K⁺Channel Facilitation of Exocytosis by Dynamic Interaction with Syntaxin

Abstract

Kv channels inhibit release indirectly by hyperpolarizing membrane potential, but the significance of Kv channel interaction with the secretory apparatus is not known. The Kv2.1 channel is commonly expressed in the soma and dendrites of neurons, where it could influence the release of neuropeptides and neurotrophins, and in neuroendocrine cells, where it could influence hormone release. Here we show that Kv2.1 channels increase dense-core vesicle (DCV)-mediated release after elevation of cytoplasmic Ca 2ϩ . This facilitation occurs even after disruption of pore function and cannot be explained by changes in membrane potential and cytoplasmic Ca 2ϩ . However, triggering release increases channel binding to syntaxin, a secretory apparatus protein. Disrupting this interaction with competing peptides or by deleting the syntaxin association domain of the channel at the C terminus blocks facilitation of release. Thus, direct association of Kv2.1 with syntaxin promotes exocytosis. The dual functioning of the Kv channel to influence release, through its pore to hyperpolarize the membrane potential and through its C-terminal association with syntaxin to directly facilitate release, reinforces the requirements for repetitive firing for exocytosis of DCVs in neuroendocrine cells and in dendrites.

Convergent Ca 2+ and Zn 2+ signaling regulates apoptotic Kv2.1 K + currents

“…In these experiments, we also confirmed the described association of syntaxin and Kv2.1 (Fig. 3A) (47)(48)(49). The increased CaMKII signal in Kv2.1-expressing cells could be the result of a direct interaction of the kinase with the K + channel.…”

“…CaMKII has been shown to regulate exocytosis via a specific interaction with syntaxin (43)(44)(45)(46). In addition, syntaxin is known to bind to the most proximal region of the Kv2.1 C terminus, termed C1a, during Ca 2+ -facilitated exocytosis in pancreatic and other nonneuronal cells (47)(48)(49). Importantly, we showed that syntaxin is also required for the membrane insertion of Kv2.1 channels during apoptosis (23).…”

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

“…Constructs, adenoviruses and recombinant peptides The Kv2.1 pore mutant (Kv2.1 W365C/Y380T ) and construct lacking the syntaxin-binding domain (Kv2.1 W365C/Y380T ΔC1a) have been described previously [20] (Fig. 1a).…”

“…The inclusion of 5% glycerol in the final wash minimised the non-specific interactions between syntaxin 1A and protein A-sepharose beads. The bound GST-fusion proteins were eluted with reduced glutathione as described [20]. For controls, cell lysates were immunoprecipitated with an irrelevant antibody (IgG) and with protein A-sepharose alone, or by performing the immunoprecipitation with Kv2.1 antibody in the presence of its antigen peptide (1:1 ratio).…”

“…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.…”