AI-Based Discovery and CryoEM Structural Elucidation of a K Channel Pharmacochaperone.

Pancreatic K channel trafficking defects underlie congenital hyperinsulinism (CHI) cases unresponsive to the K channel opener diazoxide, the mainstay medical therapy for CHI. Current clinically used K channel inhibitors have been shown to act as pharmacochaperones and restore surface expression of trafficking mutants; however, their therapeutic utility for K trafficking impaired CHI is hindered by high- affinity binding, which limits functional recovery of rescued channels. Recent structural studies of K channels employing cryo-electron microscopy (cryoEM) have revealed a promiscuous pocket where several known K pharmacochaperones bind.

Non-radioactive Rb Efflux Assay for Screening K Channel Modulators.

ATP-sensitive potassium (K) channels function as metabolic sensors that link cell membrane excitability to the cellular energy status by controlling potassium ion (K) flow across the cell membrane according to intracellular ATP and ADP concentrations. As such, K channels influence a broad spectrum of physiological processes, including insulin secretion and cardiovascular functions. K channels are hetero-octamers, consisting of four inward rectifier potassium channel subunits, Kir6.

Dynamic duo: Kir6 and SUR in K channel structure and function.

K channels are ligand-gated potassium channels that couple cellular energetics with membrane potential to regulate cell activity. Each channel is an eight subunit complex comprising four central pore-forming Kir6 inward rectifier potassium channel subunits surrounded by four regulatory subunits known as the sulfonylurea receptor, SUR, which confer homeostatic metabolic control of K gating. SUR is an ATP binding cassette (ABC) protein family homolog that lacks membrane transport activity but is essential for K expression and function.

Structure of an open K channel reveals tandem PIP binding sites mediating the Kir6.2 and SUR1 regulatory interface.

ATP-sensitive potassium (K ) channels, composed of four pore-lining Kir6.2 subunits and four regulatory sulfonylurea receptor 1 (SUR1) subunits, control insulin secretion in pancreatic β-cells. K channel opening is stimulated by PIP and inhibited by ATP.

K channel mutations in congenital hyperinsulinism: Progress and challenges towards mechanism-based therapies.

Congenital hyperinsulinism (CHI) is the most common cause of persistent hypoglycemia in infancy/childhood and is a serious condition associated with severe recurrent attacks of hypoglycemia due to dysregulated insulin secretion. Timely diagnosis and effective treatment are crucial to prevent severe hypoglycemia that may lead to life-long neurological complications. In pancreatic β-cells, adenosine triphosphate (ATP)-sensitive K (K) channels are a central regulator of insulin secretion vital for glucose homeostasis.

K channels in focus: Progress toward a structural understanding of ligand regulation.

K channels are hetero-octameric complexes of four inward rectifying potassium channels, Kir6.1 or Kir6.2, and four sulfonylurea receptors, SUR1, SUR2A, or SUR2B from the ABC transporter family.

Mechanistic insights on KATP channel regulation from cryo-EM structures.

Gated by intracellular ATP and ADP, ATP-sensitive potassium (KATP) channels couple cell energetics with membrane excitability in many cell types, enabling them to control a wide range of physiological processes based on metabolic demands. The KATP channel is a complex of four potassium channel subunits from the Kir channel family, Kir6.1 or Kir6.

Ligand-mediated Structural Dynamics of a Mammalian Pancreatic K Channel.

Regulation of pancreatic K channels involves orchestrated interactions of their subunits, Kir6.2 and SUR1, and ligands. Previously we reported K channel cryo-EM structures in the presence and absence of pharmacological inhibitors and ATP, focusing on the mechanisms by which inhibitors act as pharmacological chaperones of K channels (Martin et al.

Localized islet nuclear enlargement hyperinsulinism (LINE-HI) due to ABCC8 and GCK mosaic mutations.

Objective: Congenital hyperinsulinism (HI) is the most common cause of persistent hypoglycemia in children. In addition to typical focal or diffuse HI, some cases with diazoxide-unresponsive congenital HI have atypical pancreatic histology termed Localized Islet Nuclear Enlargement (LINE) or mosaic HI, characterized by histologic features similar to diffuse HI, but confined to only a region of pancreas. Our objective was to characterize the phenotype and genotype of children with LINE-HI.

Subcellular trafficking and endocytic recycling of K channels.

Sarcolemmal/plasmalemmal ATP-sensitive K (K) channels have key roles in many cell types and tissues. Hundreds of studies have described how the K channel activity and ATP sensitivity can be regulated by changes in the cellular metabolic state, by receptor signaling pathways and by pharmacological interventions. These alterations in channel activity directly translate to alterations in cell or tissue function, that can range from modulating secretory responses, such as insulin release from pancreatic β-cells or neurotransmitters from neurons, to modulating contractile behavior of smooth muscle or cardiac cells to elicit alterations in blood flow or cardiac contractility.

Vascular K channel structural dynamics reveal regulatory mechanism by Mg-nucleotides.

Vascular tone is dependent on smooth muscle K channels comprising pore-forming Kir6.1 and regulatory SUR2B subunits, in which mutations cause Cantú syndrome. Unique among K isoforms, they lack spontaneous activity and require Mg-nucleotides for activation.

Production and purification of ATP-sensitive potassium channel particles for cryo-electron microscopy.

ATP-sensitive potassium (K) channels are multimeric protein complexes made of four inward rectifying potassium channel (Kir6.x) subunits and four ABC protein sulfonylurea receptor (SURx) subunits. Kir6.

AKAP79/150 coordinates leptin-induced PKA signaling to regulate K channel trafficking in pancreatic β-cells.

The adipocyte hormone leptin regulates glucose homeostasis both centrally and peripherally. A key peripheral target is the pancreatic β-cell, which secretes insulin upon glucose stimulation. Leptin is known to suppress glucose-stimulated insulin secretion by promoting trafficking of K channels to the β-cell surface, which increases K conductance and causes β-cell hyperpolarization.

Functional characterization of ABCC8 variants of unknown significance based on bioinformatics predictions, splicing assays, and protein analyses: Benefits for the accurate diagnosis of congenital hyperinsulinism.

ABCC8 encodes the SUR1 subunit of the β-cell ATP-sensitive potassium channel whose loss of function causes congenital hyperinsulinism (CHI). Molecular diagnosis is critical for optimal management of CHI patients. Unfortunately, assessing the impact of ABCC8 variants on RNA splicing remains very challenging as this gene is poorly expressed in leukocytes.

Leptin modulates pancreatic β-cell membrane potential through Src kinase-mediated phosphorylation of NMDA receptors.

The adipocyte-derived hormone leptin increases trafficking of K and Kv2.1 channels to the pancreatic β-cell surface, resulting in membrane hyperpolarization and suppression of insulin secretion. We have previously shown that this effect of leptin is mediated by the NMDA subtype of glutamate receptors (NMDARs).

Structural and functional diversity calls for a new classification of ABC transporters.

Members of the ATP-binding cassette (ABC) transporter superfamily translocate a broad spectrum of chemically diverse substrates. While their eponymous ATP-binding cassette in the nucleotide-binding domains (NBDs) is highly conserved, their transmembrane domains (TMDs) forming the translocation pathway exhibit distinct folds and topologies, suggesting that during evolution the ancient motor domains were combined with different transmembrane mechanical systems to orchestrate a variety of cellular processes. In recent years, it has become increasingly evident that the distinct TMD folds are best suited to categorize the multitude of ABC transporters.

Pharmacological chaperones of ATP-sensitive potassium channels: Mechanistic insight from cryoEM structures.

ATP-sensitive potassium (K) channels are uniquely evolved protein complexes that couple cell energy levels to cell excitability. They govern a wide range of physiological processes including hormone secretion, neuronal transmission, vascular dilation, and cardiac and neuronal preconditioning against ischemic injuries. In pancreatic β-cells, K channels composed of Kir6.

Ion Channels of the Islets in Type 2 Diabetes.

Ca is an essential signal for pancreatic β-cell function. Ca plays critical roles in numerous β-cell pathways such as insulin secretion, transcription, metabolism, endoplasmic reticulum function, and the stress response. Therefore, β-cell Ca handling is tightly controlled.

Novel dominant K channel mutations in infants with congenital hyperinsulinism: Validation by in vitro expression studies and in vivo carrier phenotyping.

Inactivating mutations in the genes encoding the two subunits of the pancreatic beta-cell K channel, ABCC8 and KCNJ11, are the most common finding in children with congenital hyperinsulinism (HI). Interpreting novel missense variants in these genes is problematic, because they can be either dominant or recessive mutations, benign polymorphisms, or diabetes mutations. This report describes six novel missense variants in ABCC8 and KCNJ11 that were identified in 11 probands with congenital HI.

Mechanism of pharmacochaperoning in a mammalian K channel revealed by cryo-EM.

ATP-sensitive potassium (K) channels composed of a pore-forming Kir6.2 potassium channel and a regulatory ABC transporter sulfonylurea receptor 1 (SUR1) regulate insulin secretion in pancreatic β-cells to maintain glucose homeostasis. Mutations that impair channel folding or assembly prevent cell surface expression and cause congenital hyperinsulinism.

Leptin-induced Trafficking of K Channels: A Mechanism to Regulate Pancreatic β-cell Excitability and Insulin Secretion.

The adipocyte hormone leptin was first recognized for its actions in the central nervous system to regulate energy homeostasis but has since been shown to have direct actions on peripheral tissues. In pancreatic β-cells leptin suppresses insulin secretion by increasing K channel conductance, which causes membrane hyperpolarization and renders β-cells electrically silent. However, the mechanism by which leptin increases K channel conductance had remained unresolved for many years following the initial observation.

Functional characterization of activating mutations in the sulfonylurea receptor 1 (ABCC8) causing neonatal diabetes mellitus in Asian Indian children.

Background: Gain-of-function of ATP-sensitive K (K ) channels because of mutations in the genes encoding SUR1 (ABCC8) or Kir6.2 (KCNJ11) is a major cause of neonatal diabetes mellitus (NDM). Our aim is to determine molecular defects in K channels caused by ABCC8 mutations in Asian Indian children with NDM by in vitro functional studies.

Methods for Characterizing Disease-Associated ATP-Sensitive Potassium Channel Mutations.

The ATP-sensitive potassium (K) channel formed by the inwardly rectifying potassium channel Kir6.2 and the sulfonylurea receptor 1 (SUR1) plays a key role in regulating insulin secretion. Genetic mutations in KCNJ11 or ABCC8 which encode Kir6.