Real-time visualization of complexin during single exocytic events.
Understanding the fundamental role of soluble NSF attachment protein receptor (SNARE) complexes in membrane fusion requires knowledge of the spatiotemporal dynamics of their assembly. We visualized complexin (cplx), a cytosolic protein that binds assembled SNARE complexes, during single exocytic events in live cells. We found that cplx appeared briefly during full fusion. However, a truncated version of cplx containing only the SNARE-complex binding region persisted at fusion sites for seconds and caused fusion to be transient. Resealing pores with the mutant cplx only partially released transmitter and lipid probes, indicating that the pores are narrow and not purely lipidic in structure. Depletion of cplx similarly caused secretory cargo to be retained. These data suggest that cplx is recruited at a late step in exocytosis and modulates fusion pores composed of SNARE complexes.
An
Seong J
SJ
Department of Cellular and Molecular Physiology, Yale University School of Medicine, New Haven, Connecticut, USA. seong.an@yale.edu
Grabner
Chad P
CP
Zenisek
David
D
D033942:Adaptor Proteins, Vesicular Transport / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D004298:Dopamine / Q000378:metabolism; D005089:Exocytosis / Q000235:genetics / Q000502:physiology*; D005786:Gene Expression Regulation / Q000187:drug effects / Q000235:genetics; D049452:Green Fluorescent Proteins / Q000235:genetics; D006801:Humans; D018613:Microscopy, Confocal / Q000379:methods; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism*; D009478:Neuropeptide Y / Q000235:genetics / Q000378:metabolism; D016716:PC12 Cells; D038761:Photobleaching; D011485:Protein Binding; D011726:Pyridinium Compounds / Q000378:metabolism; D000644:Quaternary Ammonium Compounds / Q000378:metabolism; D034622:RNA Interference / Q000502:physiology; D051381:Rats; D050600:SNARE Proteins / Q000235:genetics / Q000378:metabolism; D010959:Tissue Plasminogen Activator / Q000378:metabolism; D014162:Transfection / Q000379:methods
D033942:Adaptor Proteins, Vesicular Transport; C092350:FM 4-64; D009419:Nerve Tissue Proteins; D009478:Neuropeptide Y; D011726:Pyridinium Compounds; D000644:Quaternary Ammonium Compounds; D050600:SNARE Proteins; C095993:complexin II; D049452:Green Fluorescent Proteins; D010959:Tissue Plasminogen Activator; D004298:Dopamine
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