Cysteine string protein is required for calcium secretion coupling of evoked neurotransmission in drosophila but not for vesicle recycling.
The entire deletion of the cysteine string protein (CSP) gene causes a temperature-sensitive (ts) block of evoked neurotransmission in Drosophila. CSP has been found to interact in vitro with the clathrin-uncoating ATPase HSC70, suggesting a potential role of CSP in vesicle recycling. Using FM1-43 imaging, we analyzed whether the ts block of neurotransmission in csp mutants is caused by a defect in vesicle exocytosis or vesicle recycling. We determined that FM1-43-labeled synaptic boutons of csp mutant neuromuscular junctions fail to destain at 32 degrees C after K+ depolarization, and that FM1-43 dye uptake cannot be evoked by K+ stimulation at 32 degrees C. However, when we stimulated dye uptake independent of depolarization by using black widow spider venom (BWSV), we observed endocytotic uptake of FM1-43. This suggests that endocytosis exhibits no primary ts defect. In addition, we found no ts defect of vesicle recycling at 32 degrees C that would correlate with the ts block of neurotransmission. We also discovered that BWSV and the calcium ionophore calcimycin stimulate FM1-43 destaining and quantal release in csp mutants at 32 degrees C when depolarization fails to evoke any response. The wild-type-like, calcimycin-induced response in csp null mutants indicates that some aspect of the depolarization-dependent calcium signaling pathway must be impaired, either calcium entry, calcium action, or both. Collectively, our results indicate that the csp mutation affects calcium secretion coupling of evoked exocytosis but not vesicle recycling. This supports the hypothesis that CSP links synaptic vesicles to calcium secretion coupling.
Ranjan
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Department of Neuroscience, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania 19104-6074, USA.
Bronk
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Zinsmaier
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KE
D000818:Animals; D001740:Black Widow Spider; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D004330:Drosophila / Q000235:genetics; D004594:Electrophysiology; D005071:Evoked Potentials / Q000187:drug effects / Q000502:physiology; D005089:Exocytosis / Q000502:physiology; D050956:HSP40 Heat-Shock Proteins; D007476:Ionophores / Q000494:pharmacology; D008565:Membrane Proteins; D009154:Mutation / Q000502:physiology; D009419:Nerve Tissue Proteins / Q000235:genetics*; D009474:Neurons / Q000166:cytology / Q000502:physiology; D013111:Spider Venoms / Q000494:pharmacology; D009435:Synaptic Transmission / Q000187:drug effects / Q000502:physiology*; D013572:Synaptic Vesicles / Q000187:drug effects / Q000378:metabolism*; D013696:Temperature
D050956:HSP40 Heat-Shock Proteins; D007476:Ionophores; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D013111:Spider Venoms; C088460:cysteine string protein; D000001:Calcimycin; D002118:Calcium; C038599:black widow spider venom