ATP-sensitive potassium (K ATP ) channels mediate glucoseinduced insulin secretion by coupling metabolic signals to -cell membrane potential and the secretory machinery. Reduced K ATP channel expression caused by mutations in the channel proteins: sulfonylurea receptor 1 (SUR1) and Kir6.2, results in loss of channel function as seen in congenital hyperinsulinism. Previously, we reported that sulfonylureas, oral hypoglycemic drugs widely used to treat type II diabetes, correct the endoplasmic reticulum to the plasma membrane trafficking defect caused by two SUR1 mutations, A116P and V187D. In this study, we investigated the mechanism by which sulfonylureas rescue these mutants. We found that glinides, another class of SUR-binding hypoglycemic drugs, also markedly increased surface expression of the trafficking mutants. Attenuating or abolishing the ability of mutant SUR1 to bind sulfonylureas or glinides by the following mutations: Y230A, S1238Y, or both, accordingly diminished the rescuing effects of the drugs. Interestingly, rescue of the trafficking defects requires mutant SUR1 to be co-expressed with Kir6.2, suggesting that the channel complex, rather than SUR1 alone, is the drug target. Observations that sulfonylureas also reverse trafficking defects caused by neonatal diabetes-associated Kir6.2 mutations in a way that is dependent on intact sulfonylurea binding sites in SUR1 further support this notion. Our results provide insight into the mechanistic and structural basis on which sulfonylureas rescue K ATP channel surface expression defects caused by channel mutations.Regulation of insulin secretion by blood glucose relies on expression of functional ATP-sensitive potassium (K ATP ) channels at the -cell membrane. The -cell K ATP channel is an octameric protein complex comprising four pore-forming Kir6.2 subunits and four regulatory sulfonylurea receptor 1 (SUR1) 2 subunits (1-3). Channel activity is determined by the interplay between both channel subunits and intracellular ATP and ADP: binding of ATP to the Kir6.2 subunit inhibits channel activity, whereas binding of Mg 2ϩ -complexed ATP or ADP to the SUR1 subunit stimulates channel activity (4 -6). In this way, channel activity serves as a reporter of intracellular ATP and ADP concentrations during glucose metabolism to control -cell excitability, hence insulin secretion. Dysfunction of -cell K ATP channels because of mutations in the channel subunits Kir6.2 or SUR1 underlies congenital insulin secretion disorders (4, 5). Whereas mutations causing loss of channel function lead to excessive insulin secretion and hypoglycemia as seen in patients with congenital hyperinsulinism (CHI), those causing gain of channel function lead to insufficient insulin secretion and permanent neonatal diabetes mellitus (PNDM).In congenital hyperinsulinism, two prominent mechanisms accounting for loss of channel function are loss of channel expression at the cell surface and loss of channel sensitivity to stimulation by MgADP (7). In contrast, gain of channel function a...
Heterozygous missense mutations in the pore-forming subunit Kir6.2 of ATP-sensitive K ؉ channels (K ATP channels) have recently been shown to cause permanent neonatal diabetes mellitus (PNDM). Functional studies demonstrated that PNDM mutations reduce K ATP channel sensitivity to ATP inhibition, resulting in gain of channel function. However, the impact of these mutations on channel expression has not been examined. Here, we show that PNDM mutations, including Q52R, V59G, V59M, R201C, R201H, and I296L, not only reduce channel ATP sensitivity but also impair channel expression at the cell surface to varying degrees. By tagging the PNDM Kir6.2 mutant V59G or R201H with an additional mutation, N160D, that confers voltage-dependent polyamine block of K ATP channels, we demonstrate that in simulated heterozygous state, all surface channels are either wild-type or heteromeric channels containing both wild-type and mutant Kir6.2 subunits. Comparison of the various PNDM mutations in their effects on channel nucleotide sensitivity and expression, as well as disease phenotype, suggests that both channel-gating defect and expression level may play a role in determining disease severity. Interestingly, sulfonylureas significantly increase surface expression of certain PNDM mutants, suggesting that the efficacy of sulfonylurea therapy may be compromised by the effect of these drugs on channel expression. Diabetes 55:1738 -1746, 2006 P ancreatic ATP-sensitive K ϩ channels (K ATP channels), each consisting of four pore-forming Kir6.2 subunits and four regulatory sulfonylurea receptor one (SUR1) subunits, link -cell metabolism to insulin secretion (1-3). The activity of K ATP channels is governed mainly by the dynamics of intracellular adenine nucleotides ATP and ADP at the channel site during glucose metabolism (1,4). Both nucleotides can stimulate or inhibit channel activity depending on their relative concentrations and whether Mg 2ϩ is present. Inhibition of channels by nucleotides is mediated by the Kir6.2 subunit and does not require Mg 2ϩ (5,6), whereas nucleotide stimulation is conferred by the SUR1 subunit and requires Mg 2ϩ (7,8). The physiological activity of K ATP channels in -cells is thus a balance between nucleotide inhibition and nucleotide stimulation. During glucose stimulation, ATP concentrations increase and ADP concentrations decrease, resulting in K ATP channel closure. Because K ATP channels carry the dominant conductance in high-input resistance -cells at resting state, closure of K ATP channels leads to membrane depolarization, which in turn leads to opening of voltage-gated calcium channels, calcium influx, and insulin release.Recent studies have established heterozygous missense mutations in Kir6.2 as a major cause underlying permanent neonatal diabetes mellitus (PNDM) (9 -16). As Kir6.2 is also a constituent of K ATP channel subtypes expressed outside of pancreas, including cardiac muscle, skeletal muscle, and brain, some mutations have been reported to cause muscle weakness, dysmorphic features, ...
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