ABCMdb

ABCC8 p.Asn24Lys

Predicted by SNAP2:	A: D (66%), C: D (80%), D: D (66%), E: D (63%), F: D (85%), G: D (66%), H: D (59%), I: D (85%), K: D (71%), L: D (85%), M: D (85%), P: D (66%), Q: D (63%), R: D (85%), S: N (72%), T: N (78%), V: D (85%), W: D (91%), Y: D (85%),
Predicted by PROVEAN:	A: D, C: D, D: D, E: D, F: D, G: D, H: D, I: D, K: D, L: D, M: D, P: D, Q: D, R: D, S: D, T: D, V: D, W: D, Y: D,

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[hide] Sulfonylurea receptor 1 mutations that cause oppos... J Biol Chem. 2009 Mar 20;284(12):7951-9. Epub 2009 Jan 16. Pratt EB, Yan FF, Gay JW, Stanley CA, Shyng SL
Sulfonylurea receptor 1 mutations that cause opposite insulin secretion defects with chemical chaperone exposure.
J Biol Chem. 2009 Mar 20;284(12):7951-9. Epub 2009 Jan 16., [PMID:19151370]

Abstract [show]
The beta-cell ATP-sensitive potassium (K(ATP)) channel composed of sulfonylurea receptor SUR1 and potassium channel Kir6.2 serves a key role in insulin secretion regulation by linking glucose metabolism to cell excitability. Mutations in SUR1 or Kir6.2 that decrease channel function are typically associated with congenital hyperinsulinism, whereas those that increase channel function are associated with neonatal diabetes. Here we report that two hyperinsulinism-associated SUR1 missense mutations, R74W and E128K, surprisingly reduce channel inhibition by intracellular ATP, a gating defect expected to yield the opposite disease phenotype neonatal diabetes. Under normal conditions, both mutant channels showed poor surface expression due to retention in the endoplasmic reticulum, accounting for the loss of channel function phenotype in the congenital hyperinsulinism patients. This trafficking defect, however, could be corrected by treating cells with the oral hypoglycemic drugs sulfonylureas, which we have shown previously to act as small molecule chemical chaperones for K(ATP) channels. The R74W and E128K mutants thus rescued to the cell surface paradoxically exhibited ATP sensitivity 6- and 12-fold lower than wild-type channels, respectively. Further analyses revealed a nucleotide-independent decrease in mutant channel intrinsic open probability, suggesting the mutations may reduce ATP sensitivity by causing functional uncoupling between SUR1 and Kir6.2. In insulin-secreting cells, rescue of both mutant channels to the cell surface led to hyperpolarized membrane potentials and reduced insulin secretion upon glucose stimulation. Our results show that sulfonylureas, as chemical chaperones, can dictate manifestation of the two opposite insulin secretion defects by altering the expression levels of the disease mutants.

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107 These mutations are all in the TMD0 of SUR1 (amino acids 1-196) and include G7R, N24K, F27S, R74W, A116P, E128K, and V187D. Login to comment
112 Of the five mutants, G7R, N24K, and F27S had WT-like or slightly increased ATP sensitivity, and either normal or reduced MgADP response that is commonly associated with CHI mutations (Fig. 1) (23). Login to comment
134 The R74W mutant exhibited more variable Po FIGURE1.NucleotidesensitivitiesofTMD0mutantsG7R,N24K,andF27S. Login to comment
141 Scale bars: WT: 200 pA, 10 s; G7R: 200 pA, 10 s; N24K: 20 pA, 10 s; F27S: 50 pA, 10 s. B, quantification of channel response to ATP and MgADP. Login to comment
143 The ATP sensitivity of N24K is significantly higher than WT while the MgADP sensitivity of both N24K and F27S are significantly lower than WT channels (*, p Ͻ 0.05; Student`s t test). Login to comment

[hide] Role of Hsp90 in biogenesis of the beta-cell ATP-s... Mol Biol Cell. 2010 Jun 15;21(12):1945-54. Epub 2010 Apr 28. Yan FF, Pratt EB, Chen PC, Wang F, Skach WR, David LL, Shyng SL
Role of Hsp90 in biogenesis of the beta-cell ATP-sensitive potassium channel complex.
Mol Biol Cell. 2010 Jun 15;21(12):1945-54. Epub 2010 Apr 28., [PMID:20427569]

Abstract [show]
The pancreatic beta-cell ATP-sensitive potassium (K(ATP)) channel is a multimeric protein complex composed of four inwardly rectifying potassium channel (Kir6.2) and four sulfonylurea receptor 1 (SUR1) subunits. K(ATP) channels play a key role in glucose-stimulated insulin secretion by linking glucose metabolism to membrane excitability. Many SUR1 and Kir6.2 mutations reduce channel function by disrupting channel biogenesis and processing, resulting in insulin secretion disease. To better understand the mechanisms governing K(ATP) channel biogenesis, a proteomics approach was used to identify chaperone proteins associated with K(ATP) channels. We report that chaperone proteins heat-shock protein (Hsp)90, heat-shock cognate protein (Hsc)70, and Hsp40 are associated with beta-cell K(ATP) channels. Pharmacologic inhibition of Hsp90 function by geldanamycin reduces, whereas overexpression of Hsp90 increases surface expression of wild-type K(ATP) channels. Coimmunoprecipitation data indicate that channel association with the Hsp90 complex is mediated through SUR1. Accordingly, manipulation of Hsp90 protein expression or function has significant effects on the biogenesis efficiency of SUR1, but not Kir6.2, expressed alone. Interestingly, overexpression of Hsp90 selectively improved surface expression of mutant channels harboring a subset of disease-causing SUR1 processing mutations. Our study demonstrates that Hsp90 regulates biogenesis efficiency of heteromeric K(ATP) channels via SUR1, thereby affecting functional expression of the channel in beta-cell membrane.

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177 The five mutants examined harbor mutation N24K, A116P, D310N, ⌬F1388, or D1472N in the SUR1 subunit. Login to comment
178 Of these, the N24K, D310N, and D1472N mutants exhibited improved surface expression levels in cells cotransfected with Hsp90beta cDNA relative to cells cotransfected with a control empty vector. Login to comment
180 Interestingly, the N24K, D310N and D1472N mutants have relatively milder processing/trafficking defects in that they do express at the cell surface to some extent even under control conditions, in contrast to A116P and ⌬F1388 that show virtually no surface expression. Login to comment
236 Although Hsp90beta improved surface expression of the N24K, D310N, and D1472N mutant (p ϭ 0.01, 0.01, 0.05, and 0.03 for WT, N24K, D310N, and D1472N, respectively), it did not significantly increase surface expression of the A116P or ⌬F1388 mutants. Login to comment

[hide] Congenital hyperinsulinism associated ABCC8 mutati... Diabetes. 2007 Sep;56(9):2339-48. Epub 2007 Jun 15. Yan FF, Lin YW, MacMullen C, Ganguly A, Stanley CA, Shyng SL
Congenital hyperinsulinism associated ABCC8 mutations that cause defective trafficking of ATP-sensitive K+ channels: identification and rescue.
Diabetes. 2007 Sep;56(9):2339-48. Epub 2007 Jun 15., [PMID:17575084]

Abstract [show]
Congenital hyperinsulinism (CHI) is a disease characterized by persistent insulin secretion despite severe hypoglycemia. Mutations in the pancreatic ATP-sensitive K(+) (K(ATP)) channel proteins sulfonylurea receptor 1 (SUR1) and Kir6.2, encoded by ABCC8 and KCNJ11, respectively, is the most common cause of the disease. Many mutations in SUR1 render the channel unable to traffic to the cell surface, thereby reducing channel function. Previous studies have shown that for some SUR1 trafficking mutants, the defects could be corrected by treating cells with sulfonylureas or diazoxide. The purpose of this study is to identify additional mutations that cause channel biogenesis/trafficking defects and those that are amenable to rescue by pharmacological chaperones. Fifteen previously uncharacterized CHI-associated missense SUR1 mutations were examined for their biogenesis/trafficking defects and responses to pharmacological chaperones, using a combination of immunological and functional assays. Twelve of the 15 mutations analyzed cause reduction in cell surface expression of K(ATP) channels by >50%. Sulfonylureas rescued a subset of the trafficking mutants. By contrast, diazoxide failed to rescue any of the mutants. Strikingly, the mutations rescued by sulfonylureas are all located in the first transmembrane domain of SUR1, designated as TMD0. All TMD0 mutants rescued to the cell surface by the sulfonylurea tolbutamide could be subsequently activated by metabolic inhibition on tolbutamide removal. Our study identifies a group of CHI-causing SUR1 mutations for which the resulting K(ATP) channel trafficking and expression defects may be corrected pharmacologically to restore channel function.

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47 TABLE 1 Genetic and clinical information on patients carrying the CHI mutations Mutation Disease Haplotype Diazoxide response References G7R Focal G7R No 44 N24K Diffuse N24K/R1215W No Not reported F27S Focal F27S No 39 R74W Focal R74W/R1215Q No 39,45,46 E128K Diffuse E128K No Not reported R495Q Diffuse R495Q/R1215Q No 39 E501K Focal E501K No 39 L503P Focal L503P No 44 F686S Focal F686S No 39 G716V* Diffuse G716V/G716V No 47,48 K1337N Not done g3992-9a/K1337N Yes 39 L1350Q Focal L1350Q No 44 S1387F Diffuse S1387F/NA No 9,24 L1390P NA L1390P/NA No Not reported D1472H Diffuse ⌬F1388/D1472H No 39 *Patient was from consanguineous mating and therefore was homozygous for the G716V mutation (48). Login to comment
94 The first group, including G7R, N24K, F27S, R74W, and E128K, is located in the first transmembrane domain TMD0; the second group, including R495Q, E501K, L503P, F686S, and G716V, is located in the second transmembrane domain TMD1 extending through the first nucleotide binding domain; the third group, including K1337N, L1350Q, S1387F, L1390P, and D1472H, is clustered in the second nucleotide binding domain and the COOH terminus of the protein. Login to comment
102 The mutations that have not been previously reported in the literature include N24K, E128K, and L1390P. Login to comment
118 Results from this assay showed that F27S, R74W, E128K, R495Q, E501K, L503P, F686S, G716V, L1350Q, and D1472H mutant channels had greatly reduced surface expression (Ͻ20% of wild-type level)-whereas G7R and N24K mutant channels displayed modestly decreased surface expression level (Ͼ30% but Ͻ50% of wild-type level) and K1337N, S1378F, and L1390P exhibited normal or mildly reduced expression (Ͼ60% of wild-type level; Fig. 3A). Login to comment
130 In Western blots, several mutants, including G7R, N24K, F27S, R74W, and E128K, all located in TMD0, exhibited increased complex-glycosylated SUR1 in cells coexpressing Kir6.2 on overnight treatment with 1 ␮mol/l glibenclamide (Fig. 5A). Login to comment
48 TABLE 1 Genetic and clinical information on patients carrying the CHI mutations Mutation Disease Haplotype Diazoxide response References G7R Focal G7R No 44 N24K Diffuse N24K/R1215W No Not reported F27S Focal F27S No 39 R74W Focal R74W/R1215Q No 39,45,46 E128K Diffuse E128K No Not reported R495Q Diffuse R495Q/R1215Q No 39 E501K Focal E501K No 39 L503P Focal L503P No 44 F686S Focal F686S No 39 G716V* Diffuse G716V/G716V No 47,48 K1337N Not done g3992-9a/K1337N Yes 39 L1350Q Focal L1350Q No 44 S1387F Diffuse S1387F/NA No 9,24 L1390P NA L1390P/NA No Not reported D1472H Diffuse èc;F1388/D1472H No 39 *Patient was from consanguineous mating and therefore was homozygous for the G716V mutation (48). Login to comment
95 The first group, including G7R, N24K, F27S, R74W, and E128K, is located in the first transmembrane domain TMD0; the second group, including R495Q, E501K, L503P, F686S, and G716V, is located in the second transmembrane domain TMD1 extending through the first nucleotide binding domain; the third group, including K1337N, L1350Q, S1387F, L1390P, and D1472H, is clustered in the second nucleotide binding domain and the COOH terminus of the protein. Login to comment
103 The mutations that have not been previously reported in the literature include N24K, E128K, and L1390P. Login to comment
119 Results from this assay showed that F27S, R74W, E128K, R495Q, E501K, L503P, F686S, G716V, L1350Q, and D1472H mutant channels had greatly reduced surface expression (b0d;20% of wild-type level)-whereas G7R and N24K mutant channels displayed modestly decreased surface expression level (b0e;30% but b0d;50% of wild-type level) and K1337N, S1378F, and L1390P exhibited normal or mildly reduced expression (b0e;60% of wild-type level; Fig. 3A). Login to comment

[hide] Carbamazepine as a novel small molecule corrector ... J Biol Chem. 2013 Jul 19;288(29):20942-54. doi: 10.1074/jbc.M113.470948. Epub 2013 Jun 6. Chen PC, Olson EM, Zhou Q, Kryukova Y, Sampson HM, Thomas DY, Shyng SL
Carbamazepine as a novel small molecule corrector of trafficking-impaired ATP-sensitive potassium channels identified in congenital hyperinsulinism.
J Biol Chem. 2013 Jul 19;288(29):20942-54. doi: 10.1074/jbc.M113.470948. Epub 2013 Jun 6., [PMID:23744072]

Abstract [show]
ATP-sensitive potassium (KATP) channels consisting of sulfonylurea receptor 1 (SUR1) and the potassium channel Kir6.2 play a key role in insulin secretion by coupling metabolic signals to beta-cell membrane potential. Mutations in SUR1 and Kir6.2 that impair channel trafficking to the cell surface lead to loss of channel function and congenital hyperinsulinism. We report that carbamazepine, an anticonvulsant, corrects the trafficking defects of mutant KATP channels previously identified in congenital hyperinsulinism. Strikingly, of the 19 SUR1 mutations examined, only those located in the first transmembrane domain of SUR1 responded to the drug. We show that unlike that reported for several other protein misfolding diseases, carbamazepine did not correct KATP channel trafficking defects by activating autophagy; rather, it directly improved the biogenesis efficiency of mutant channels along the secretory pathway. In addition to its effect on channel trafficking, carbamazepine also inhibited KATP channel activity. Upon subsequent removal of carbamazepine, however, the function of rescued channels was recovered. Importantly, combination of the KATP channel opener diazoxide and carbamazepine led to enhanced mutant channel function without carbamazepine washout. The corrector effect of carbamazepine on mutant KATP channels was also demonstrated in rat and human beta-cells with an accompanying increase in channel activity. Our findings identify carbamazepine as a novel small molecule corrector that may be used to restore KATP channel expression and function in a subset of congenital hyperinsulinism patients.

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125 At 10 òe;M, the F27S and E128K mutations exhibited the greatest improvement to nearly the level seen with 5 òe;M glibenclamide; R74W, A116P, and V187D showed moderate responses; whereas G7R and N24K, which have less severe processing defects (31), had weak responses (Fig. 1C). Login to comment
127 At 10 òe;M, the F27S and E128K mutations exhibited the greatest improvement to nearly the level seen with 5 òe;M glibenclamide; R74W, A116P, and V187D showed moderate responses; whereas G7R and N24K, which have less severe processing defects (31), had weak responses (Fig. 1C). Login to comment

[hide] Pharmacological rescue of trafficking-impaired ATP... Front Physiol. 2013 Dec 24;4:386. doi: 10.3389/fphys.2013.00386. Martin GM, Chen PC, Devaraneni P, Shyng SL
Pharmacological rescue of trafficking-impaired ATP-sensitive potassium channels.
Front Physiol. 2013 Dec 24;4:386. doi: 10.3389/fphys.2013.00386., [PMID:24399968]

Abstract [show]
ATP-sensitive potassium (KATP) channels link cell metabolism to membrane excitability and are involved in a wide range of physiological processes including hormone secretion, control of vascular tone, and protection of cardiac and neuronal cells against ischemic injuries. In pancreatic beta-cells, KATP channels play a key role in glucose-stimulated insulin secretion, and gain or loss of channel function results in neonatal diabetes or congenital hyperinsulinism, respectively. The beta-cell KATP channel is formed by co-assembly of four Kir6.2 inwardly rectifying potassium channel subunits encoded by KCNJ11 and four sulfonylurea receptor 1 subunits encoded by ABCC8. Many mutations in ABCC8 or KCNJ11 cause loss of channel function, thus, congenital hyperinsulinism by hampering channel biogenesis and hence trafficking to the cell surface. The trafficking defects caused by a subset of these mutations can be corrected by sulfonylureas, KATP channel antagonists that have long been used to treat type 2 diabetes. More recently, carbamazepine, an anticonvulsant that is thought to target primarily voltage-gated sodium channels has been shown to correct KATP channel trafficking defects. This article reviews studies to date aimed at understanding the mechanisms by which mutations impair channel biogenesis and trafficking and the mechanisms by which pharmacological ligands overcome channel trafficking defects. Insight into channel structure-function relationships and therapeutic implications from these studies are discussed.

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218 Mutation Domain Rescue Rescue Gating References by SU by CBZ property SUR1 G7R TMD0 Yes Yes Normal Yan et al., 2007 N24K TMD0 Yes Yes Normal Yan et al., 2007 F27S TMD0 Yes Yes Normal Yan et al., 2007 R74W TMD0 Yes Yes ATP-insensitive Yan et al., 2007 A116P TMD0 Yes Yes Normal Yan et al., 2004 E128K TMD0 Yes Yes ATP-insensitive Yan et al., 2007 V187D TMD0 Yes Yes Normal Yan et al., 2004 R495Q TMD1 Yes Yes Unknown Yan et al., 2007 E501K TMD1 Yes Yes Unknown Yan et al., 2007 L503P TMD1 No No Unknown Yan et al., 2007 F686S NBD1 No No Unknown Yan et al., 2007 G716V NBD1 No No Unknown Yan et al., 2007 E1324K TMD2 N.D.3 N.D. Login to comment