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Thierry Galli - One of the best experts on this subject based on the ideXlab platform.
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Soluble N‐ethylmaleimide‐sensitive factor attachment protein receptors required during Trypanosoma cruzi parasitophorous vacuole development
Cellular Microbiology, 2017Co-Authors: Juan Agustín Cueto, María Cristina Vanrell, Betiana Nebaí Salassa, Sébastien Nola, Maria I. Colombo, Thierry Galli, Patricia Silvia RomanoAbstract:Trypanosoma cruzi, the etiologic agent of Chagas disease, is an obligate intracellular parasite that exploits different host vesicular pathways to invade the target cells. Vesicular and target SNAREs are key proteins of the intracellular membrane fusion machinery. During the early times of T. cruzi infection, several vesicles are attracted to the parasite contact sites in the plasma membrane. Fusion of these vesicles promotes the formation of the parasitic vacuole and parasite entry. In this work, we study the requirement and the nature of SNAREs involved in the fusion events which take place during T. cruzi infection. Our results show that inhibition of NSF, a protein required for SNARE complex disassembly, impairs T. cruzi infection. Both TI-VAMP/VAMP7 and cellubrevin/VAMP3, two v-SNAREs of the endocytic and exocytic pathways, are specifically recruited to the parasitophorous vacuole membrane in a synchronized manner but, although VAMP3 is acquired earlier than VAMP7, impairment of VAMP3 by tetanus neurotoxin fails to reduce T. cruzi infection. In contrast, reduction of VAMP7 activity by expression of VAMP7's Longin domain, depletion by siRNA or knock out, significantly decreases T. cruzi infection susceptibility as a result of a minor acquisition of lysosomal components to the parasitic vacuole. In addition, overexpression of the VAMP7 partner Vti1b, increases the infection whereas expression of a KIF5 kinesin mutant reduces VAMP7 recruitment to vacuole and concomitantly, T. cruzi infection. Altogether, these data support a key role of TI-VAMP/VAMP7 in the fusion events that culminate in the T. cruzi parasitophorous vacuole development.
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role of tetanus neurotoxin insensitive vesicle associated membrane protein in membrane domains transport and homeostasis
Cellular logistics, 2015Co-Authors: Diana Molino, Sébastien Nola, Veronique Prouxgillardeaux, Agathe Verraes, Lydia Danglot, Gaelle Boncompain, Franck Perez, Markus R Wenk, Guanghou Shui, Thierry GalliAbstract:Biological membranes in eukaryotes contain a large variety of proteins and lipids often distributed in domains in plasma membrane and endomembranes. Molecular mechanisms responsible for the transport and the organization of these membrane domains along the secretory pathway still remain elusive. Here we show that vesicular SNARE TI-VAMP/VAMP7 plays a major role in membrane domains composition and transport. We found that the transport of exogenous and endogenous GPI-anchored proteins was altered in fibroblasts isolated from VAMP7-knockout mice. Furthermore, disassembly and reformation of the Golgi apparatus induced by Brefeldin A treatment and washout were impaired in VAMP7-depleted cells, suggesting that loss of VAMP7 expression alters biochemical properties and dynamics of the Golgi apparatus. In addition, lipid profiles from these knockout cells indicated a defect in glycosphingolipids homeostasis. We conclude that VAMP7 is required for effective transport of GPI–anchored proteins to cell surface and t...
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role of vamp3 and vamp7 in the commitment of yersinia pseudotuberculosis to lc3 associated pathways involving single or double membrane vacuoles
Autophagy, 2014Co-Authors: Laure-anne Ligeon, Nicolas Barois, Antonino Bongiovanni, Elisabeth Werkmeister, Veronique Prouxgillardeaux, Delphine Armelle Lacorre, Kevin Moreau, Thierry Galli, Frank LafontAbstract:Yersinia pseudotuberculosis can replicate inside macrophages by hijacking autophagy and blocking autophagosome acidification. In bone marrow-derived macrophages, the bacteria are mainly observed inside double-membrane vacuoles positive for LC3, a hallmark of autophagy. Here, we address the question of the membrane traffic during internalization of Yersinia investigating the role of vesicle- associated membrane proteins (VAMPs). First, we show that as in epithelial cells, Yersinia pseudotuberculosis replicates mainly in nonacidic LC3-positive vacuoles. Second, in these cells, we unexpectedly found that VAMP3 localizes preferentially to Yersinia-containing vacuoles (YCVs) with single membranes using correlative light-electron microscopy. Third, we reveal the precise kinetics of VAMP3 and VAMP7 association with YCVs positive for LC3. Fourth, we show that VAMP7 knockdown alters LC3′s association with single-and multimembrane-YCVs. Finally, in uninfected epithelial cells stimulated for autophagy, VAMP3 overexp...
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Increased activity of the Vesicular Soluble N -Ethylmaleimide-sensitive Factor Attachment Protein Receptor TI-VAMP/VAMP7 by Tyrosine Phosphorylation in the Longin Domain
Journal of Biological Chemistry, 2013Co-Authors: Andrea Burgo, Sébastien Nola, Alessandra Maria Casano, Agathe Verraes, Aurelia Kuster, Florent Dingli, Damarys Loew, Stefan Arold, Guan Wang, Thierry GalliAbstract:Vesicular (v)- and target (t)-SNAREs play essential roles in intracellular membrane fusion through the formation of cytoplasmic α-helical bundles. Several v-SNAREs have a Longin N-terminal extension that, by promoting a closed conformation, plays an autoinhibitory function and decreases SNARE complex formation and membrane fusion efficiency. The molecular mechanism leading to Longin v-SNARE activation is largely unknown. Here we find that exocytosis mediated by the Longin v-SNARE TI-VAMP/VAMP7 is activated by tonic treatment with insulin and insulin-like growth factor-1 but not by depolarization and intracellular calcium rise. In search of a potential downstream mechanism, we found that TI-VAMP is phosphorylated in vitro by c-Src kinase on tyrosine 45 of the Longin domain. Accordingly, a mutation of tyrosine 45 into glutamate, but not phenylalanine, activates both t-SNARE binding and exocytosis. Activation of TI-VAMP-mediated exocytosis thus relies on tyrosine phosphorylation.
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A Molecular Network for the Transport of the TI-VAMP/VAMP7 Vesicles from Cell Center to Periphery
Developmental Cell, 2012Co-Authors: Andrea Burgo, Véronique Proux-gillardeaux, Emmanuel Sotirakis, Alessandra Maria Casano, Agathe Verraes, Maïté Coppey-moisan, Ronald K.h. Liem, Etienne Formstecher, Thierry GalliAbstract:The compartmental organization of eukaryotic cells is maintained dynamically by vesicular trafficking. SNARE proteins play a crucial role in intracellular membrane fusion and need to be targeted to their proper donor or acceptor membrane. The molecular mechanisms that allow for the secretory vesicles carrying the v-SNARE TI-VAMP/VAMP7 to leave the cell center, load onto microtubules, and reach the periphery to mediate exocytosis are largely unknown. Here, we show that the TI-VAMP/VAMP7 partner Varp, a Rab21 guanine nucleotide exchange factor, interacts with GolginA4 and the kinesin 1 Kif5A. Activated Rab21-GTP in turn binds to MACF1, an actin and microtubule regulator, which is itself a partner of GolginA4. These components are required for directed movement of TI-VAMP/VAMP7 vesicles from the cell center to the cell periphery. The molecular mechanisms uncovered here suggest an integrated view of the transport of vesicles carrying a specific v-SNARE toward the cell surface.
Martin Oheim - One of the best experts on this subject based on the ideXlab platform.
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astrocyte vamp3 vesicles undergo ca2 independent cycling and modulate glutamate transporter trafficking
The Journal of Physiology, 2015Co-Authors: Karine Herault, Kathleen Zylbersztejn, Martin Oheim, Marcel A. Lauterbach, Marc Guillon, Dongdong LiAbstract:Key points Mouse cortical astrocytes express VAMP3 but not VAMP2. VAMP3 vesicles undergo Ca2+-independent exo- and endocytotic cycling at the plasma membrane. VAMP3 vesicle traffic regulates the recycling of plasma membrane glutamate transporters. cAMP modulates VAMP3 vesicle cycling and glutamate uptake. Abstract Previous studies suggest that small synaptic-like vesicles in astrocytes carry vesicle-associated vSNARE proteins, VAMP3 (cellubrevin) and VAMP2 (synaptobrevin 2), both contributing to the Ca2+-regulated exocytosis of gliotransmitters, thereby modulating brain information processing. Here, using cortical astrocytes taken from VAMP2 and VAMP3 knock-out mice, we find that astrocytes express only VAMP3. The morphology and function of VAMP3 vesicles were studied in cultured astrocytes at single vesicle level with stimulated emission depletion (STED) and total internal reflection fluorescence (TIRF) microscopies. We show that VAMP3 antibodies label small diameter (∼80 nm) vesicles and that VAMP3 vesicles undergo Ca2+-independent exo-endocytosis. We also show that this pathway modulates the surface expression of plasma membrane glutamate transporters and the glutamate uptake by astrocytes. Finally, using pharmacological and optogenetic tools, we provide evidence suggesting that the cytosolic cAMP level influences astrocytic VAMP3 vesicle trafficking and glutamate transport. Our results suggest a new role for VAMP3 vesicles in astrocytes.
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Astrocyte VAMP3 vesicles undergo Ca2+ -independent cycling and modulate glutamate transporter trafficking.
The Journal of Physiology, 2015Co-Authors: Karine Herault, Kathleen Zylbersztejn, Martin Oheim, Marcel A. Lauterbach, Marc Guillon, Nicole RopertAbstract:Mouse cortical astrocytes express VAMP3 but not VAMP2. VAMP3 vesicles undergo Ca(2+) -independent exo- and endocytotic cycling at the plasma membrane. VAMP3 vesicle traffic regulates the recycling of plasma membrane glutamate transporters. cAMP modulates VAMP3 vesicle cycling and glutamate uptake. Previous studies suggest that small synaptic-like vesicles in astrocytes carry vesicle-associated vSNARE proteins, VAMP3 (cellubrevin) and VAMP2 (synaptobrevin 2), both contributing to the Ca(2+) -regulated exocytosis of gliotransmitters, thereby modulating brain information processing. Here, using cortical astrocytes taken from VAMP2 and VAMP3 knock-out mice, we find that astrocytes express only VAMP3. The morphology and function of VAMP3 vesicles were studied in cultured astrocytes at single vesicle level with stimulated emission depletion (STED) and total internal reflection fluorescence (TIRF) microscopies. We show that VAMP3 antibodies label small diameter (∼80 nm) vesicles and that VAMP3 vesicles undergo Ca(2+) -independent exo-endocytosis. We also show that this pathway modulates the surface expression of plasma membrane glutamate transporters and the glutamate uptake by astrocytes. Finally, using pharmacological and optogenetic tools, we provide evidence suggesting that the cytosolic cAMP level influences astrocytic VAMP3 vesicle trafficking and glutamate transport. Our results suggest a new role for VAMP3 vesicles in astrocytes.
Kathleen Zylbersztejn - One of the best experts on this subject based on the ideXlab platform.
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astrocyte vamp3 vesicles undergo ca2 independent cycling and modulate glutamate transporter trafficking
The Journal of Physiology, 2015Co-Authors: Karine Herault, Kathleen Zylbersztejn, Martin Oheim, Marcel A. Lauterbach, Marc Guillon, Dongdong LiAbstract:Key points Mouse cortical astrocytes express VAMP3 but not VAMP2. VAMP3 vesicles undergo Ca2+-independent exo- and endocytotic cycling at the plasma membrane. VAMP3 vesicle traffic regulates the recycling of plasma membrane glutamate transporters. cAMP modulates VAMP3 vesicle cycling and glutamate uptake. Abstract Previous studies suggest that small synaptic-like vesicles in astrocytes carry vesicle-associated vSNARE proteins, VAMP3 (cellubrevin) and VAMP2 (synaptobrevin 2), both contributing to the Ca2+-regulated exocytosis of gliotransmitters, thereby modulating brain information processing. Here, using cortical astrocytes taken from VAMP2 and VAMP3 knock-out mice, we find that astrocytes express only VAMP3. The morphology and function of VAMP3 vesicles were studied in cultured astrocytes at single vesicle level with stimulated emission depletion (STED) and total internal reflection fluorescence (TIRF) microscopies. We show that VAMP3 antibodies label small diameter (∼80 nm) vesicles and that VAMP3 vesicles undergo Ca2+-independent exo-endocytosis. We also show that this pathway modulates the surface expression of plasma membrane glutamate transporters and the glutamate uptake by astrocytes. Finally, using pharmacological and optogenetic tools, we provide evidence suggesting that the cytosolic cAMP level influences astrocytic VAMP3 vesicle trafficking and glutamate transport. Our results suggest a new role for VAMP3 vesicles in astrocytes.
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Astrocyte VAMP3 vesicles undergo Ca2+ -independent cycling and modulate glutamate transporter trafficking.
The Journal of Physiology, 2015Co-Authors: Karine Herault, Kathleen Zylbersztejn, Martin Oheim, Marcel A. Lauterbach, Marc Guillon, Nicole RopertAbstract:Mouse cortical astrocytes express VAMP3 but not VAMP2. VAMP3 vesicles undergo Ca(2+) -independent exo- and endocytotic cycling at the plasma membrane. VAMP3 vesicle traffic regulates the recycling of plasma membrane glutamate transporters. cAMP modulates VAMP3 vesicle cycling and glutamate uptake. Previous studies suggest that small synaptic-like vesicles in astrocytes carry vesicle-associated vSNARE proteins, VAMP3 (cellubrevin) and VAMP2 (synaptobrevin 2), both contributing to the Ca(2+) -regulated exocytosis of gliotransmitters, thereby modulating brain information processing. Here, using cortical astrocytes taken from VAMP2 and VAMP3 knock-out mice, we find that astrocytes express only VAMP3. The morphology and function of VAMP3 vesicles were studied in cultured astrocytes at single vesicle level with stimulated emission depletion (STED) and total internal reflection fluorescence (TIRF) microscopies. We show that VAMP3 antibodies label small diameter (∼80 nm) vesicles and that VAMP3 vesicles undergo Ca(2+) -independent exo-endocytosis. We also show that this pathway modulates the surface expression of plasma membrane glutamate transporters and the glutamate uptake by astrocytes. Finally, using pharmacological and optogenetic tools, we provide evidence suggesting that the cytosolic cAMP level influences astrocytic VAMP3 vesicle trafficking and glutamate transport. Our results suggest a new role for VAMP3 vesicles in astrocytes.
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Absence of TI-VAMP/Vamp7 leads to increased anxiety in mice.
Journal of Neuroscience, 2012Co-Authors: Lydia Danglot, Kathleen Zylbersztejn, Maja Petkovic, Maxime Gauberti, Hamid Meziane, Roy Combe, Marie-france Champy, Marie-christine Birling, Guillaume Pavlovic, Jean-charles BizotAbstract:Vesicular (v)- and target (t)-SNARE proteins assemble in SNARE complex to mediate membrane fusion. Tetanus neurotoxin-insensitive vesicular-associated membrane protein (TI-VAMP/VAMP7), a vesicular SNARE expressed in several cell types including neurons, was previously shown to play a major role in exocytosis involved in neurite growth in cultured neurons. Here we generated a complete constitutive knock-out by deleting the exon 3 of Vamp7. Loss of TI-VAMP expression did not lead to any striking developmental or neurological defect. Knock-out mice displayed decreased brain weight and increased third ventricle volume. Axon growth appeared normal in cultured knock-out neurons. Behavioral characterization unraveled that TI-VAMP knock-out was associated with increased anxiety. Our results thus suggest compensatory mechanisms allowing the TI-VAMP knock-out mice to fulfill major developmental processes. The phenotypic traits unraveled here further indicate an unexpected role of TI-VAMP-mediated vesicular traffic in anxiety and suggest a role for TI-VAMP in higher brain functions.
Dongdong Li - One of the best experts on this subject based on the ideXlab platform.
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astrocyte vamp3 vesicles undergo ca2 independent cycling and modulate glutamate transporter trafficking
The Journal of Physiology, 2015Co-Authors: Karine Herault, Kathleen Zylbersztejn, Martin Oheim, Marcel A. Lauterbach, Marc Guillon, Dongdong LiAbstract:Key points Mouse cortical astrocytes express VAMP3 but not VAMP2. VAMP3 vesicles undergo Ca2+-independent exo- and endocytotic cycling at the plasma membrane. VAMP3 vesicle traffic regulates the recycling of plasma membrane glutamate transporters. cAMP modulates VAMP3 vesicle cycling and glutamate uptake. Abstract Previous studies suggest that small synaptic-like vesicles in astrocytes carry vesicle-associated vSNARE proteins, VAMP3 (cellubrevin) and VAMP2 (synaptobrevin 2), both contributing to the Ca2+-regulated exocytosis of gliotransmitters, thereby modulating brain information processing. Here, using cortical astrocytes taken from VAMP2 and VAMP3 knock-out mice, we find that astrocytes express only VAMP3. The morphology and function of VAMP3 vesicles were studied in cultured astrocytes at single vesicle level with stimulated emission depletion (STED) and total internal reflection fluorescence (TIRF) microscopies. We show that VAMP3 antibodies label small diameter (∼80 nm) vesicles and that VAMP3 vesicles undergo Ca2+-independent exo-endocytosis. We also show that this pathway modulates the surface expression of plasma membrane glutamate transporters and the glutamate uptake by astrocytes. Finally, using pharmacological and optogenetic tools, we provide evidence suggesting that the cytosolic cAMP level influences astrocytic VAMP3 vesicle trafficking and glutamate transport. Our results suggest a new role for VAMP3 vesicles in astrocytes.
Pablo A Ortiz - One of the best experts on this subject based on the ideXlab platform.
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decreased surface nkcc2 expression and increased urinary excretion of water and ions in vamp3 knockout mice
The FASEB Journal, 2015Co-Authors: Pablo A Ortiz, Mohammed Ziaul Haque, Paulo S. CaceresAbstract:Trafficking of the NKCC2 cotransporter to the apical surface of thick ascending limbs (TAL) regulates excretion of urine, ions and blood pressure. We showed that vesicle fusion proteins VAMP2 and VAMP3 mediate NKCC2 exocytosis to the TAL plasma membrane. VAMP3 also mediates NKCC2 expression. We showed that VAMP3-/- mice have lower blood pressure in normal and low-Na diet. However, it is not know whether surface NKCC2 is decreased in VAMP3-/- mice and the contribution of VAMP3 to renal function is unclear. We hypothesize that VAMP3-/- mice have decreased surface NKCC2 expression and increased excretion of water and ions. First we studied VAMP3-/- kidney histology and we found no abnormalities. Next, we measured surface NKCC2 in TAL suspensions by surface biotinylation. Surface NKCC2 was decreased by 46 ± 4% compared to wt. We also observed 23% higher urine volume (wt= 1.3 ± 0.1 ml/day vs VAMP3-/- = 1.6 ± 0.2 ml/day) and lower urine osmolality (wt= 3433 ± 166 mOsm vs VAMP3-/- = 2924 ± 131 mOsm). UNa, UK and...
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vesicle associated membrane protein 2 VAMP2 but not vamp3 mediates camp stimulated trafficking of the renal na k 2cl co transporter nkcc2 in thick ascending limbs
Journal of Biological Chemistry, 2014Co-Authors: Mariela Mendez, Pablo A Ortiz, Paulo S. CaceresAbstract:In the kidney, epithelial cells of the thick ascending limb (TAL) reabsorb NaCl via the apical Na+/K+/2Cl− co-transporter NKCC2. Steady-state surface NKCC2 levels in the apical membrane are maintained by a balance between exocytic delivery, endocytosis, and recycling. cAMP is the second messenger of hormones that enhance NaCl absorption. cAMP stimulates NKCC2 exocytic delivery via protein kinase A (PKA), increasing steady-state surface NKCC2. However, the molecular mechanism involved has not been studied. We found that several members of the SNARE family of membrane fusion proteins are expressed in TALs. Here we report that NKCC2 co-immunoprecipitates with VAMP2 in rat TALs, and they co-localize in discrete domains at the apical surface. cAMP stimulation enhanced VAMP2 exocytic delivery to the plasma membrane of renal cells, and stimulation of PKA enhanced VAMP2-NKCC2 co-immunoprecipitation in TALs. In vivo silencing of VAMP2 but not VAMP3 in TALs blunted cAMP-stimulated steady-state surface NKCC2 expression and completely blocked cAMP-stimulated NKCC2 exocytic delivery. VAMP2 was not involved in constitutive NKCC2 delivery. We concluded that VAMP2 but not VAMP3 selectively mediates cAMP-stimulated NKCC2 exocytic delivery and surface expression in TALs. We also demonstrated that cAMP stimulation enhances VAMP2 exocytosis and promotes VAMP2 interaction with NKCC2.
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Vesicle-associated membrane protein 2 (VAMP2) but Not VAMP3 mediates cAMP-stimulated trafficking of the renal Na+-K+-2Cl- co-transporter NKCC2 in thick ascending limbs.
Journal of Biological Chemistry, 2014Co-Authors: Paulo S. Caceres, Mariela Mendez, Pablo A OrtizAbstract:In the kidney, epithelial cells of the thick ascending limb (TAL) reabsorb NaCl via the apical Na+/K+/2Cl− co-transporter NKCC2. Steady-state surface NKCC2 levels in the apical membrane are maintained by a balance between exocytic delivery, endocytosis, and recycling. cAMP is the second messenger of hormones that enhance NaCl absorption. cAMP stimulates NKCC2 exocytic delivery via protein kinase A (PKA), increasing steady-state surface NKCC2. However, the molecular mechanism involved has not been studied. We found that several members of the SNARE family of membrane fusion proteins are expressed in TALs. Here we report that NKCC2 co-immunoprecipitates with VAMP2 in rat TALs, and they co-localize in discrete domains at the apical surface. cAMP stimulation enhanced VAMP2 exocytic delivery to the plasma membrane of renal cells, and stimulation of PKA enhanced VAMP2-NKCC2 co-immunoprecipitation in TALs. In vivo silencing of VAMP2 but not VAMP3 in TALs blunted cAMP-stimulated steady-state surface NKCC2 expression and completely blocked cAMP-stimulated NKCC2 exocytic delivery. VAMP2 was not involved in constitutive NKCC2 delivery. We concluded that VAMP2 but not VAMP3 selectively mediates cAMP-stimulated NKCC2 exocytic delivery and surface expression in TALs. We also demonstrated that cAMP stimulation enhances VAMP2 exocytosis and promotes VAMP2 interaction with NKCC2.
