title: SNARE proteins are critical for regulated exocytosis of ECP from human eosinophils.
abstract: The SNARE hypothesis, describing a protein assembly-disassembly pathway, was recently proposed for the sequential steps of synaptic vesicle docking, activation and fusion. To determine if SNARE proteins are involved in regulated exocytosis in eosinophils, the presence and functional role of SNAREs was examined in human blood eosinophils. Immunoblotting, subcellular fractionation, and immunocytochemistry documented that vesicle-associated membrane protein-2 (VAMP-2), a vesicle-SNARE, was expressed in human eosinophils. Syntaxin 4 and SNAP-25 were also detected. Sequencing of cloned RT-PCR products amplified from a domain conserved among VAMP isoforms revealed identity only to VAMP-2 but not to VAMP-1 or cellubrevin. Functional experiments revealed that tetanus toxin pretreatment, which cleaved VAMP-2 in eosinophils, significantly inhibited both IgE receptor- and phorbol ester-mediated exocytosis of eosinophil cationic protein (ECP) from streptolysin-O-permeabilized eosinophils. Thus, these results strongly suggest a critical role of SNAREs in regulated exocytosis in eosinophils.
authors.affiliation: Allergy and Lung Research Laboratory, University Hospital of Aarhus, DK-8000 Aarhus C, Denmark.
authors.forename: H J
authors.initials: HJ
authors.lastname: Hoffmann
authors.forename: T
authors.initials: T
authors.lastname: Bjerke
authors.forename: M
authors.initials: M
authors.lastname: Karawajczyk
authors.forename: R
authors.initials: R
authors.lastname: Dahl
authors.forename: M A
authors.initials: MA
authors.lastname: Knepper
authors.forename: S
authors.initials: S
authors.lastname: Nielsen
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001798:Blood Proteins / Q000378:metabolism*; D015153:Blotting, Western; D001921:Brain / Q000502:physiology*; D047091:Eosinophil Granule Proteins; D004804:Eosinophils / Q000502:physiology*; D005089:Exocytosis; D006801:Humans; D008565:Membrane Proteins / Q000097:blood / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D020033:Protein Isoforms / Q000097:blood / Q000235:genetics / Q000378:metabolism; D050683:R-SNARE Proteins; D051381:Rats; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012260:Ribonucleases; D050600:SNARE Proteins; D013572:Synaptic Vesicles / Q000502:physiology; D033921:Vesicular Transport Proteins
chemical_list: D001798:Blood Proteins; D047091:Eosinophil Granule Proteins; D008565:Membrane Proteins; D020033:Protein Isoforms; D050683:R-SNARE Proteins; D050600:SNARE Proteins; D033921:Vesicular Transport Proteins; D012260:Ribonucleases