F, Gröne HJ, Johannes* L, Shafaq-Zadah*# M (2021 Feb
9)
Glycolipid-dependent and lectin-driven transcytosis in mouse enterocytes
COMMUNICATIONS BIOLOGYNature : 4 : 173 (2021) : DOI : 10.1038
Résumé
Glycoproteins and glycolipids at the plasma membrane contribute to a range of functions
from growth factor signaling to cell adhesion and migration. Glycoconjugates undergo
endocytic trafficking. According to the glycolipid-lectin (GL-Lect) hypothesis, the construction
of tubular endocytic pits is driven in a glycosphingolipid-dependent manner by sugar-binding
proteins of the galectin family. Here, we provide evidence for a function of the GL-Lect
mechanism in transcytosis across enterocytes in the mouse intestine. We show that
galectin-3 (Gal3) and its newly identified binding partner lactotransferrin are transported in a
glycosphingolipid-dependent manner from the apical to the basolateral membrane.
Transcytosis of lactotransferrin is perturbed in Gal3 knockout mice and can be rescued by
exogenous Gal3. Inside enterocytes, Gal3 is localized to hallmark structures of the GL-Lect
mechanism, termed clathrin-independent carriers. These data pioneer the existence of GL-
Lect endocytosis in vivo and strongly suggest that polarized trafficking across the intestinal
barrier relies on this mechanism.
Karaki S, Blanc C, Tran T, Galy-Fauroux I, Mougel A, Dransart E, Anson M, Tanchot C, Gibault L,
Lepimpec-Barthes F, Darmotte D, Fabre E, Golub* R, Johannes* L, Tartour* E (2021 Jan 24)
CXCR6 deficiency impairs cancer vaccine efficacy and CD8+ resident memory T-
cell recruitment in head and neck and lung tumors
J Immunother CancerBMJ Journals : DOI : 10.1136/jitc-2020-001948
Résumé
Background Resident memory T lymphocytes (TRM) are located in tissues and play an
important role in immunosurveillance against tumors. The presence of TRM prior to treatment
or their induction is associated to the response to anti-Programmed cell death protein 1
(PD-1)/Programmed death-ligand 1 (PD-L1) immunotherapy and the efficacy of cancer
vaccines. Previous work by our group and others has shown that the intranasal route of
vaccination allows more efficient induction of these cells in head and neck and lung mucosa,
resulting in better tumor protection. The mechanisms of in vivo migration of these cells
remains largely unknown, apart from the fact that they express the chemokine receptor
CXCR6.
Methods We used CXCR6-deficient mice and an intranasal tumor vaccination model
targeting the Human Papillomavirus (HPV) E7 protein expressed by the TC-1 lung cancer
epithelial cell line. The role of CXCR6 and its ligand, CXCL16, was analyzed using
INSTITUT CURIE, 20 rue d’Ulm, 75248 Paris Cedex 05, France | 1Publications de l’équipe
Trafic endocytique et ciblage intracellulaire
multiparametric cytometric techniques and Luminex assays.
Human biopsies obtained from patients with lung cancer were also included in this study.
Results We showed that CXCR6 was preferentially expressed by CD8+ TRM after vaccination
+
in mice and also on intratumoral CD8 TRM derived from human lung cancer. We also
demonstrate that vaccination of Cxcr6-deficient mice induces a defect in the lung
recruitment of antigen-specific CD8+ T cells, preferentially in the TRM subsets. In addition, we
found that intranasal vaccination with a cancer vaccine is less effective in these Cxcr6-
deficient mice compared with wild-type mice, and this loss of efficacy is associated with
decreased recruitment of local antitumor CD8+ TRM. Interestingly, intranasal, but not
intramuscular vaccination induced higher and more sustained concentrations of CXCL16,
compared with other chemokines, in the bronchoalveolar lavage fluid and pulmonary
parenchyma.
Conclusions This work demonstrates the in vivo role of CXCR6-CXCL16 axis in the migration
+
of CD8 resident memory T cells in lung mucosa after vaccination, resulting in the control of
tumor growth. This work reinforces and explains why the intranasal route of vaccination is
the most appropriate strategy for inducing these cells in the head and neck and pulmonary
mucosa, which remains a major objective to overcome resistance to anti-PD-1/PD-L1,
especially in cold tumors.
Année de publication : 2020
Forrester A, Rathjen SJ, Garcia Castillo MD, Bachert C, Couhert A, Tepshi L, Pichard S, Martinez J,
Renard H-F, Valades Cruz CA, Dingli F, Loew D, Lamaze C, Cintrat JC, Linstedt AD, Gillet D,
Barbier J, Johannes L (2020 May 29)
Functional dissection of the retrograde Shiga toxin trafficking inhibitor Retro-2
Nature Chemical BiologyNature : 16 : 327,336 : DOI : 10.1038/s41589-020-0474-4
Résumé
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.
INSTITUT CURIE, 20 rue d’Ulm, 75248 Paris Cedex 05, France | 2Publications de l’équipe
Trafic endocytique et ciblage intracellulaire
Année de publication : 2019
Erik B. Watkins, Jaroslaw Majewski, Eva Y. Chi, Haifei Gao, Jean-Claude Florent, Ludger Johannes
(2019 Sep 20)
Shiga Toxin Induces Lipid Compression: A Mechanism for Generating Membrane
Curvature
Nano letters : 19 : DOI : 10.1021/acs.nanolett.9b03001
Résumé
Biomembranes are hard to compress laterally, and membrane area compressibility has not
been associated with biological processes. Using X-ray surface scattering, we observed that
bacterial Shiga toxin compresses lipid packing in a gel phase monolayer upon binding to its
cellular receptor, the glycolipid Gb3. This toxin-induced reorganization of lipid packing
reached beyond the immediate membrane patch that the protein was bound to, and linkers
separating the Gb3 carbohydrate and ceramide moieties modulated the toxin’s capacity to
compress the membrane. Within a natural membrane, asymmetric compression of the toxin-
bound leaflet could provide a mechanism to initiate narrow membrane bending, as observed
upon toxin entry into cells. Such lipid compression and long-range membrane reorganization
by glycolipid-binding proteins represent novel concepts in membrane biology that have
direct implications for the construction of endocytic pits in clathrin-independent endocytosis.
François Legoux, Déborah Bellet, Celine Daviaud, Yara El Morr, Aurelie Darbois, Kristina Niort,
Emanuele Procopio, Marion Salou, Jules Gilet, Bernhard Ryffel, Aurélie Balvay, Anne Foussier,
Manal Sarkis, Ahmed El Marjou, Frederic Schmidt, Sylvie Rabot, Olivier Lantz (2019 Aug 31)
Microbial metabolites control the thymic development of mucosal-associated
invariant T cells.
Science (New York, N.Y.) : DOI : eaaw2719
Résumé
How the microbiota modulate immune functions remains poorly understood. Mucosal-
associated invariant T (MAIT) cells are implicated in mucosal homeostasis and absent in
germ-free mice. Here, we show that commensal bacteria govern murine MAIT intrathymic
development, as MAIT cells did not recirculate to the thymus. MAIT development required
expression in bacteria, indicating that production of the MAIT antigen 5-(2-
oxopropylideneamino)-6-d-ribitylaminouracil (5-OP-RU) was necessary. 5-OP-RU rapidly
traveled from mucosal surfaces to the thymus, where it was captured by the major
histocompatibility complex class Ib molecule MR1. This led to increased numbers of the
earliest MAIT precursors and the expansion of more mature receptor-related orphan receptor
γt-positive MAIT cells. Thus, a microbiota-derived metabolite controls development of
mucosally targeted T cells, in a process blurring the distinction between exogenous and self-
antigens.
INSTITUT CURIE, 20 rue d’Ulm, 75248 Paris Cedex 05, France | 3Publications de l’équipe
Trafic endocytique et ciblage intracellulaire
Année de publication : 2018
Patricia Bassereau, Rui Jin, Tobias Baumgart, Markus Deserno, Rumiana Dimova, Vadim A.
Frolov, Pavel V. Bashkirov, Helmut Grubmüller, Reinhard Jahn, H. Jelger Risselada, Ludger
Johannes, Michael M. Kozlov, Reinhard Lipowsky, Thomas J. Pucadyil, Wade F. Zeno, Jeanne C.
Stachowiak, Dimitrios Stamou, Artù Breuer, Line Lauritsen, Camille Simon, Cécile Sykes, Gregory
A. Voth, Thomas R Weikl (2018 Jul 20)
The 2018 biomembrane curvature and remodeling roadmap.
Journal of Physics D: Applied Physics : 51 : 343001 : DOI : 10.1088/1361-6463/aacb98
Résumé
The importance of curvature as a structural feature of biological membranes has been
recognized for many years and has fascinated scientists from a wide range of different
backgrounds. On the one hand, changes in membrane morphology are involved in a plethora
of phenomena involving the plasma membrane of eukaryotic cells, including endo- and
exocytosis, phagocytosis and filopodia formation. On the other hand, a multitude of
intracellular processes at the level of organelles rely on generation, modulation, and
maintenance of membrane curvature to maintain the organelle shape and functionality. The
contribution of biophysicists and biologists is essential for shedding light on the mechanistic
understanding and quantification of these processes.
Given the vast complexity of phenomena and mechanisms involved in the coupling between
membrane shape and function, it is not always clear in what direction to advance to
eventually arrive at an exhaustive understanding of this important research area. The 2018
Biomembrane Curvature and Remodeling Roadmap of Journal of Physics D: Applied
Physics addresses this need for clarity and is intended to provide guidance both for students
who have just entered the field as well as established scientists who would like to improve
their orientation within this fascinating area.
Silvia Serra, Ahmed Alouane, Thomas Le Saux, Steve Huvelle, Raphaël Plasson, Frédéric
Schmidt, Ludovic Jullien, Raphaël Labruère (2018 Jun 7)
A chemically encoded timer for dual molecular delivery at tailored ranges and
concentrations.
Chemical communications (Cambridge, England) : 6396-6399 : DOI : 10.1039/c8cc03253j
Résumé
Spatiotemporal control of molecular distribution is much in demand in many fields of
chemistry. To address this goal, we exploit a low molecular weight branched self-immolative
architecture, which acts as a triggerable chemically encoded timer for autonomous
sequential release of two chemicals. Using a light-activated model liberating two distinct
fluorophores, we generated a tunable spatially contrasted molecular distribution.
INSTITUT CURIE, 20 rue d’Ulm, 75248 Paris Cedex 05, France | 4Publications de l’équipe
Trafic endocytique et ciblage intracellulaire
Johannes Ludger, Pezeshkian W, Ipsen JH, Shillcock J (2018 May 7)
Clustering on Membranes: Fluctuations and More
Trends Cell BiolTrends Cell Biol : 28 : 405 415 : DOI : 10.1016/j.tcb.2018.01.009
Résumé
Clustering of extracellular ligands and proteins on the plasma membrane is required to
perform specific cellular functions, such as signaling and endocytosis. Attractive forces that
originate in perturbations of the membrane’s physical properties contribute to this
clustering, in addition to direct protein-protein interactions. However, these membrane-
mediated forces have not all been equally considered, despite their importance. In this
review, we describe how line tension, lipid depletion, and membrane curvature contribute to
membrane-mediated clustering. Additional attractive forces that arise from protein-induced
perturbation of a membrane’s fluctuations are also described. This review aims to provide a
survey of the current understanding of membrane-mediated clustering and how this supports
precise biological functions.
Année de publication : 2017
Steve Huvelle, Ahmed Alouane, Thomas Le Saux, Ludovic Jullien, Frédéric Schmidt (2017 Mar 31)
Syntheses and kinetic studies of cyclisation-based self-immolative spacers.
Organic & biomolecular chemistry : 3435-3443 : DOI : 10.1039/c7ob00121e
Résumé
Kinetic analysis of the disassembly of self-immolative spacers based on cyclisation processes
was performed. Five compounds were synthesized belonging to two different series, and
their kinetic constants were determined. Electron-donating substituents gave a slight
acceleration but the main effect was steric, and the Thorpe-Ingold effect was indeed
particularly effective. Comparison with the self-immolative spacers based on elimination
processes showed that cyclisations gave comparable or lower rate, but the corresponding
spacers are more difficult to modulate.
Weria Pezeshkian, Haifei Gao, Senthil Arumugam, Ulrike Becken, Patricia Bassereau, Jean-Claude
Florent, John Hjort Ipsen, Ludger Johannes, Julian C Shillcock (2017 Jan 24)
Mechanism of Shiga Toxin Clustering on Membranes.
ACS Nano : 11 : 314-324 : DOI : 10.1021/acsnano.6b05706
Résumé
The bacterial Shiga toxin interacts with its cellular receptor, the glycosphingolipid
globotriaosylceramide (Gb3 or CD77), as a first step to entering target cells. Previous studies
have shown that toxin molecules cluster on the plasma membrane, despite the apparent
lack of direct interactions between them. The precise mechanism by which this clustering
INSTITUT CURIE, 20 rue d’Ulm, 75248 Paris Cedex 05, France | 5Publications de l’équipe
Trafic endocytique et ciblage intracellulaire
occurs remains poorly defined. Here, we used vesicle and cell systems and computer
simulations to show that line tension due to curvature, height, or compositional mismatch,
and lipid or solvent depletion cannot drive the clustering of Shiga toxin molecules. By
contrast, in coarse-grained computer simulations, a correlation was found between clustering
and toxin nanoparticle-driven suppression of membrane fluctuations, and experimentally we
observed that clustering required the toxin molecules to be tightly bound to the membrane
surface. The most likely interpretation of these findings is that a membrane fluctuation-
induced force generates an effective attraction between toxin molecules. Such force would
be of similar strength to the electrostatic force at separations around 1 nm, remain strong at
distances up to the size of toxin molecules (several nanometers), and persist even beyond.
This force is predicted to operate between manufactured nanoparticles providing they are
sufficiently rigid and tightly bound to the plasma membrane, thereby suggesting a route for
the targeting of nanoparticles to cells for biomedical applications.
Année de publication : 2016
Ludger Johannes, Christian Wunder, Massiullah Shafaq-Zadah (2016 Dec 17)
Glycolipids and Lectins in Endocytic Uptake Processes.
Journal of molecular biology : DOI : S0022-2836(16)30453-3
Résumé
A host of endocytic processes has been described at the plasma membrane of eukaryotic
cells. Their categorization has most commonly referenced cytosolic machinery, of which the
clathrin coat has occupied a preponderant position. In what concerns intra-membrane
constituents, the focus of interest has been on phosphatidylinositol lipids and their capacity
to orchestrate endocytic events on the cytosolic leaflet of the membrane. The contribution of
extracellular determinants to the construction of endocytic pits has received much less
attention, depite the fact that (glyco)sphingolipids are exoplasmic leaflet fabric of membrane
domains, termed rafts, whose contributions to predominantly clathrin-independent
internalization processes is well recognized. Furthermore, sugar modifications on
extracellular domains of proteins, and sugar-binding proteins, termed lectins, have also been
linked to the uptake of endocytic cargoes at the plasma membrane. In this review, we first
summarize these contributions by extracellular determinants to the endocytic process. We
thus propose a molecular hypothesis – termed the GL-Lect hypothesis – on how GlycoLipids
and Lectins drive the formation of compositional nanoenvrionments from which the
endocytic uptake of glycosylated cargo proteins is operated via clathrin-independent
carriers. Finally, we position this hypothesis within the global context of endocytic pathway
proposals that have emerged in recent years.
Daniela Chmiest, Nanaocha Sharma, Natacha Zanin, Christine Viaris de Lesegno, Massiullah
Shafaq-Zadah, Vonick Sibut, Florent Dingli, Philippe Hupé, Stephan Wilmes, Jacob Piehler,
Damarys Loew, Ludger Johannes, Gideon Schreiber, Christophe Lamaze (2016 Dec 6)
INSTITUT CURIE, 20 rue d’Ulm, 75248 Paris Cedex 05, France | 6Publications de l’équipe
Trafic endocytique et ciblage intracellulaire
Spatiotemporal control of interferon-induced JAK/STAT signalling and gene
transcription by the retromer complex.
Nature communications : 13476 : DOI : 10.1038/ncomms13476
Résumé
Type-I interferons (IFNs) play a key role in the immune defences against viral and bacterial
infections, and in cancer immunosurveillance. We have established that clathrin-dependent
endocytosis of the type-I interferon (IFN-α/β) receptor (IFNAR) is required for JAK/STAT
signalling. Here we show that the internalized IFNAR1 and IFNAR2 subunits of the IFNAR
complex are differentially sorted by the retromer at the early endosome. Binding of the
retromer VPS35 subunit to IFNAR2 results in IFNAR2 recycling to the plasma membrane,
whereas IFNAR1 is sorted to the lysosome for degradation. Depletion of VPS35 leads to
abnormally prolonged residency and association of the IFNAR subunits at the early
endosome, resulting in increased activation of STAT1- and IFN-dependent gene transcription.
These experimental data establish the retromer complex as a key spatiotemporal regulator
of IFNAR endosomal sorting and a new factor in type-I IFN-induced JAK/STAT signalling and
gene transcription.
Ludger Johannes, Christian Wunder (2016 Dec 3)
Retromer Sets a Trap for Endosomal Cargo Sorting.
Cell : 1452-1454 : DOI : S0092-8674(16)31598-7
Résumé
Membrane trafficking from endosomes to the trans-Golgi network or the plasma membrane
is driven by the retromer complex. Through structural analysis of the cargo-bound complex,
Lucas et al. describe a mechanism by which endosomal membrane recruitment and cargo
recognition are integrated through cooperative interactions between retromer subunits.
Romain Daillère, Marie Vétizou, Nadine Waldschmitt, Takahiro Yamazaki, Christophe Isnard,
Vichnou Poirier-Colame, Connie P M Duong, Caroline Flament, Patricia Lepage, Maria Paula
Roberti, Bertrand Routy, Nicolas Jacquelot, Lionel Apetoh, Sonia Becharef, Sylvie Rusakiewicz,
Philippe Langella, Harry Sokol, Guido Kroemer, David Enot, Antoine Roux, Alexander Eggermont,
Eric Tartour, Ludger Johannes, Paul-Louis Woerther, Elisabeth Chachaty, Jean-Charles Soria,
Encouse Golden, Silvia Formenti, Magdalena Plebanski, Mutsa Madondo, Philip Rosenstiel, Didier
Raoult, Vincent Cattoir, Ivo Gomperts Boneca, Mathias Chamaillard, Laurence Zitvogel (2016 Oct
9)
Enterococcus hirae and Barnesiella intestinihominis Facilitate
Cyclophosphamide-Induced Therapeutic Immunomodulatory Effects.
Immunity : DOI : S1074-7613(16)30378-8
INSTITUT CURIE, 20 rue d’Ulm, 75248 Paris Cedex 05, France | 7Publications de l’équipe
Trafic endocytique et ciblage intracellulaire
Résumé
The efficacy of the anti-cancer immunomodulatory agent cyclophosphamide (CTX) relies on
intestinal bacteria. How and which relevant bacterial species are involved in tumor
immunosurveillance, and their mechanism of action are unclear. Here, we identified two
bacterial species, Enterococcus hirae and Barnesiella intestinihominis that are involved
during CTX therapy. Whereas E. hirae translocated from the small intestine to secondary
lymphoid organs and increased the intratumoral CD8/Treg ratio, B. intestinihominis
accumulated in the colon and promoted the infiltration of IFN-γ-producing γδT cells in cancer
lesions. The immune sensor, NOD2, limited CTX-induced cancer immunosurveillance and the
bioactivity of these microbes. Finally, E. hirae and B. intestinihominis specific-memory Th1
cell immune responses selectively predicted longer progression-free survival in advanced
lung and ovarian cancer patients treated with chemo-immunotherapy. Altogether, E. hirae
and B. intestinihominis represent valuable « oncomicrobiotics » ameliorating the efficacy of
the most common alkylating immunomodulatory compound.
N Gupta, R Noël, A Goudet, K Hinsinger, A Michau, V Pons, H Abdelkafi, T Secher, A Shima, O
Shtanko, Y Sakurai, S Cojean, S Pomel, V Liévin-Le Moal, V Leignel, J-A Herweg, A Fischer, L
Johannes, Kate Harrison, Philippa M Beard, P Clayette, R Le Grand, J O Rayner, T Rudel, J Vacus, P
M Loiseau, R A Davey, E Oswald, J-C Cintrat, J Barbier, D Gillet (2016 Oct 8)
Inhibitors of retrograde trafficking active against ricin and Shiga toxins also
protect cells from several viruses, Chlamydiales and Leishmania.
Chemico-biological interactions : DOI : S0009-2797(16)30427-6
Résumé
Medical countermeasures to treat biothreat agent infections require broad-spectrum
therapeutics that do not induce agent resistance. A cell-based high-throughput screen (HTS)
against ricin toxin combined with hit optimization allowed selection of a family of compounds
that meet these requirements. The hit compound Retro-2 and its derivatives have been
demonstrated to be safe in vivo in mice even at high doses. Moreover, Retro-2 is an inhibitor
of retrograde transport that affects syntaxin-5-dependent toxins and pathogens. As a
consequence, it has a broad-spectrum activity that has been demonstrated both in vitro and
in vivo against ricin, Shiga toxin-producing O104:H4 entero-hemorrhagic E. coli and
Leishmania sp. and in vitro against Ebola, Marburg and poxviruses and Chlamydiales. An
effect is anticipated on other toxins or pathogens that use retrograde trafficking and
syntaxin-5. Since Retro-2 targets cell components of the host and not directly the pathogen,
no selection of resistant pathogens is expected. These lead compounds need now to be
developed as drugs for human use.
INSTITUT CURIE, 20 rue d’Ulm, 75248 Paris Cedex 05, France | 8You can also read
