Non-conducting function of the Kv2.1 channel enables it to recruit vesicles for release in neuroendocrine and nerve cells

Abstract

SummaryRegulation of exocytosis by voltage-gated K + channels has classically been viewed as inhibition mediated by K + fluxes. We recently identified a new role for Kv2.1 in facilitating vesicle release from neuroendocrine cells, which is independent of K + flux. Here, we show that Kv2.1-induced facilitation of release is not restricted to neuroendocrine cells, but also occurs in the somatic-vesicle release from dorsal-root-ganglion neurons and is mediated by direct association of Kv2.1 with syntaxin. We further show in adrenal chromaffin cells that facilitation induced by both wild-type and non-conducting mutant Kv2.1 channels in response to long stimulation persists during successive stimulation, and can be attributed to an increased number of exocytotic events and not to changes in single-spike kinetics. Moreover, rigorous analysis of the pools of released vesicles reveals that Kv2.1 enhances the rate of vesicle recruitment during stimulation with high Ca 2+ , without affecting the size of the readily releasable vesicle pool. These findings place a voltage-gated K + channel among the syntaxin-binding proteins that directly regulate pre-fusion steps in exocytosis.

Pancreatic β-cell prosurvival effects of the incretin hormones involve post-translational modification of Kv2.1 delayed rectifier channels

“…Transfected INS-1 b-cells were treated with GIP or GLP-1 and solubilized extracts immunoprecipitated with phosphoserine/threonine or acetyl-lysine antibodies, followed by immunoblotting with Kv2.1 antibody. GIP (1-42, 100 nM) and GLP-1 (7-36, 100 nM) increased phosphorylation (Figures 3a and b) and acetylation (Figures 3c and d) of Kv2.1, whereas truncated forms of the peptides, GIP (19)(20)(21)(22)(23)(24)(25)(26)(27)(28)(29)(30) and GLP-1 , that do not exhibit insulinotropic activity, had no effect. These results indicate that the incretin hormones are capable of increasing both phosphorylation and acetylation of Kv2.1 protein in pancreatic b-cells.…”

“…12,23 Additionally, the cytoplasmic C-terminus of Kv2.1 binds to syntaxin 1A in b-cells and their interaction is involved in channel gating, trafficking and insulin exocytosis. 27,28 Multifunctional roles have also been shown in other cell types, and Kv2.1 facilitation of secretory vesicle recruitment in neuroendocrine cells, 29 and the clustering of cell surface Kv2.1 channels in hippocampal neurons 30 were shown to be independent from K þ conductance. In view of its additional pro-apoptotic role in neurons, we focused on the potential role of Kv2.1 in b-cell apoptosis and the ability of GIP and GLP-1 to modulate its action.…”

“…(e) Effects of GIP and GLP-1 on Kv2.1 surface expression. Human islet cells were treated with 100 nM of indicated peptides, GIP (1-42), GIP analog (19)(20)(21)(22)(23)(24)(25)(26)(27)(28)(29)(30), GLP-1 (7-36) and GLP-1 analog (9-36) for 1 h. Proteinase K was applied for 30 min, and cell lysates were analyzed for Kv 2.1 by SDS-PAGE and IB as described in Proteinase K digestion experiments of Materials and methods. The arrow labeled S indicates the immunoblot position of surface Kv2.1, and the arrow labeled I indicates the intracellular Kv2.1.…”

The voltage-dependent potassium channel subunit Kv2.1 regulates insulin secretion from rodent and human islets independently of its electrical function

“…Only recently has the converse been examined, namely the question of whether this interaction controls exocytosis itself. Increasing Kv2.1 enhances exocytosis in PC12 and bovine chromaffin cells due to an interaction between the channel C-terminus (specifically, the C1a domain) and syntaxin 1A [20,22]. This effect is lost upon disruption of Kv2.1-syntaxin 1A binding, either by deletion of the C1a domain in the channel or introduction of a small peptide (i.e.…”

“…We previously reported that interactions between Kv2.1 and the SNARE proteins synaptosomalassociated protein 25 (SNAP-25) [17] and syntaxin 1A [18] modulate channel electrical function. More recently, we showed that syntaxin 1A binding to a C-terminal domain of Kv2.1 directly facilitates exocytosis in PC12, bovine chromaffin and rat dorsal root ganglion cells following Kv2.1 upregulation [20][21][22], and that disruption of Kv2.1-syntaxin 1A interaction impairs noradrenaline (norepinephrine) release from permeabilised PC12 cells [23]. We therefore sought to determine whether Kv2.1 plays a direct role in insulin exocytosis as such in rodent and human beta cells.…”