Identification of SNAREs involved in regulated exocytosis in the pancreatic acinar cell.
The molecular basis of exocytotic membrane fusion in the pancreatic acinar cell was investigated using an in vitro assay that measures both zymogen granule-plasma membrane fusion and granule-granule fusion. These two fusion events were differentially sensitive to Ca(2+), suggesting that they are controlled by different Ca(2+)-sensing mechanisms. Botulinum neurotoxin C (BoNT/C) treatment of the plasma membranes caused cleavage of syntaxin 2, the apical isoform of this Q-SNARE, but did not affect syntaxin 4, the basolateral isoform. BoNT/C also cleaved syntaxin 3, the zymogen granule isoform. BoNT/C treatment of plasma membranes abolished granule-plasma membrane fusion, whereas toxin treatment of the granules reduced granule-plasma membrane fusion and abolished granule-granule fusion. Tetanus toxin cleaved granule-associated synaptobrevin 2 but caused only a small reduction in both granule-plasma membrane fusion and granule-granule fusion. Our results indicate that syntaxin 2 is the isoform that mediates fusion between zymogen granules and the apical plasma membrane of the acinar cell. Syntaxin 3 mediates granule-granule fusion, which might be involved in compound exocytosis. In contrast, the major R-SNARE on the zymogen granule remains to be identified.
Hansen
N J
NJ
Department of Pharmacology, University of Cambridge, Cambridge, CB2 1QJ, United Kingdom.
Antonin
W
W
Edwardson
J M
JM
D000595:Amino Acid Sequence; D001905:Botulinum Toxins / Q000494:pharmacology; D002118:Calcium / Q000494:pharmacology; D002413:Cations, Divalent / Q000494:pharmacology; D002451:Cell Compartmentation; D002458:Cell Fractionation; D002462:Cell Membrane / Q000737:chemistry / Q000502:physiology; D003594:Cytoplasmic Granules / Q000737:chemistry / Q000502:physiology; D004792:Enzyme Precursors; D005089:Exocytosis / Q000502:physiology*; D008274:Magnesium / Q000494:pharmacology; D008561:Membrane Fusion / Q000187:drug effects / Q000502:physiology; D008565:Membrane Proteins / Q000302:isolation & purification* / Q000378:metabolism; D008673:Metals, Alkaline Earth / Q000494:pharmacology; D008969:Molecular Sequence Data; D010179:Pancreas / Q000166:cytology / Q000502:physiology*; D020033:Protein Isoforms / Q000378:metabolism; D050765:Qa-SNARE Proteins; D050600:SNARE Proteins; D033921:Vesicular Transport Proteins
D002413:Cations, Divalent; D004792:Enzyme Precursors; D008565:Membrane Proteins; D008673:Metals, Alkaline Earth; D020033:Protein Isoforms; D050765:Qa-SNARE Proteins; D050600:SNARE Proteins; D033921:Vesicular Transport Proteins; D001905:Botulinum Toxins; C096324:botulinum toxin type C; D008274:Magnesium; D002118:Calcium