Modulation of Kv2.1 channel gating and TEA sensitivity by distinct domains of SNAP-25.
Distinct domains within the SNARE (soluble N-ethylmaleimide-sensitive fusion protein attachment protein receptor) proteins, STX1A (syntaxin 1A) and SNAP-25 (synaptosome-associated protein-25 kDa), regulate hormone secretion by their actions on the cell's exocytotic machinery, as well as voltage-gated Ca2+ and K+ channels. We examined the action of distinct domains within SNAP-25 on Kv2.1 (voltage gated K+ 2.1) channel gating. Dialysis of N-terminal SNAP-25 domains, S197 (SNAP-25(1-197)) and S180 (SNAP-25(1-180)), but not S206 (full-length SNAP-25(1-206)) increased the rate of Kv2.1 channel activation and slowed channel inactivation. Remarkably, these N-terminal SNAP-25 domains, acting on the Kv2.1 cytoplasmic N-terminus, potentiated the external TEA (tetraethylammonium)-mediated block of Kv2.1. To further examine whether these are effects of the channel pore domain, internal K+ was replaced with Na+ and external K+ was decreased from 4 to 1 mM, which decreased the IC50 of the TEA block from 6.8+/-0.9 mM to >100 mM. Under these conditions S180 completely restored TEA sensitivity (7.9+/-1.5 mM). SNAP-25 C-terminal domains, SNAP-25(198-206) and SNAP-25(181-197), had no effect on Kv2.1 gating kinetics. We conclude that different domains within SNAP-25 can form distinct complexes with Kv2.1 to execute a fine allosteric regulation of channel gating and the architecture of the outer pore structure in order to modulate cell excitability.
He
Yan
Y
School of Public Health and Family Medicine, Capital University of Medical Sciences, Beijing, China.
Kang
Youhou
Y
Leung
Yuk-Man
YM
Xia
Fuzhen
F
Gao
Xiaodong
X
Xie
Huanli
H
Gaisano
Herbert Y
HY
Tsushima
Robert G
RG
D000494:Allosteric Regulation; D000818:Animals; D018726:Anti-Dyskinesia Agents / Q000378:metabolism / Q000494:pharmacology; D001905:Botulinum Toxins / Q000378:metabolism / Q000494:pharmacology; D019274:Botulinum Toxins, Type A / Q000378:metabolism / Q000494:pharmacology; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D004594:Electrophysiology; D006801:Humans; D015640:Ion Channel Gating / Q000187:drug effects; D010455:Peptides / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012680:Sensitivity and Specificity; D051669:Shab Potassium Channels / Q000235:genetics / Q000378:metabolism*; D050825:Synaptosomal-Associated Protein 25 / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D019789:Tetraethylammonium / Q000378:metabolism / Q000494:pharmacology*; D014162:Transfection
D018726:Anti-Dyskinesia Agents; C496991:Kcnb1 protein, rat; D010455:Peptides; D011993:Recombinant Fusion Proteins; D051669:Shab Potassium Channels; C494210:Snap25 protein, rat; D050825:Synaptosomal-Associated Protein 25; D019789:Tetraethylammonium; D001905:Botulinum Toxins; D019274:Botulinum Toxins, Type A; C096326:botulinum toxin type E