N-ETHYLMALEIMIDE-SENSITIVE FACTOR ACTS AT A PROFUSION ATP-DEPENDENT STEP IN CA2-ACTIVATED EXOCYTOSIS()

An ATP-dependent activity of NSF (N-ethylmaleimide-sensitive factor) t hat rearranges soluble NSF attachment protein (SNAP) receptor (SNARE) protein complexes was proposed to be the driving force for membrane fu sion. The Ca2+-activated fusion of secretory vesicles with the plasma membrane in permeable PC12 cells requires ATP; however, the ATP requir ement is for a priming step that precedes the Ca2+-triggered fusion re action. While phosphoinositide phosphorylation is a key reaction requi red for priming, additional ATP-dependent reactions are also necessary . Here we report that the NSF-catalyzed rearrangement of SNARE protein complexes occurs during ATP-dependent priming, NSF with alpha-SNAP (s oluble NSF attachment protein) were required for ATP-dependent priming but not Ca2+-triggered fusion, indicating that NSF acts at an ATP-dep endent prefusion step rather than at fusion itself. NSF-catalyzed acti vation of SNARE proteins may reorganize membranes to generate a vesicl e-plasma membrane prefusion intermediate that is poised for conversion to full fusion by Ca2+-dependent mechanisms.