Botulinum neurotoxin A and neurotoxin E cleavage products of synaptosome-associated protein of 25 kd exhibit distinct actions on pancreatic islet beta-cell Kv2.1 channel gating

• Published in: Pancreas Vol. 36; no. 1; p. 10
• Main Authors: He, Yan, Elias, Chadwick L, Huang, Ya-Chi, Gao, Xiaodong, Leung, Yuk-Man, Kang, Youhou, Xie, Huanli, Chaddock, John A, Tsushima, Robert G, Gaisano, Herbert Y
• Format: Journal Article
• Language: English
• Published: United States 01.01.2008
• Subjects: Animals; Blotting, Western; Botulinum Toxins - metabolism; Botulinum Toxins, Type A - metabolism; Cell Line; Glutathione Transferase - genetics; Insulin - metabolism; Insulin Secretion; Insulin-Secreting Cells - chemistry; Insulin-Secreting Cells - physiology; Ion Channel Gating - drug effects; Microscopy, Confocal; Patch-Clamp Techniques; Peptide Fragments - genetics; Peptide Fragments - pharmacology; Rats; Recombinant Fusion Proteins - genetics; Shab Potassium Channels - drug effects; Shab Potassium Channels - physiology; Synaptosomal-Associated Protein 25 - genetics; Synaptosomal-Associated Protein 25 - metabolism; Synaptosomal-Associated Protein 25 - pharmacology; Transfection
• ISSN: 1536-4828
• DOI: 10.1097/mpa.0b013e31812eee28

Synaptosome-associated protein of 25 kd (SNAP-25) regulates pancreatic islet beta-cell-delayed rectifier K channels (Kv2.1) in addition to insulin exocytosis. Botulinum neurotoxin A (BoNT/A) and E (BoNT/E) cleavage and presumed deletion of SNAP-25 have been used to examine SNAP-25 function. We hypothesized that proteolytic products of SNAP-25 (206 amino acids) resulting from BoNT/A and BoNT/E cleavage, SNAP-25(1-197) and SNAP-25(1-180), have independent actions on beta-cell Kv gating. We examined by confocal microscopy and immunoblotting BoNT/A and BoNT/E cleavage of SNAP-25 to these N-terminal fragments, and the consequent effects of these BoNTs and SNAP-25 fragments on Kv currents in rat beta cells and MIN6 cells by patch clamp electrophysiology. Confocal microscopy and immunoblotting showed that MIN6 cells transfected with BoNT/A or BoNT/E generated SNAP-25(1-197) and SNAP-25(1-180) fragments that were retained in the cytosol. Both BoNTs caused increased rate of channel activation and slowed channel inactivation, mimicked by these SNAP-25 fragments, but not full-length SNAP-25. These SNAP-25 fragments potentiated tetraethylammonium block of beta-cell Kv currents. BoNT/A or BoNT/E treatment of beta cells generates N-terminal SNAP-25 fragments that are retained in beta cells to directly influence Kv channel gating in a manner distinct from full-length SNAP-25, contributing to overall actions of these BoNTs on insulin secretion.