Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0848283 (rundown)
 502 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We examined the role of SNAPs, soluble proteins that attach N-ethylmaleimide-sensitive factor (NSF), in regulating exocytosis in single rat adrenal chromaffin cells. Whole-cell dialysis of Ca2+-buffered solution or photolysis of caged-Ca2+ was used to manipulate cytosolic Ca2+ concentration ([Ca2+]i), whereas exocytosis was measured via carbon fiber amperometry or membrane capacitance. Buffering [Ca2+]i to approximately 170 nm produced a mean rate of exocytosis of approximately one amperometric event per minute. Including alpha-SNAP (60 or 500 nm) in the intracellular solution dramatically increased the mean rate of exocytosis. The stimulatory action of alpha-SNAP requires ATP hydrolysis mediated via NSF, because this action was blocked by intracellular dialysis of ATP-gamma-S (2 mm) and could not be mimicked by a mutant alpha-SNAP that does not stimulate the ATPase activity of NSF. This action of alpha-SNAP was significant only at [Ca2+]i between 100 and 300 nm and was not shared by beta-SNAP (500 nm), suggesting that alpha-SNAP enhanced a component of exocytosis that is regulated by a high-affinity Ca2+ sensor. In cells dialyzed with both alpha- and beta-SNAP, the rate of exocytosis was smaller than that produced by alpha-SNAP alone, suggesting that alpha- and beta-SNAP interact competitively. Although only alpha-SNAP stimulated exocytosis at [Ca2+]i between 100 and 300 nm, both alpha- and beta-SNAP isoforms equally slowed the time-dependent rundown of the exocytic response. Our results indicate that alpha- and beta-SNAP have different actions in exocytosis. Thus, the ratio of different isoforms of SNAPs can determine release probability at the levels of [Ca2+]i that are involved in regulation of exocytosis.
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PMID:Differential regulation of exocytosis by alpha- and beta-SNAPs. 1175 88

We investigated the functional and structural implications of SNAP25 having two SNARE motifs (SN1 and SN2). A membrane-bound, intramolecular FRET probe was constructed to report on the folding of N-terminal SN1 and C-terminal SN2 in living cells. Membrane-bound constructs containing either or both SNARE motifs were also singly labeled with donor or acceptor fluorophores. Interaction of probes with other SNAREs was monitored by the formation of SDS-resistant complexes and by changes in FRET measured in vitro using spectroscopy and in the plasma membrane of living cells using TIRF microscopy. The probes formed the predicted SDS-resistant SNARE complexes. FRET measurements revealed that syntaxin induced a close association of the N-termini of SN1 and SN2. This association required that the SNARE motifs reside in the same molecule. Unexpectedly, the syntaxin-induced FRET was prevented by VAMP. Both full-length SNAP25 constructs and the combination of its separated, membrane-bound constituent chains supported secretion in permeabilized chromaffin cells that had been allowed to rundown. However, only full-length SNAP25 constructs enabled robust secretion from intact cells or permeabilized cells before rundown. The experiments suggest that the bidentate structure permits specific conformations in complexes with syntaxin and VAMP and facilitates the function of SN1 and SN2 in exocytosis.
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PMID:The structural and functional implications of linked SNARE motifs in SNAP25. 1859 34