Nathalie Sauvonnet
Pasteur Institute
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Featured researches published by Nathalie Sauvonnet.
Nature | 2015
Emmanuel Boucrot; Antonio P.A. Ferreira; Leonardo Almeida-Souza; Sylvain Debard; Yvonne Vallis; Gillian Howard; Laetitia Bertot; Nathalie Sauvonnet; Harvey T. McMahon
Endocytosis is required for internalization of micronutrients and turnover of membrane components. Endophilin has been assigned as a component of clathrin-mediated endocytosis. Here we show in mammalian cells that endophilin marks and controls a fast-acting tubulovesicular endocytic pathway that is independent of AP2 and clathrin, activated upon ligand binding to cargo receptors, inhibited by inhibitors of dynamin, Rac, phosphatidylinositol-3-OH kinase, PAK1 and actin polymerization, and activated upon Cdc42 inhibition. This pathway is prominent at the leading edges of cells where phosphatidylinositol-3,4-bisphosphate—produced by the dephosphorylation of phosphatidylinositol-3,4,5-triphosphate by SHIP1 and SHIP2—recruits lamellipodin, which in turn engages endophilin. This pathway mediates the ligand-triggered uptake of several G-protein-coupled receptors such as α2a- and β1-adrenergic, dopaminergic D3 and D4 receptors and muscarinic acetylcholine receptor 4, the receptor tyrosine kinases EGFR, HGFR, VEGFR, PDGFR, NGFR and IGF1R, as well as interleukin-2 receptor. We call this new endocytic route fast endophilin-mediated endocytosis (FEME).
Gene | 1997
Anthony P. Pugsley; Olivera Francetic; Odile Possot; Nathalie Sauvonnet; Kim R. Hardie
The main terminal branch (MTB) of the general secretory pathway is used by a wide variety of Gram- bacteria to transport exoproteins from the periplasm to the outside milieu. Recent work has led to the identification of the function of two of its 14 (or more) components: an enzyme with type-IV prepilin peptidase activity and a chaperone-like protein required for the insertion of another of the MTB components into the outer membrane. Despite these important discoveries, little tangible progress has been made towards identifying MTB components that determine secretion specificity (presumably by binding to cognate exoproteins) or which form the putative channel through which exoproteins are transported across the outer membrane. However, the idea that the single integral outer membrane component of the MTB could line the wall of this channel, and the intriguing possibility that other components of the MTB form a rudimentary type-IV pilus-like structure that might span the periplasm both deserve more careful examination. Although Escherichia coli K-12 does not normally secrete exoproteins, its chromosome contains an apparently complete set of genes coding for MTB components. At least two of these genes code for functional proteins, but the operon in which twelve of the genes are located does not appear to be expressed. We are currently searching for conditions which allow these genes to be expressed with the eventual aim of identifying the protein(s) that E. coli K-12 can secrete.
Molecular Microbiology | 1996
Nathalie Sauvonnet; Anthony P. Pugsley
Pullulanase (PulA) is a 116kDa amylolytic lipoprotein secreted by the Gram‐negative bacterium Klebsiella oxytoca via the general secretory pathway. A deletion strategy was used in an attempt to determine the nature and the location of the secretion signal(s) in PulA presumed to be necessary for its specific secretion. The starting material was a gene fusion coding for an efficiently secreted PulA‐β‐lactamase hybrid protein. Successive series of exonuclease III‐generated deletions were used to remove internal segments of PulA from this hybrid. A simple plate test allowed the identification of truncated hybrids that retained β‐lactamase activity and that were secreted. Two non‐adjacent regions, A and B (78 and 80 amino acids, respectively), were together necessary and sufficient to promote β‐lactamase translocation across the outer membrane. Secretion of PulA itself was markedly reduced when either of these regions was deleted, and was completely abolished when both regions were eliminated.
EMBO Reports | 2008
Alexandre Grassart; Annick Dujeancourt; Paul B. Lazarow; Alice Dautry-Varsat; Nathalie Sauvonnet
There are several endocytic pathways, which are either dependent on or independent of clathrin. This study focuses on a poorly characterized mechanism—clathrin‐ and caveolae‐independent endocytosis—used by the interleukin‐2 receptor β (IL‐2Rβ). We address the question of its regulation in comparison with the clathrin‐dependent pathway. First, we show that Ras‐related C3 botulinum toxin substrate 1 (Rac1) is specifically required for IL‐2Rβ entry, and we identify p21‐activated kinases (Paks) as downstream targets. By RNA interference, we show that Pak1 and Pak2 are both necessary for IL‐2Rβ uptake, in contrast to the clathrin‐dependent route. We observe that cortactin, a partner of actin and dynamin—two essential endocytic factors—is required for IL‐2Rβ uptake. Furthermore, we find that cortactin acts downstream from Paks, suggesting control of its function by these kinases. Thus, we describe a cascade composed of Rac1, Paks and cortactin specifically regulating IL‐2Rβ internalization. This study indicates Paks as the first specific regulators of the clathrin‐independent endocytosis pathway.
Journal of Bacteriology | 2000
Nathalie Sauvonnet; Pierre Gounon; Anthony P. Pugsley
Escherichia coli K-12 possesses at least 16 chromosomal genes related to genes involved in the formation of type IV pili in other gram-negative bacteria. However, E. coli K-12 does not produce type IV pili when grown under standard laboratory conditions. The results of reverse transcription-PCR, operon fusion analysis, and immunoblotting demonstrated that several of the putative E. coli piliation genes are expressed at very low levels. Increasing the level of expression of the major pilin gene (ppdD) and the linked assembly genes hofB and hofC (homologues of the Pseudomonas aeruginosa type IV pilus assembly genes pilB and pilC) did not lead to pilus production. However, expression of the ppdD gene in P. aeruginosa led to assembly of PpdD into pili that were recognized by antibodies directed against the PpdD protein. Assembly of PpdD into pili in P. aeruginosa was dependent on the expression of the pilB and pilC genes and independent of expression of the P. aeruginosa pilin structural gene pilA.
Science | 2014
Mathieu Boissan; Guillaume Montagnac; Qinfang Shen; Lorena Griparic; Jérôme Guitton; Maryse Romao; Nathalie Sauvonnet; Thibault Lagache; Ioan Lascu; Graça Raposo; Céline Desbourdes; Uwe Schlattner; Marie-Lise Lacombe; Simona Polo; Alexander M. van der Bliek; Aurélien Roux; Philippe Chavrier
Supplying power: Right time, right place Cell membranes are very flexible and easily molded to shape; however, to physically pinch off a membrane vesicle from a membrane tube still requires power. A type of molecular machine known as dynamin is involved in this sort of membrane remodeling. Dynamins use guanosine triphosphate (GTP) rather than the more commonly used cellular energy source adenosine triphosphate to work. Boissan et al. now show that two separate dynamins found in the cytoplasm or the mitochondria both use the same sort of enzyme—nucleoside diphosphate kinases—to provide GTP at just the right time and the right place to power membrane fission. Science, this issue p. 1510 During membrane fission, molecular motors are provided with a local energy source. Dynamin superfamily molecular motors use guanosine triphosphate (GTP) as a source of energy for membrane-remodeling events. We found that knockdown of nucleoside diphosphate kinases (NDPKs) NM23-H1/H2, which produce GTP through adenosine triphosphate (ATP)–driven conversion of guanosine diphosphate (GDP), inhibited dynamin-mediated endocytosis. NM23-H1/H2 localized at clathrin-coated pits and interacted with the proline-rich domain of dynamin. In vitro, NM23-H1/H2 were recruited to dynamin-induced tubules, stimulated GTP-loading on dynamin, and triggered fission in the presence of ATP and GDP. NM23-H4, a mitochondria-specific NDPK, colocalized with mitochondrial dynamin-like OPA1 involved in mitochondria inner membrane fusion and increased GTP-loading on OPA1. Like OPA1 loss of function, silencing of NM23-H4 but not NM23-H1/H2 resulted in mitochondrial fragmentation, reflecting fusion defects. Thus, NDPKs interact with and provide GTP to dynamins, allowing these motor proteins to work with high thermodynamic efficiency.
Traffic | 2010
Alexandre Grassart; Vannary Meas-Yedid; Alexandre Dufour; Jean-Christophe Olivo-Marin; Alice Dautry-Varsat; Nathalie Sauvonnet
Growing evidence indicates that kinases are central to the regulation of endocytic pathways. Previously, we identified p21‐activated kinase 1 (Pak1) as the first specific regulator of clathrin‐ and caveolae‐independent endocytosis used by the interleukin 2 receptor subunit (IL‐2R). Here, we address the mechanism by which Pak1 regulates IL‐2Rβ endocytosis. First, we show that Pak1 phosphorylates an activator of actin polymerization, cortactin, on its serine residues 405 and 418. Consistently, we observe a specific inhibition of IL‐2Rβ endocytosis when cells overexpress a cortactin, wherein these serine residues have been mutated. In addition, we show that the actin polymerization enhancer, neuronal Wiskott–Aldrich syndrome protein (N‐WASP), is involved in IL‐2Rβ endocytosis. Strikingly, we find that Pak1 phosphorylation of cortactin on serine residues 405 and 418 increases its association with N‐WASP. Thus, Pak1, by controlling the interaction between cortactin and N‐WASP, could regulate the polymerization of actin during clathrin‐independent endocytosis.
Cell Host & Microbe | 2012
Joëlle Mounier; Gaelle Boncompain; Lidija Senerovic; Thibault Lagache; Fabrice Chrétien; Franck Perez; Michael Kolbe; Jean-Christophe Olivo-Marin; Philippe J. Sansonetti; Nathalie Sauvonnet
Shigella infection causes destruction of the human colonic epithelial barrier. The Golgi network and recycling endosomes are essential for maintaining epithelial barrier function. Here we show that Shigella epithelial invasion induces fragmentation of the Golgi complex with consequent inhibition of both secretion and retrograde transport in the infected host cell. Shigella induces tubulation of the Rab11-positive compartment, thereby affecting cell surface receptor recycling. The molecular process underlying the observed damage to the Golgi complex and receptor recycling is a massive redistribution of plasma membrane cholesterol to the sites of Shigella entry. IpaB, a virulence factor of Shigella that is known to bind cholesterol, is necessary and sufficient to induce Golgi fragmentation and reorganization of the recycling compartment. Shigella infection-induced Golgi disorganization was also observed in vivo, suggesting that this mechanism affecting the sorting of cell surface molecules likely contributes to host epithelial barrier disruption associated with Shigella pathogenesis.
Journal of Bacteriology | 2001
Anthony P. Pugsley; Nicolas Bayan; Nathalie Sauvonnet
When expressed in Escherichia coli, the 15 Klebsiella oxytoca pul genes that encode the so-called Pul secreton or type II secretion machinery promote pullulanase secretion and the assembly of one of the secreton components, PulG, into pili. Besides these pul genes, efficient pullulanase secretion also requires the host dsbA gene, encoding a periplasmic disulfide oxidoreductase, independently of disulfide bond formation in pullulanase itself. Two secreton components, the secretin pilot protein PulS and the minor pseudopilin PulK, were each shown to posses an intramolecular disulfide bond whose formation was catalyzed by DsbA. PulS was apparently destabilized by the absence of its disulfide bond, whereas PulK stability was not dramatically affected either by a dsbA mutation or by the removal of one of its cysteines. The pullulanase secretion defect in a dsbA mutant was rectified by overproduction of PulK, indicating reduced disulfide bond formation in PulK as the major cause of the secretion defect under the conditions tested (in which PulS is probably present in considerable excess of requirements). PulG pilus formation was independent of DsbA, probably because PulK is not needed for piliation.
PLOS ONE | 2012
Darran J. Wigelsworth; Gordon Ruthel; Leonie Schnell; Peter Herrlich; Josip Blonder; Timothy D. Veenstra; Robert J. Carman; Tracy D. Wilkins; Guy Tran Van Nhieu; Serge Pauillac; Nathalie Sauvonnet; Bradley G. Stiles; Michel R. Popoff; Holger Barth
Various pathogenic clostridia produce binary protein toxins associated with enteric diseases of humans and animals. Separate binding/translocation (B) components bind to a protein receptor on the cell surface, assemble with enzymatic (A) component(s), and mediate endocytosis of the toxin complex. Ultimately there is translocation of A component(s) from acidified endosomes into the cytosol, leading to destruction of the actin cytoskeleton. Our results revealed that CD44, a multifunctional surface protein of mammalian cells, facilitates intoxication by the iota family of clostridial binary toxins. Specific antibody against CD44 inhibited cytotoxicity of the prototypical Clostridium perfringens iota toxin. Versus CD44+ melanoma cells, those lacking CD44 bound less toxin and were dose-dependently resistant to C. perfringens iota, as well as Clostridium difficile and Clostridium spiroforme iota-like, toxins. Purified CD44 specifically interacted in vitro with iota and iota-like, but not related Clostridium botulinum C2, toxins. Furthermore, CD44 knockout mice were resistant to iota toxin lethality. Collective data reveal an important role for CD44 during intoxication by a family of clostridial binary toxins.