TALK channel control of β-cell Ca2+ handling and hormone secretion

Abstract

TALK-1 and TALK-2 channels are therapeutic targets for diabetes owing to their low conductance, high and islet-enriched expression in β-cells, and genetic/causal linkage to diabetes. TALK-1 is a regulator of ꞵ-cell electrical excitability, Ca2+ handling, and glucose-stimulated insulin secretion (GSIS). Genetic evidence links KCNK16 (encoding TALK‑1) to type 2 diabetes (T2D) and maturity-onset diabetes of the young. Moreover, ablation of β-cell TALK-1 improves glucose homeostasis in the context of diet-induced obesity. However, it remains unclear if TALK-1 can be pharmacologically targeted to enhance β-cell function. Here, we identify the first selective inhibitors of TALK-1 utilizing a high-throughput fluorescence-based thallium flux assay. A top hit, VU0544194 (K2P16-i7), was further evaluated based on its potency and TALK-1 selectivity. K2P16‑i7 increased endoplasmic reticulum (ER) Ca2+ (Ca2+ER) storage, inhibited mouse β‑cell K2P currents, and enhanced islet insulin secretion and Ca2+ entry. These findings establish the first selective TALK‑1 inhibitors and highlight an islet‑selective therapeutic strategy for diabetes. TALK‑1 and TALK‑2 share high sequence homology, and polymorphisms in or near KCNK17 (encoding TALK‑2) have been linked to increased T2D risk. Despite this, the β‑cell functions of TALK‑2 remain undefined. Thus, this project also examined how TALK‑2 influences β‑cell Ca2+ handling and insulin secretion. TALK-2 localized to the ER and plasma membrane. Heterologous TALK-2 expression resulted in reduced Ca2+ER storage, enhanced store-operated Ca2+ entry, and increased basal cytosolic Ca2+. Human islet TALK-2 was examined utilizing magnetically sorted β-cells with shRNA-mediated TALK-2 knockdown (TALK-2-KD). ER membrane single channel recordings confirmed functional TALK 2 channels on the human β-cell ER. TALK-2-KD increased Ca2+ER stores compared to Scramble shRNA controls. Additionally, cytosolic Ca2+ entry was reduced in TALK-2-KD β-cells under euglycemic conditions and increased under hyperglycemic conditions. Human pseudoislets with β-cell-specific TALK-2-KD displayed reduced basal insulin secretion and enhanced GSIS. These data suggest that TALK-2 promotes β-cell Ca2+ER release, which increases basal insulin release and reduces GSIS. Therefore, polymorphisms in KCNK17 that increase TALK-2 activity or expression would be predicted to increase T2D risk by blunting β-cell glucose-stimulated Ca2+ influx, limiting GSIS, promoting Ca2+ER leak and elevating basal insulin secretion.