K2.1 clusters on β-cell plasma membrane act as reservoirs that replenish pools of newcomer insulin granule through their interaction with syntaxin-3.

The voltage-dependent K (K) channel K2.1 is a major delayed rectifier in many secretory cells, including pancreatic β cells. In addition, K2.1 has a direct role in exocytosis at an undefined step, involving SNARE proteins, that is independent of its ion-conducting pore function. Here, we elucidated the precise step in exocytosis. We previously reported that syntaxin-3 (Syn-3) is the key syntaxin that mediates exocytosis of newcomer secretory granules that spend minimal residence time on the plasma membrane before fusion. Using high-resolution total internal reflection fluorescence microscopy, we now show that K2.1 forms reservoir clusters on the β-cell plasma membrane and binds Syn-3 via its C-terminal C1b domain, which recruits newcomer insulin secretory granules into this large reservoir. Upon glucose stimulation, secretory granules were released from this reservoir to replenish the pool of newcomer secretory granules for subsequent fusion, occurring just adjacent to the plasma membrane K2.1 clusters. C1b deletion blocked the aforementioned K2.1-Syn-3-mediated events and reduced fusion of newcomer secretory granules. These insights have therapeutic implications, as K2.1 overexpression in type-2 diabetes rat islets restored biphasic insulin secretion.