A novel tetanus neurotoxin-insensitive vesicle-associated membrane protein in SNARE complexes of the apical plasma membrane of epithelial cells.

The importance of soluble N-ethyl maleimide (NEM)-sensitive fusion protein (NSF) attachment protein (SNAP) receptors (SNAREs) in synaptic vesicle exocytosis is well established because it has been demonstrated that clostridial neurotoxins (NTs) proteolyze the vesicle SNAREs (v-SNAREs) vesicle-associated membrane protein (VAMP)/brevins and their partners, the target SNAREs (t-SNAREs) syntaxin 1 and SNAP25. Yet, several exocytotic events, including apical exocytosis in epithelial cells, are insensitive to numerous clostridial NTs, suggesting the presence of SNARE-independent mechanisms of exocytosis. In this study we found that syntaxin 3, SNAP23, and a newly identified VAMP/brevin, tetanus neurotoxin (TeNT)-insensitive VAMP (TI-VAMP), are insensitive to clostridial NTs. In epithelial cells, TI-VAMP-containing vesicles were concentrated in the apical domain, and the protein was detected at the apical plasma membrane by immunogold labeling on ultrathin cryosections. Syntaxin 3 and SNAP23 were codistributed at the apical plasma membrane where they formed NEM-dependent SNARE complexes with TI-VAMP and cellubrevin. We suggest that TI-VAMP, SNAP23, and syntaxin 3 can participate in exocytotic processes at the apical plasma membrane of epithelial cells and, more generally, domain-specific exocytosis in clostridial NT-resistant pathways.

[1]  T. Südhof,et al.  Synaptic vesicle membrane fusion complex: action of clostridial neurotoxins on assembly. , 1994, The EMBO journal.

[2]  J. Gordon,et al.  A strategy for isolation of cDNAs encoding proteins affecting human intestinal epithelial cell growth and differentiation: characterization of a novel gut-specific N-myristoylated annexin , 1992, The Journal of cell biology.

[3]  M. Bennett,et al.  Distinct cellular locations of the syntaxin family of proteins in rat pancreatic acinar cells. , 1996, Molecular biology of the cell.

[4]  F. Benfenati,et al.  Tetanus and Botulinum Neurotoxins Are Zinc Proteases Specific for Components of the Neuroexocytosis Apparatus a , 1994, Annals of the New York Academy of Sciences.

[5]  P. De Camilli,et al.  Tetanus toxin-mediated cleavage of cellubrevin impairs exocytosis of transferrin receptor-containing vesicles in CHO cells , 1994, The Journal of cell biology.

[6]  P. D. Smith,et al.  Structural evolution of the annexin supergene family. , 1994, Trends in genetics : TIG.

[7]  P. Roche,et al.  Identification of a Novel Syntaxin- and Synaptobrevin/VAMP-binding Protein, SNAP-23, Expressed in Non-neuronal Tissues* , 1996, The Journal of Biological Chemistry.

[8]  William J. Norton.  Part III , 1916, Transactions of the American Institute of Electrical Engineers.

[9]  R. Jahn,et al.  Botulinum neurotoxin C1 blocks neurotransmitter release by means of cleaving HPC‐1/syntaxin. , 1993, The EMBO journal.

[10]  F. Benfenati,et al.  Tetanus and botulinum-B neurotoxins block neurotransmitter release by proteolytic cleavage of synaptobrevin , 1992, Nature.

[11]  C. Hopkins,et al.  Endocytic pathways in polarized Caco-2 cells: identification of an endosomal compartment accessible from both apical and basolateral surfaces , 1990, The Journal of cell biology.

[12]  P. De Camilli,et al.  Cellubrevin and synaptobrevins: similar subcellular localization and biochemical properties in PC12 cells , 1995, The Journal of cell biology.

[13]  H. Schägger,et al.  Tricine-sodium dodecyl sulfate-polyacrylamide gel electrophoresis for the separation of proteins in the range from 1 to 100 kDa. , 1987, Analytical biochemistry.

[14]  Giampietro Schiavo,et al.  SNARE motif and neurotoxins , 1994, Nature.

[15]  T. Weimbs,et al.  Apical targeting in polarized epithelial cells: There's more afloat than rafts. , 1997, Trends in cell biology.

[16]  J. Rothman,et al.  Vesicular transport between the endoplasmic reticulum and the Golgi stack requires the NEM-sensitive fusion protein , 1989, Nature.

[17]  Thomas C. Südhof,et al.  Botulinum neurotoxin A selectively cleaves the synaptic protein SNAP-25 , 1993, Nature.

[18]  R. Parton,et al.  Annexin XIIIb: a novel epithelial specific annexin is implicated in vesicular traffic to the apical plasma membrane , 1995, The Journal of cell biology.

[19]  T. Weimbs,et al.  Differential localization of syntaxin isoforms in polarized Madin-Darby canine kidney cells. , 1996, Molecular biology of the cell.

[20]  A. Klip,et al.  Tissue distribution of SNAP-23 and its subcellular localization in 3T3-L1 cells. , 1997, Biochemical and biophysical research communications.

[21]  R. Scheller,et al.  Botulinum Neurotoxin Type C Cleaves a Single Lys-Ala Bond within the Carboxyl-terminal Region of Syntaxins (*) , 1995, The Journal of Biological Chemistry.

[22]  G. Schiavo,et al.  Common and distinct fusion proteins in axonal growth and transmitter release , 1996, The Journal of comparative neurology.

[23]  S. Nielsen,et al.  Syntaxin-4 is localized to the apical plasma membrane of rat renal collecting duct cells: possible role in aquaporin-2 trafficking. , 1996, The Journal of clinical investigation.

[24]  B. Dasgupta,et al.  N-Ethylmaleimide-sensitive Factor Acts at a Prefusion ATP-dependent Step in Ca2+-activated Exocytosis* , 1996, The Journal of Biological Chemistry.

[25]  H. Schulman,et al.  Nitric Oxide Modulates Synaptic Vesicle Docking/Fusion Reactions , 1996, Neuron.

[26]  A. Le Bivic,et al.  Human syntaxin 3 is localized apically in human intestinal cells. , 1997, Journal of cell science.

[27]  R. Scheller,et al.  A New Syntaxin Family Member Implicated in Targeting of Intracellular Transport Vesicles* , 1996, Journal of Biological Chemistry.

[28]  Paul Tempst,et al.  SNAP receptors implicated in vesicle targeting and fusion , 1993, Nature.

[29]  R. Scheller,et al.  Protein-protein interactions contributing to the specificity of intracellular vesicular trafficking. , 1994, Science.

[30]  J. Changeux,et al.  Golgi apparatus in chick skeletal muscle: changes in its distribution during end plate development and after denervation. , 1989, Proceedings of the National Academy of Sciences of the United States of America.

[31]  E. Ikonen,et al.  Functional rafts in cell membranes , 1997, Nature.

[32]  P. Novick,et al.  Homologs of the synaptobrevin/VAMP family of synaptic vesicle proteins function on the late secretory pathway in S. cerevisiae , 1993, Cell.

[33]  A. Lupas,et al.  Predicting coiled coils from protein sequences , 1991, Science.

[34]  L. Johannes,et al.  Exocytosis: SNAREs drum up! , 1998, The European journal of neuroscience.

[35]  R. Jahn,et al.  Clostridial neurotoxins: new tools for dissecting exocytosis. , 1994, Trends in cell biology.

[36]  Mark K. Bennett,et al.  A protein assembly-disassembly pathway in vitro that may correspond to sequential steps of synaptic vesicle docking, activation, and fusion , 1993, Cell.

[37]  I. Pastan,et al.  Identification of two lysosomal membrane glycoproteins , 1985, The Journal of cell biology.

[38]  R. Kelly,et al.  Mutational analysis of VAMP domains implicated in Ca2+‐induced insulin exocytosis. , 1996, The EMBO journal.

[39]  R. Scheller,et al.  The Mammalian Brain rsec6/8 Complex , 1996, Neuron.

[40]  R. Scheller,et al.  The syntaxin family of vesicular transport receptors , 1993, Cell.

[41]  Jonathan Pevsner,et al.  Specificity and regulation of a synaptic vesicle docking complex , 1994, Neuron.

[42]  T. Südhof,et al.  Cellubrevin is a ubiquitous tetanus-toxin substrate homologous to a putative synaptic vesicle fusion protein , 1993, Nature.

[43]  H. Hauri,et al.  Sorting of endogenous plasma membrane proteins occurs from two sites in cultured human intestinal epithelial cells (Caco-2) , 1990, Cell.

[44]  M. D'Esposito,et al.  A synaptobrevin–like gene in the Xq28 pseudoautosomal region undergoes X inactivation , 1996, Nature Genetics.

[45]  H. Pollard,et al.  Annexins: the problem of assessing the biological role for a gene family of multifunctional calcium- and phospholipid-binding proteins. , 1994, Biochimica et biophysica acta.

[46]  E. Ikonen,et al.  Different requirements for NSF, SNAP, and Rab proteins in apical and basolateral transport in MDCK cells , 1995, Cell.

[47]  C. Montecucco,et al.  Structural Determinants of the Specificity for Synaptic Vesicle-associated Membrane Protein/Synaptobrevin of Tetanus and Botulinum Type B and G Neurotoxins* , 1996, The Journal of Biological Chemistry.

[48]  J. Pevsner,et al.  Human syntaxin 7: a Pep12p/Vps6p homologue implicated in vesicle trafficking to lysosomes. , 1997, Gene.