ABCMdb

ABCC8 p.Val187Asp

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Publications

PMID: 21567408 [PubMed] Nakagawa H et al: "Ubiquitin-mediated proteasomal degradation of ABC transporters: a new aspect of genetic polymorphisms and clinical impacts."

No. Sentence Comment

155 Effect of Mutations and Nonsynonymous SNPs on Protein Trafficking, Maturation, or ERAD of ABC Transporters Protein AA Mutation/SNP Effect on Protein Reference ABCA1 W590S Mutation Functional defect 115 R587W Mutation Impaired glycol processing 115 Q597R Mutation Impaired glycol processing, ERAD 115,116 Y1532C Mutation Altered protein trafficking 117 R1925Q Mutation Altered protein trafficking 118 ABCA3 R43L Mutation Altered protein trafficking 119 L101P Mutation Altered protein trafficking 119 R280C Mutation Altered protein trafficking 119 ABCA4 L541P Mutation Mislocalization 120 R602W Mutation Mislocalization 120 A1038V Mutation Mislocalization 120 C1490Y Mutation Mislocalization 120 ABCB1a G268V Mutation ERAD 121 G341C Mutation ERAD 121 I1196S Mutation Reduced glycosylation 122 ABCB4 I541F Mutation Accumulation in ER 123 ABCB11a E135K Mutation Reduced level of mature protein 124 L198P Mutation Reduced level of mature protein 124 E297G Mutation Reduced level of mature protein 124 L413W Mutation Reduced level of mature protein 124 R432T Mutation Reduced level of mature protein 124 D482G Mutation Immature protein in ER 124,125 N490D Mutation Reduced level of mature protein 124 A570T Mutation Reduced level of mature protein 124 T655I Mutation Reduced level of mature protein 124 Y818F SNP Moderate reduction of protein 124 G982R Mutation Retention in ER 125 R1153C Mutation ERAD 125 R1286Q Mutation Retention in ER 125 ABCC2a R768W Mutation Impaired protein trafficking 126 I1173F Mutation Impaired protein maturation 127 R1392 Mutation Impaired protein maturation 128 M1393 Mutation Impaired protein maturation 129 ABCC4a E757K SNP Altered protein trafficking 23 ABCC7 F508 Mutation Misfolding, ERAD 36-39,130 G85E Mutation Impaired protein maturation 130-132 G91R Mutation Impaired protein maturation 130-132 N1303K Mutation Impaired protein maturation 130-132 ABCC8 WT Wild type Ubiquitin-proteasome degradation 133 A116P Mutation Ubiquitin-proteasome degradation 133 V187D Mutation Ubiquitin-proteasome degradation 133 F1388 Mutation Impaired protein trafficking 134 L1544P Mutation Impaired protein trafficking 135,136 ABCC11a G180R SNP Ubiquitin-proteasome degradation 50 27 Mutation Ubiquitin-proteasome degradation 50 ABCG2a V12M SNP Altered protein localization 96 Q141K SNP Ubiquitin-proteasome degradation 102 F208S SNP Ubiquitin-proteasome degradation 78,99 S441N SNP Ubiquitin-proteasome degradation 78,99 Mutations of ABCA1, ABCA3, ABCA4, ABCB4, ABCB11, ABCC2, ABCC7 (CFTR), and ABCC8 are associated with Tangier disease, fatal surfactant deficiency, Stargardt disease, progressive familial intrahepatic cholestasis type 3 (PFIC-3), progressive familial intrahepatic cholestasis type 2 (PFIC-2), Dubin-Johnson syndrome, cystic fibrosis, and familial hyperinsulinism, respectively. Login to comment

PMID: 15987767 [PubMed] Yan FF et al: "Role of ubiquitin-proteasome degradation pathway in biogenesis efficiency of {beta}-cell ATP-sensitive potassium channels."

No. Sentence Comment

228 Three SUR1 mutations, A116P, V187D, and ⌬F1388, which we have previously shown to result in ER retention and surface expression defects of KATP channels, were tested. Login to comment
231 To further test this hypothesis, we examined the combined effect of glibenclamide and MG132 on surface expression of the A116P and V187D mutants. Login to comment
232 Sulfonylureas such as glibenclamide have previously been shown to significantly increase surface expression of the A116P and V187D mutants, presumably by acting as pharmacological chaperones to help mutant SUR1 fold more efficiently (42). Login to comment
233 We found that pretreatment of COS cells expressing the A116P or the V187D mutant with 5 ␮M glibenclamide led to a significant increase in mutant channel surface expression (P Ͻ 0.01) on subsequent exposure to the proteasome inhibitor MG132 (Fig. 7B). Login to comment
245 A: COSm6 cells transiently coexpressing Kir6.2 and wild-type (WT) fSUR1 or fSUR1 bearing the A116P, V187D, or ⌬F1388 mutation were treated with or without 10 ␮M MG132 for 6 h and processed for chemiluminescence assays to quantify channel expression level at the cell surface. Login to comment
248 B: cells expressing Kir6.2 and WT-, A116P-, or V187D-fSUR1 were treated for 24 h with (Glib) or without (Control) 5 ␮M glibenclamide. Login to comment
260 Among them, ⌬F1388 has been proposed to cause severe folding defects, whereas A116P and V187D appear to have milder defects that can be partially overcome by sulfonylurea treatment. Login to comment
262 However, after pretreatment with glibenclamide, the A116P and V187D mutant channels responded to proteasome inhibitors with a statistically significant increase in surface expression (Fig. 7B). Login to comment
229 Three SUR1 mutations, A116P, V187D, and èc;F1388, which we have previously shown to result in ER retention and surface expression defects of KATP channels, were tested. Login to comment
234 We found that pretreatment of COS cells expressing the A116P or the V187D mutant with 5 òe;M glibenclamide led to a significant increase in mutant channel surface expression (P b0d; 0.01) on subsequent exposure to the proteasome inhibitor MG132 (Fig. 7B). Login to comment
246 A: COSm6 cells transiently coexpressing Kir6.2 and wild-type (WT) fSUR1 or fSUR1 bearing the A116P, V187D, or èc;F1388 mutation were treated with or without 10 òe;M MG132 for 6 h and processed for chemiluminescence assays to quantify channel expression level at the cell surface. Login to comment
249 B: cells expressing Kir6.2 and WT-, A116P-, or V187D-fSUR1 were treated for 24 h with (Glib) or without (Control) 5 òe;M glibenclamide. Login to comment
264 However, after pretreatment with glibenclamide, the A116P and V187D mutant channels responded to proteasome inhibitors with a statistically significant increase in surface expression (Fig. 7B). Login to comment

PMID: 19151370 [PubMed] Pratt EB et al: "Sulfonylurea receptor 1 mutations that cause opposite insulin secretion defects with chemical chaperone exposure."

No. Sentence Comment

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
108 The functional properties of rescued A116P and V187D mutant channels had been characterized in detail and shown to be normal (13). Login to comment
272 If R74W and E128K cause functional uncoupling between TMD0-SUR1 and Kir6.2, one might ask if the mutations also result in reduced physical association between the two subunits. Login to comment
273 Several SUR1-TMD0 mutations have been reported to reduce physical association between TMD0 and Kir6.2 in co-immunoprecipitation experiments, including CHI-causing A116P and V187D mutations and PNDM-causing F132L mutation (10, 30). Login to comment

PMID: 15640549 [PubMed] Fournet JC et al: "Genetics of congenital hyperinsulinism."

No. Sentence Comment

73 Morerecently,afoundermutation(V187D) was found in Finnish patients with HI (29%) [12]. Login to comment

PMID: 16380471 [PubMed] Otonkoski T et al: "Noninvasive diagnosis of focal hyperinsulinism of infancy with [18F]-DOPA positron emission tomography."

No. Sentence Comment

83 The two previously detected founder mutations SUR1-V187D (5) and SUR1-E1506K (24) were screened by direct sequencing in all Finnish patients. Login to comment
104 Genetic analysis showed that five of the patients were paternal heterozygotes for the Finnish major founder ABCC8 mutation V187D (Table 1). Login to comment
118 Representative illustrations of a patient with focal CHI who carries the paternally inherited SUR1-V187D mutation (patient 1, Table 1). Login to comment
100 In one of these (case no. 7), a pancreatic biopsy with diffuse-type pathol- TABLE 1 Clinical and genetic characteristics of the patients, together with the findings in PET scan, pancreatic catheterization, surgery, and histology Patient Age at diagnosis/PET ABCC8 mutation Response to medication Glucose need (mg/kg/min) PET PVS/PACS Surgery/histology 1 Neonatal/6 months V187D (561Tb0e;A) (Paternal) Dzxafa;, Octr af9; 12.3 focal/head focal/head focal resection/posterior neck PAD: focus 6.8 afb; 4 mm 2 4 months/13 months V187D (561Tb0e;A) (Paternal) Dzxafa;, Octr af9; 9.5 focal/body focal/body focal resection/body PAD: focus 5 afb; 4 mm 3 Neonatal/6 months G1469V (4408Gb0e;T) (Paternal) Dzxafa;, Octr af9; 12.7 focal/head focal/head focal resection/head PAD: focus 8 afb; 5 mm 4 Neonatal/3.5 years V187D (561Tb0e;A) (Paternal) Dzxafa;, Octr af9; 12.7 diffuse diffuse* ND 5 Neonatal/6 months A113V (338Cb0e;T) (Paternal) Dzxafa;, Octr af9; 12.7 diffuse diffuse Near-total pancreatectomy PAD: diffuse histology 6 Neonatal/5 years No mutations Dzx af9; 6.5 diffuse diffuse* ND 7 3 months/4 years No mutations Dzx af9; 6.5 diffuse ND Pancreas biopsy PAD: diffuse histology 8 Neonatal/9 months G92D (275Gb0e;A) Dzxafa;, Octr af9; 10.3 diffuse ND Near-total pancreatectomy PAD: diffuse histology 9 Neonatal/1 month V187D (561Tb0e;A) (Paternal) Dzxafa;, Octrafa; 20 focal/head ND focal resection/uncinate process PAD: focus 8 afb; 4 mm 10 Neonatal/2 months No mutations Dzx partial, Octr af9; 6.2 diffuse ND ND 11 5 months/13 months V187D (561Tb0e;A) (Paternal) Dzxafa;, Octr af9; 6.4 diffuse diffuse ND 12 Neonatal/1.5 months G474A (de novo) Dzxafa;, Octr af9; 15.9 diffuse ND Near-total pancreatectomy PAD diffuse; 13 Neonatal/3 months C418R (1252Tb0e;C) (Maternal) Dzxafa;, Octr af9; 26 focal/body ND focal resection/body PAD: focus 10 afb; 6 mm 14 Neonatal/6 months A113V (338Cb0e;T) (Paternal) Dzxafa;, Octr af9; 13 diffuse ND ND The glucose need refers to the glucose infusion rate that was required to maintain normoglycemia at the time of the PET scan. Login to comment
105 Genetic analysis showed that five of the patients were paternal heterozygotes for the Finnish major founder ABCC8 mutation V187D (Table 1). Login to comment
119 Representative illustrations of a patient with focal CHI who carries the paternally inherited SUR1-V187D mutation (patient 1, Table 1). Login to comment

PMID: 17575084 [PubMed] Yan FF et al: "Congenital hyperinsulinism associated ABCC8 mutations that cause defective trafficking of ATP-sensitive K+ channels: identification and rescue."

No. Sentence Comment

129 We first examined the effects of sulfonylureas, which were shown previously to improve surface expression of the A116P- and V187D-SUR1 mutants (16), on the trafficking mutants identified in this study (those that had surface expression Ͻ50% of wild type based on chemiluminescence assays shown in Fig. 3A). Login to comment
178 The first two trafficking mutations that we reported to be rescued by sulfonylurea drugs are A116P and V187D, both located in TMD0 of SUR1 (16). Login to comment
184 First, a truncated SUR1 of TMD0 alone containing the A116P or V187D trafficking mutations failed to respond to sulfonylurea rescue. Login to comment
177 The first two trafficking mutations that we reported to be rescued by sulfonylurea drugs are A116P and V187D, both located in TMD0 of SUR1 (16). Login to comment
183 First, a truncated SUR1 of TMD0 alone containing the A116P or V187D trafficking mutations failed to respond to sulfonylurea rescue. Login to comment

PMID: 10866047 [PubMed] Macfarlane WM et al: "Sulfonylurea receptor 1 and Kir6.2 expression in the novel human insulin-secreting cell line NES2Y."

No. Sentence Comment

188 Indeed, recent studies have demonstrated that even as V187D-SUR1 mutations in PHHI beta-cells led to the appearance of KATP channels in patient tissue, expression of the same mutation in Xenopus oocytes failed to reconstitute the channels (8). Login to comment
189 Indeed, recent studies have demonstrated that even as V187D-SUR1 mutations in PHHI b-cells led to the appearance of KATP channels in patient tissue, expression of the same mutation in Xenopus oocytes failed to reconstitute the channels (8). Login to comment

PMID: 15978902 [PubMed] Shi NQ et al: "Function and distribution of the SUR isoforms and splice variants."

No. Sentence Comment

69 When the RKR motif in a DF1388 mutant of SUR1 (a PHHI mutation) was mutated to AAA, the re-constituted DF1388SUR1AAA/KIR6.2 channel was somewhat active and could be expressed at the cell surface partially [22,23].A most recent study in trafficking issue of other mutant KATP channels revealed that the A116P and V187D mutants (mutations cause congenital hyperinsulinism) of SUR1 could be rescued to the cell membrane surface by sulfonylureas [24]. Login to comment

PMID: 12110524 [PubMed] Huopio H et al: "K(ATP) channels and insulin secretion disorders."

No. Sentence Comment

134 The recessively inherited missense mutation V187D, located in a transmembrane domain of SUR1, leads to severe early-onset HI (46). Login to comment
136 Interestingly, the disease phenotype is almost as severe in patients homozygous or heterozygous for the mutation; even a single copy of the V187D mutation seems to lead to a severe drug-unresponsive form of HI in compound heterozygotes. Login to comment
138 Functional studies (intact cell recordings, cell-free inside-out patches) of beta-cells isolated from an HI patient homozygous for the V187D mutation, as well as the results of recombinant KATP channel experiments, are consistent with the phenotype and show that mutation SUR1[V187D] leads to a loss of functional KATP channels that are not activated by diazoxide or somatostatin. Login to comment
164 Birthplaces of parents of HI patients with founder mutations SUR1(E1506K) (E) and SUR1(V187D) (F) are indicated (8, 40). Login to comment
123 The recessively inherited missense mutation V187D, located in a transmembrane domain of SUR1, leads to severe early-onset HI (46). Login to comment
125 Interestingly, the disease phenotype is almost as severe in patients homozygous or heterozygous for the mutation; even a single copy of the V187D mutation seems to lead to a severe drug-unresponsive form of HI in compound heterozygotes. Login to comment
127 Functional studies (intact cell recordings, cell-free inside-out patches) of beta-cells isolated from an HI patient homozygous for the V187D mutation, as well as the results of recombinant KATP channel experiments, are consistent with the phenotype and show that mutation SUR1[V187D] leads to a loss of functional KATP channels that are not activated by diazoxide or somatostatin. Login to comment
153 Birthplaces of parents of HI patients with founder mutations SUR1(E1506K) (E) and SUR1(V187D) (F) are indicated (8, 40). Login to comment

PMID: 21422196 [PubMed] Park SE et al: "Characterization of ABCC8 and KCNJ11 gene mutations and phenotypes in Korean patients with congenital hyperinsulinism."

No. Sentence Comment

28 Founder mutations are reported in certain populations, such as certain ABCC8 gene mutations (c.3992-9GOA and F1388del) in Ashkenazi Jews and V187D and E1507K mutations among the Finnish population. Login to comment

PMID: 16613899 [PubMed] Proks P et al: "A heterozygous activating mutation in the sulphonylurea receptor SUR1 (ABCC8) causes neonatal diabetes."

No. Sentence Comment

140 Two mutations in TMD0 (A116P and V187D), which cause congenital hyperinsulinism, abrogate the association of SUR1 and Kir6.2 and lead to loss of KATP channel function (27,35). Login to comment

PMID: 11772909 [PubMed] Huopio H et al: "Carriers of an inactivating beta-cell ATP-sensitive K(+) channel mutation have normal glucose tolerance and insulin sensitivity and appropriate insulin secretion."

No. Sentence Comment

5 RESEARCH DESIGN AND METHODS - We studied 8 parents of CHI patients, all 8 of whom were heterozygous for the inactivating SUR1 mutation V187D, and 10 matched control subjects. Login to comment
7 RESULTS - Carriers of the V187D substitution had normal glucose tolerance, normal tissue sensitivity to insulin, and no signs of inappropriate insulin secretion. Login to comment
8 The normal insulin response to tolbutamide indicated that heterozygosity for the V187D mutation did not impair KATP channel function. Login to comment
31 We have identified a missense mutation, V187D, in the SUR1 gene that is responsible for the majority of severe CHI cases in Finland. Login to comment
35 To determine in vivo the effects of the heterozygous state of this mutation on the regulation of beta-cell secretion, whole-body glucose metabolism, and glucose homeostasis, including the counterregulatory system against hypoglycemia, we studied eight parents of CHI patients, all eight of whom are carriers of the V187D substitution. Login to comment
37 The subjects for the present study were the parents of five patients with diffuse CHI caused by the homozygous SUR1 mutation V187D. Login to comment
39 Genetic analysis confirmed that all eight parents were heterozygous carriers of the V187D mutation. Login to comment
49 Several weeks after the other tests, four carriers of the V187D mutation and six control subjects participated in the tolbutamide test. Login to comment
73 The tolbutamide test was performed in four V187D heterozygotes and six control subjects to investigate the beta-cell response to the KATP channel antagonist tolbutamide. Login to comment
75 It was thus hypothesized that the increment in insulin secretion would be lower in the V187D carriers if their KATP channels are defective. Login to comment
98 Altogether, 7 of 8 V187D carriers and 4 of 10 control subjects had suffered from hypoglycemic symptoms. Login to comment
99 OGTT All individuals in both study groups had normal glucose tolerance according to World Health Organization criteria (16), as determined by OGTT (fasting blood glucose: V187D heterozygotes 4.2-5.5 mmol/l, control subjects 2.8-6.2 mmol/l; 2-h blood glucose: V187D heterozygotes 2.8-6.2 mmol/l, control subjects 3.46.4 mmol/l). Login to comment
101 The blood glucose levels were similar in both groups (V187D heterozygotes and control subjects) at all time points measured after the oral glucose load. Login to comment
104 Moreover, the plasma insulin response, expressed as the incremental insulin area under the curve, was similar in the two groups (V187D heterozygotes 519 Ϯ 98 pmol/l ⅐ h, control subjects 553 Ϯ 89 pmol/l ⅐ h). Login to comment
106 Similarly, the incremental C-peptide area under the curve was comparable between the study groups (V187D hetrozygotes 3,125 Ϯ Figure 1-Blood glucose (A), plasma insulin (B), and plasma C-peptide (C) concentrations during the OGTT. Login to comment
107 E, Control subjects; f, SUR1 V187D heterozygotes. Login to comment
108 Table 1-Clinical and biochemical characteristics of the study groups Control subjects V187D carriers P n 10 8 Age (years) 35.6 Ϯ 1.1 35.3 Ϯ 0.9 NS Sex (M/F) 5/5 4/4 NS BMI (kg/m2 ) 23.2 Ϯ 0.6 24.0 Ϯ 1.2 NS Fasting blood glucose (mmol/l) 4.3 Ϯ 0.2 4.7 Ϯ 0.2 NS Fasting insulin (pmol/l) 49.2 Ϯ 6.6 42.0 Ϯ 5.4 NS Fasting C-peptide (pmol/l) 530 Ϯ 35 450 Ϯ 51 NS Systolic blood pressure (mmHg) 128 Ϯ 4 129 Ϯ 3 NS Diastolic blood pressure (mmHg) 84 Ϯ 2 76 Ϯ 2 NS HbA1c (%) 5.4 Ϯ 0.1 5.4 Ϯ 0.1 NS Fasting hepatic insulin extraction (pmol C-peptide/pmol P-insulin) 10.7 Ϯ 0.3 10.7 Ϯ 0.4 NS Data are means Ϯ SEM or n. Login to comment
113 Furthermore, the incremental glucose area under the curve was similar in both study groups (71.1 Ϯ 3.9 and 70.2 Ϯ 2.7 mmol/l ⅐ min for the V187D heterozygote group and control subjects, respectively). Login to comment
117 Finally, neither the counterregulatory hormone responses in normoglycemia (serum glucagon 84.3 Ϯ 5.5 vs. 81.0 Ϯ 9.9 pmol/l, serum epinephrine 0.14 Ϯ 0.03 vs. 0.24 Ϯ 0.05 nmol/l, serum norepinephrine 1.5 Ϯ 0.2 vs. 1.8 Ϯ 0.3 nmol/l, serum cortisol 213.3 Ϯ 37.4 vs. 253.8 Ϯ 30.7 nmol/l, and serum growth hormone 0.88 Ϯ 0.13 vs. 0.46 Ϯ 0.15 ␮g/l in the V187D heterozygote group and in the control group, respectively) and in hypoglycemia (serum glucagon 95.1 Ϯ 4.2 vs. 120.7 Ϯ 18.9 pmol/l, serum epinephrine 1.3 Ϯ 0.4 vs. 1.6 Ϯ 0.3 nmol/l, serum norepinephrine 1.9 Ϯ 0.3 vs. 2.1 Ϯ 0.3 nmol/l, serum cortisol 355.9 Ϯ 85.4 vs. 493.4 Ϯ 59.3 nmol/l, and serum growth hormone 11.4 Ϯ 2.9 vs. 14.7 Ϯ 3.9 ␮g/l in the V187D heterozygote group and control subjects, respectively) nor the symptoms of hypoglycemia evaluated during the hypoglycemic clamp differed significantly between the groups. Login to comment
118 Tolbutamide test Figure 4 shows that plasma insulin and C-peptide responses to the tolbutamide injection were similar in V187D heterozygotes and control subjects when expressed as the difference between the hormone levels measured at 0 and 3 min after the tolbutamide bolus. Login to comment
119 The incremental areas under the curve (0-10 min) did not differ, either (P-insulin 1,744 Ϯ 338 vs. 2,226 Ϯ 491 pmol/l ⅐ min and C-peptide 5,950 Ϯ 1,266 vs. 6,607 Ϯ 870 pmol/l ⅐ min for V187D carriers and control subjects, respectively). Login to comment
124 In this study, we investigated in detail the glucose metabolism of such individuals, who carried the previously described SUR1 loss of function mutation V187D (9). Login to comment
131 E, Control subjects; f, SUR1 V187D heterozygotes. Login to comment
133 E And Ⅺ, control subjects; f, SUR1 V187D heterozygotes. Login to comment
136 Our present findings, demonstrating normal insulin and C-peptide responses after tolbutamide injection, indicate that normal KATP channel function is maintained in the beta-cells of the SUR1 V187D heterozygotes. Login to comment
144 In our study, only one of the mothers of V187D homozygous patients had transiently elevated blood glucose levels during pregnancy. Login to comment
145 All V187D mutation carriers had normal glucose tolerance and normal rates of whole-body glucose uptake. Login to comment
146 In addition, first-phase insulin secretion, which is known to be impaired in individuals at high risk for type 1 (24) and type 2 (25,26) diabetes, was not impaired in subjects with the V187D substitution. Login to comment
147 Our results indicate that the carriers of the V187D substitution do not have any features of type 2 diabetes. Login to comment
150 In our study, 7 of the 8 V187D carriers but only 4 of the 10 control individuals had suffered from such symptoms. Login to comment
151 Therefore, we determined whether insulin levels of the V187D carriers were higher after overnight fasting and during hypoglycemia. Login to comment
160 In the present study, carriers of the V187D substitution had quite normal counterregulatory system function. Login to comment
163 Some mutations impair the function of KATP channels only slightly, whereas the V187D mutation leads to total inactivation of pancreatic beta-cell KATP channels. Login to comment

PMID: 16023110 [PubMed] Yang K et al: "Low temperature completely rescues the function of two misfolded K ATP channel disease-mutants."

No. Sentence Comment

1 Two SUR1 mutations, A116P and V187D, reduce channel activity causing persistent hyperinsulinemic hypoglycemia of infancy. Login to comment
17 Two SUR1 mutations, A116P and V187D, have been reported to cause PHHI (Fig. 1) [7,8]. Login to comment
60 Maturation of A116P and V187D SUR1 is temperature sensitive and requires Kir6.2 We tested whether the maturation of A116P and V187D SUR1 was temperature sensitive by co-expressing them with Kir6.2 at three different temperatures. Login to comment
65 Only the lower band was found for either A116P or V187D, indicating that they were retained in the ER. Login to comment
67 At 30 °C, both mutants could form the mature upper band but the percentage of the upper band normalized to that of the WT was only 33% for A116P and 86% for V187D (Fig. 2A and B). Login to comment
68 Interestingly, at 18 °C, similar proportions of WT and mutants matured to form the upper bands (relative to WT, 96% for A116P and 99% for V187D). Login to comment
70 The reduced rescue in the processing of A116P compared to V187D at Fig. 1. Login to comment
73 SUR1 is made up of TMD0 and the core domain connected together through the cytoplasmic linker L0. A116P and V187D are two mutations that cause PHHI. Login to comment
76 Processing and misfolding of A116P and V187D mutants are temperature sensitive. Login to comment
81 However, the upper band was not detected for A116P-AAA and V187D-AAA. Login to comment
86 When expressed at 37 °C in the absence of Kir6.2, WT SUR1 but neither A116P nor V187D have been shown to be mature glycosylated when their RKR motifs were mutated to AAA (three alanines) [9]. Login to comment
87 We found that even at 18 °C, A116P and V187D SUR1-AAA could not mature to form the upper band (Fig. 2B) in the absence of Kir6.2. Login to comment
90 A116P and V187D disrupt heteromeric subunits interaction at 37 °C, but not at 18 °C, by causing misfolding One explanation for the temperature sensitive processing of these mutants is that the mutations cause misfolding in SUR1 that is temperature sensitive. Login to comment
102 We have previously shown that A116P or V187D mutation abolished the association between TMD0 and Kir6.2 at 18 °C [5], which seems to contradict with our current result obtained at the same temperature. Login to comment
106 Mutant and the WT channels show indistinguishable channel properties at 18 °C suggesting folding defect is completely corrected It is possible that A116P and V187D mutations still cause small conformational changes in SUR1 that are not detected by immunoprecipitation when expressed at 18 °C. Login to comment
115 t1/2 for azide activation is the time required to reach half of the maximally azide activated current (t1/2 for WT = 264 ± 10 s; A116P = 246 ± 10 s; and V187D = 277 ± 10 s; n = 6). Login to comment
116 Azide activated current is the maximum current obtained in the presence of azide minus the background current remained in the presence of glibenclamide (Iaz for WT = À14.53 ± 2.66 lA; A116P = À15.96 ± 3.98 lA; V187D = À14.18 ± 2.80 lA). Login to comment
117 Diazoxide/azide activation is the ratio of the current activated by diazoxide divided by the current activated by azide (Idzx/Iaz for WT = 0.95 ± 0.08; A116P = 0.92 ± 0.13; V187D = 1.12 ± 0.10). Login to comment
127 Lastly, we investigated the effect of A116P and V187D mutations on the single channel characteristics of the SUR1/Kir6.2 channels. Login to comment
134 (A) Representative current traces obtained at À100 mV from patches excised from oocytes expressing WT or A116P SUR1/Kir6.2 channels (trace for V187D mutant channels were not shown). Login to comment
139 The obtained values for k (lM) and n are: 16.71 ± 0.88 (k) and 1.43 ± 0.09 (n) for WT; 12.17 ± 0.25 and 1.17 ± 0.03 for A116P; 11.21 ± 0.22 and 1.42 ± 0.04 for V187D. Login to comment
146 Table 1 Single channel parameters for WT, A116P and V187D SUR1/Kir6.2 channelsa SUR1 + Kir6.2 A116P + Kir6.2 V187D + Kir6.2 sc1 (ms) 0.36 ± 0.002 0.43 ± 0.007 0.38 ± 0.007 sc2 (ms) 9.36 ± 2.238 7.28 ± 1.711 10.37 ± 2.895 sc3 (ms) 63.12 ± 14.543 42.92 ± 7.702 71.31 ± 37.949 ac1 0.969 ± 0.006 0.961 ± 0.008 0.975 ± 0.0006 ac2 0.025 ± 0.007 0.025 ± 0.005 0.016 ± 0.003 ac3 0.006 ± 0.002 0.014 ± 0.003 0.009 ± 0.003 so (ms) 1.31 ± 0.042 1.47 ± 0.027 1.46 ± 0.056 sb (ms) 67.66 ± 11.93 68.92 ± 17.77 77.96 ± 4.06 Po 0.595 ± 0.034 0.554 ± 0.047 0.583 ± 0.022 i (pA) 6.05 ± 0.43 6.07 ± 0.06 6.26 ± 0.17 a Explanations of the symbols can be found in the legends of Figs. 5 and 6. Login to comment
148 Conclusion Our data prove that A116P and V187D disrupt the association between the two KATP channel subunits by causing misfolding in SUR1 at physiological temperature (37 °C). Login to comment
82 However, the upper band was not detected for A116P-AAA and V187D-AAA. Login to comment
88 When expressed at 37 C in the absence of Kir6.2, WT SUR1 but neither A116P nor V187D have been shown to be mature glycosylated when their RKR motifs were mutated to AAA (three alanines) [9]. Login to comment
89 We found that even at 18 C, A116P and V187D SUR1-AAA could not mature to form the upper band (Fig. 2B) in the absence of Kir6.2. Login to comment
92 A116P and V187D disrupt heteromeric subunits interaction at 37 C, but not at 18 C, by causing misfolding One explanation for the temperature sensitive processing of these mutants is that the mutations cause misfolding in SUR1 that is temperature sensitive. Login to comment
110 Mutant and the WT channels show indistinguishable channel properties at 18 C suggesting folding defect is completely corrected It is possible that A116P and V187D mutations still cause small conformational changes in SUR1 that are not detected by immunoprecipitation when expressed at 18 C. Login to comment
119 t1/2 for azide activation is the time required to reach half of the maximally azide activated current (t1/2 for WT = 264 &#b1; 10 s; A116P = 246 &#b1; 10 s; and V187D = 277 &#b1; 10 s; n = 6). Login to comment
120 Azide activated current is the maximum current obtained in the presence of azide minus the background current remained in the presence of glibenclamide (Iaz for WT = 14.53 &#b1; 2.66 lA; A116P = 15.96 &#b1; 3.98 lA; V187D = 14.18 &#b1; 2.80 lA). Login to comment
121 Diazoxide/azide activation is the ratio of the current activated by diazoxide divided by the current activated by azide (Idzx/Iaz for WT = 0.95 &#b1; 0.08; A116P = 0.92 &#b1; 0.13; V187D = 1.12 &#b1; 0.10). Login to comment
131 Lastly, we investigated the effect of A116P and V187D mutations on the single channel characteristics of the SUR1/Kir6.2 channels. Login to comment
144 The obtained values for k (lM) and n are: 16.71 &#b1; 0.88 (k) and 1.43 &#b1; 0.09 (n) for WT; 12.17 &#b1; 0.25 and 1.17 &#b1; 0.03 for A116P; 11.21 &#b1; 0.22 and 1.42 &#b1; 0.04 for V187D. Login to comment
151 Table 1 Single channel parameters for WT, A116P and V187D SUR1/Kir6.2 channelsa SUR1 + Kir6.2 A116P + Kir6.2 V187D + Kir6.2 sc1 (ms) 0.36 &#b1; 0.002 0.43 &#b1; 0.007 0.38 &#b1; 0.007 sc2 (ms) 9.36 &#b1; 2.238 7.28 &#b1; 1.711 10.37 &#b1; 2.895 sc3 (ms) 63.12 &#b1; 14.543 42.92 &#b1; 7.702 71.31 &#b1; 37.949 ac1 0.969 &#b1; 0.006 0.961 &#b1; 0.008 0.975 &#b1; 0.0006 ac2 0.025 &#b1; 0.007 0.025 &#b1; 0.005 0.016 &#b1; 0.003 ac3 0.006 &#b1; 0.002 0.014 &#b1; 0.003 0.009 &#b1; 0.003 so (ms) 1.31 &#b1; 0.042 1.47 &#b1; 0.027 1.46 &#b1; 0.056 sb (ms) 67.66 &#b1; 11.93 68.92 &#b1; 17.77 77.96 &#b1; 4.06 Po 0.595 &#b1; 0.034 0.554 &#b1; 0.047 0.583 &#b1; 0.022 i (pA) 6.05 &#b1; 0.43 6.07 &#b1; 0.06 6.26 &#b1; 0.17 a Explanations of the symbols can be found in the legends of Figs. 5 and 6. Login to comment
153 Conclusion Our data prove that A116P and V187D disrupt the association between the two KATP channel subunits by causing misfolding in SUR1 at physiological temperature (37 C). Login to comment

PMID: 15562009 [PubMed] Henwood MJ et al: "Genotype-phenotype correlations in children with congenital hyperinsulinism due to recessive mutations of the adenosine triphosphate-sensitive potassium channel genes."

No. Sentence Comment

116 Whereas one patient who was homozygous for the common Finnish SUR1 V187D mutation had a negative AIR to tolbutamide (0.14 ␮U/ml), a second patient with the same mutations had a modest response (11.7 ␮U/ml); the third patient with compound heterozygosity for Kir6.2 (c 1-54 t)/K67N mutations had an AIR to tolbutamide of 68 ␮U/ml, implying considerable residual channel function. Login to comment

PMID: 16416420 [PubMed] Gloyn AL et al: "Mutations in the genes encoding the pancreatic beta-cell KATP channel subunits Kir6.2 (KCNJ11) and SUR1 (ABCC8) in diabetes mellitus and hyperinsulinism."

No. Sentence Comment

240 population two founder mutations have been reported (V187D and E1507 K) [Huopio et al., 2000; Otonkoski et al., 1999]. Login to comment
241 The V187D mutation is associated with 50% of HI in this population [Otonkoski et al., 1999]. Login to comment

PMID: 14707124 [PubMed] Yan F et al: "Sulfonylureas correct trafficking defects of ATP-sensitive potassium channels caused by mutations in the sulfonylurea receptor."

No. Sentence Comment

2 We report here that sulfonylureas also function as chemical chaperones to rescue KATP channel trafficking defects caused by two SUR1 mutations, A116P and V187D, identified in patients with congenital hyperinsulinism. Login to comment
3 Sulfonylureas markedly increased cell surface expression of the A116P and V187D mutants by stabilizing the mutant SUR1 proteins and promoting their maturation. Login to comment
5 Importantly, both mutant channels rescued to the cell surface have normal ATP, MgADP, and diazoxide sensitivities, demonstrating that SUR1 harboring either the A116P or the V187D mutation is capable of associating with Kir6.2 to form functional KATP channels. Thus, sulfonylureas may be used to treat congenital hyperinsulinism caused by certain KATP channel trafficking mutations. Login to comment
43 Here, we report that two PHHI-associated SUR1 mutations, A116P and V187D (2, 21, 29, 37), located in the first transmembrane domain (TM0), prevent trafficking of KATP channels from the ER to the plasma membrane. Login to comment
93 RESULTS Both the A116P and V187D Mutations in SUR1 Prevent Normal Cell Surface Expression of KATP Channels-Several recent studies have shown that defective KATP channel trafficking is an underlying mechanism of congenital hyperinsulinism. Login to comment
95 To examine whether mutations located in other parts of the molecule also affect channel trafficking, we focused our attention to two mutations, A116P and V187D, that are located in the first transmembrane domain, or TM0, of SUR1 (Fig. 1), and that have previously been reported to not form functional channels when co-expressed with Kir6.2 (6, 21, 37). Login to comment
96 To investigate how the A116P and V187D mutations lead to loss of functional KATP channels, we first performed Western blot analysis. Login to comment
101 Fig. 2A shows that, although both the immature and mature forms were seen with cells co-expressing WT-fSUR1 and Kir6.2, only the immature form was evident in cells co-expressing Kir6.2 and the A116P- or the V187D-fSUR1 mutants. Login to comment
104 In contrast to the abundant surface staining observed in cells transfected with Kir6.2 and WT-fSUR1, surface staining in cells transfected with Kir6.2 and A116P-fSUR1 or V187D-fSUR1 was barely detectable (Fig. 2B, top panels). Login to comment
106 These results led us to conclude that the A116P and V187D mutations cause loss of functional KATP channels by preventing channels from trafficking to the cell surface. Login to comment
107 The Trafficking Defects of the A116P and V187D Mutants Are Intrinsic to SUR1-One potential explanation for the trafficking defects seen with the A116P- and V187D-fSUR1 mutations is that the SUR1 mutants are unable to associate with Kir6.2. Login to comment
109 To address this possibility, we used a heterotandem dimer construct in which the C terminus of the mutant fSUR1 has been fused to the N terminus of Kir6.2 (referred to as A116P- or V187D-fSUR1/Kir6.2 fusion) to achieve obligatory physical association between the two subunits; similar SUR1/ Kir6.2 fusion constructs have been used previously by a number of groups for structure-function and trafficking studies (7, 9, 10, 28, 34). Login to comment
111 Although WT fSUR1/Kir6.2 fusion protein was expressed at a level comparable with that observed in cells transfected with WT-fSUR1 and Kir6.2 as individual subunits, fusion proteins carrying the A116P- or V187D-SUR1 mutation had poor surface expression, ϳ10 and 20% that of WT fSUR1/ Kir6.2 fusion, respectively (Fig. 3A). Login to comment
114 Analysis of the A116- and V187D-SUR1 mutants by immunoblotting and immunofluorescent staining experiments. Login to comment
117 In contrast, only the immature band is observed in cells expressing Kir6.2 and A116P- or V187D-fSUR1. Login to comment
118 The total steady-state protein level of A116P- and V187D-fSUR1 also appears less than that of WT-fSUR1. Login to comment
120 B, top panels, surface staining of COSm6 cells transiently transfected with Kir6.2 and either WT-, A116P-, or V187D-fSUR1 using the M2 anti-FLAG mouse monoclonal antibodies followed by Cy-3-conjugated anti-mouse secondary antibody. Login to comment
122 Whereas cells expressing WT-fSUR1 channels have abundant surface staining, those expressing A116P- or V187D-fSUR1 channels have barely detectable staining. Login to comment
125 Both A116P- and V187D-fSUR1 were detected inside the cell, with a perinuclear staining pattern. Login to comment
126 fWT, WT-fSUR1; fA116P, A116P-fSUR1; fV187D, V187D-fSUR1. Login to comment
128 The A116P and V187D mutations in SUR1. Login to comment
129 The locations of the A116P and V187D mutations in SUR1 are shown. Login to comment
133 The surface expression levels of A116P-fSUR1AAA and V187D-fSUR1AAA are 10 and 20% that of WT-fSUR1AAA, respectively (Fig. 3B). Login to comment
135 In addition, although the surface expression levels of the A116P and V187D mutant channels were very low (7 and 19% of WT; see Figs. Login to comment
139 Taken together, the results led us to propose that the A116P and V187D mutations cause trafficking defects in SUR1, possibly by promoting protein misfolding, but that the mutant proteins retain the ability to associate with Kir6.2 to form functional channels. Login to comment
140 Another potential mechanism for the deficient surface expression of KATP channels is that the A116P and V187D mutations interfere with proper shielding of the RKR signals in the channel complex. Login to comment
142 We found that inactivation of the RKR signal in SUR1 slightly increased surface expression of the A116P mutant channels (from 6 to 18% of normal expression level) but not the V187D mutant, and removal of the RKR signal in Kir6.2 (Kir6.2⌬C25) also had very little effect on the surface expression of either mutant (Fig. 3C). Login to comment
144 Glibenclamide Corrects the Channel Trafficking Defects Caused by the A116P and V187D Mutations in SUR1-The data presented so far suggest that the two mutations likely cause defective channel trafficking by promoting misfolding of SUR1. Login to comment
151 We found that treating cells with 5% glycerol for 24 h slightly improved surface expression of both the A116P and V187D mutants as well as the WT channel (Fig. 4A). Login to comment
152 Treating cells with 5 ␮M glibenclamide, however, dramatically increased surface expression of A116P-fSUR1, from 5 to 55%, and of V187D-fSUR1, from 19% to 70% of normal WT channel expression level (Fig. 4A). Login to comment
154 The effect of glibenclamide on the A116P and V187D mu- FIG. 3. Login to comment
155 The trafficking defects of the A116P- and V187D-fSUR1 mutants are intrinsic to SUR1. Login to comment
156 A, obligatory association between SUR1 and Kir6.2 does not overcome trafficking defects caused by A116P or V187D. Login to comment
157 Fusion fSUR1/Kir6.2 constructs containing either A116P or V187D mutation still exhibit poor surface expression compared with the WT fusion construct. Login to comment
162 B, the A116P and V187D mutations cause trafficking defects in SUR1. Login to comment
164 However, introducing A116P or V187D to fSUR1AAA (fA116PAAA and fV187DAAA) abolishes the ability of the proteins to traffic to the cell surface. Login to comment
165 C, trafficking defects caused by A116P and V187D do not involve improper shielding of RKR signals in the channel complex. Login to comment
166 Inactivation of the RKR signal in SUR1 (SUR1RKR/AAA) only slightly improved surface expression of A116P but not V187D, whereas removal of RKR in Kir6.2 (Kir6.2⌬C25) slightly improved surface expression of V187D but not A116P. Login to comment
167 fWT, WT-fSUR1; fA116P, A116P-fSUR1; fV187D, V187D-fSUR1. Login to comment
170 The response of the A116P- and the V187D-fSUR1 mutants to glibenclamide was specific; another SUR1 ligand, FIG. 4. Login to comment
171 Glibenclamide rescues surface expression of the A116P and V187D mutant KATPchannels. A, cells co-expressing Kir6.2 and WT-, A116P-, or V187D-fSUR1 were subjected to the different drug treatments indicated for 24 h, and surface expression of fSUR1 was quantified using the chemiluminescence assay as described in Fig. 3. Login to comment
172 Without any drug treatment, expression levels of the A116P and V187D mutants are 6.4 Ϯ 1.2 and 19.1 Ϯ 4.8% that of WT, respectively. Login to comment
174 Glibenclamide treatment at 5 ␮M for 24 h dramatically improved surface expression of both A116P- and V187D-fSUR1 (to 55.4 Ϯ 4.8 and 70.4 Ϯ 14.5% of WT, respectively) but only slightly increased WT expression (by 6.6 Ϯ 2.8%). Login to comment
176 B, Western blots showing that in cells expressing Kir6.2 and the A116P or V187D mutant fSUR1, treatment with 5 ␮M glibenclamide for 24 h led to appearance of the mature band, which was not detected in untreated cells (Fig. 2A). Login to comment
178 Top panels, surface staining of COSm6 cells transfected with Kir6.2 and either WT-, A116P-, or V187D-fSUR1, as described for Fig. 2B. Login to comment
180 In contrast to the data shown in Fig. 2B, cells expressing A116P- or V187D-fSUR1 mutant channels had strong surface staining that is nearly comparable with cells expressing WT-fSUR1 channels. Login to comment
183 fWT, WT-fSUR1; fA116P, A116P-fSUR1; fV187D, V187D-fSUR1; Glib, glibenclamide. Login to comment
185 In fact, diazoxide slightly decreased the surface expression of both A116P and V187D mutants. Login to comment
196 Cells expressing A116P-fSUR1 alone were pulse-labeled for 30 min and chased for up to 18 h in the presence or absence of 5 ␮M glibenclamide. Login to comment
199 Tolbutamide Also Rescues SUR1 A116P Mutant Channels to the Cell Surface, and the Expressed Channels Are Fully Functional after Tolbutamide Washout-Following our observation that glibenclamide significantly improves cell surface expression of the A116P and V187D mutants, the question arises as to whether the rescued channels are still glibenclamide-bound and whether they are physiologically functional. Login to comment
204 Concentration and time dependence of the effect of glibenclamide on surface expression of A116P- and V187D-fSUR1 mutant KATPchannels. A, cells transfected with Kir6.2 and WT-, A116P-, or V187D-fSUR1 were treated with different concentrations of glibenclamide for 24 h, and the surface expression of fSUR1 was quantified by the chemiluminescence assay. Login to comment
210 The cells expressing A116P- or V187D-fSUR1 mutant channels were treated with 5 ␮M for different periods of time as indicated, and surface expression of the mutant channel was quantified by chemiluminescence assays. Login to comment
212 Each data point represents the average from 2-3 experiments, and the error bar is the deviation from the average or the S.E. fWT, WT-fSUR1; fA116P, A116P-fSUR1; fV187D, V187D-fSUR1. Login to comment
216 Fig. 7A shows that in chemiluminescence assays, tolbutamide also increases surface expression of both A116P- and V187D-fSUR1 mutant channels, at concentrations of 100 and 300 ␮M that we tested (only 300 ␮M is shown). Login to comment
217 Consistently, in inside-out patch clamp recording experiments, tolbutamide treatment led to parallel increases in the current size of both mutant channels; the average patch current amplitudes in K-INT for A116P- and V187D-fSUR1 channels are 3.24 Ϯ 0.75 nA (n ϭ 12) and 5.40 Ϯ 1.19 nA (n ϭ 13), respectively, compared with 7.05 Ϯ 1.02 nA (n ϭ 17) for control WT-fSUR1 channels. Login to comment
231 In this study, we show that two SUR1 point mutations, A116P and V187D, identified in patients with congenital hyperinsulinism (2, 37) cause defective trafficking and a lack of cell surface expression of KATP channels. Login to comment
233 Mechanisms of Trafficking Defects Caused by the A116P and V187D Mutations-Multiple steps are involved in the proper expression of KATP channels on the cell surface. Login to comment
252 fWT, WT-fSUR1; fA116P, A116P-fSUR1. Login to comment
253 ported by our metabolic pulse-chase labeling experiments, which showed that glibenclamide slowed the degradation rate of the A116P mutant SUR1 and, in the presence of Kir6.2, promoted maturation of the mutant protein. Login to comment
255 This conclusion differs somewhat from that reached by Chan et al. (29), who proposed that the A116P and V187D mutations cause PHHI by preventing association between SUR1 and FIG. 7. Login to comment
256 Tolbutamide rescues functional A116P and V187D mutant channels to the cell surface. Login to comment
257 A, cells transfected with Kir6.2 and A116P- or V187D-fSUR1 were treated with 300 ␮M tolbutamide for 24 h, and surface expression of channels was measured by chemiluminescence assays. Login to comment
258 At 300 ␮M, tolbutamide was nearly as effective as 5 ␮M glibenclamide and restored surface expression of A116P- and V187D-fSUR1 channels from 6.4 Ϯ 1.2 to 49.9 Ϯ 7.6% and from 19.1 Ϯ 4.8 to 48.9 Ϯ 6.0% of normal levels, respectively. Login to comment
267 C, representative KATP current traces recorded from inside-out membrane patches containing WT-fSUR1, A116P-fSUR1, or V187D-fSUR1 channels 2 h after tolbutamide removal. Login to comment
272 fWT, WT-fSUR1; fA116P, A116P-fSUR1; fV187D, V187D-fSUR1. Login to comment
284 However, the first transmembrane domain (TM0) where the A116P and V187D mutations are located has not been implicated in glibenclamide binding. Login to comment
292 By contrast, diazoxide, which too binds SUR1 but is structurally quite different from sulfonylureas and results in channel stimulation, does not correct the trafficking defect of either A116P- or V187D-fSUR1. Login to comment
293 Comparison with Other Trafficking Mutants-A number of missense or point deletion mutations in SUR1 have been reported to reduce or prevent cell surface expression of KATP channels, including ⌬F1388, R1394H, L1544P, A1457T, V1550D, and L1551V (34-36, 50). Login to comment
296 Although like A116P and V187D they all result in a lack of surface channel expression phenotype, the mechanisms leading to this phenotype differ, as revealed by their responses to the different rescuing strategies. Login to comment
300 Although sulfonylureas rescue the surface expression of mutant channels bearing the A116P or the V187D mutation, they do not rescue surface expression of either ⌬F1388 or L1544P mutant channels (36). Login to comment
307 Although genetic and clinical data on the A116P mutation have not been published, the V187D mutation has been shown to account for the majority of PHHI cases in Finland (37). Login to comment
309 The results presented in this study show that sulfonylureas have rapid, potent, and long lasting (for at least 12 h after drug removal) effects on rescuing the A116P and V187D mutant channels to the cell surface. Login to comment
310 Most importantly, both A116P- and V187D-SUR1 mutant channels rescued to the cell surface by tolbutamide are fully functional upon drug removal and respond to MgADP and diazoxide stimulation like WT channels. Thus, mutant channels rescued to the surface will be able to respond to metabolic signals and to diazoxide treatment. Login to comment
42 Here, we report that two PHHI-associated SUR1 mutations, A116P and V187D (2, 21, 29, 37), located in the first transmembrane domain (TM0), prevent trafficking of KATP channels from the ER to the plasma membrane. Login to comment
90 The data were presented as the means afe; S.E. RESULTS Both the A116P and V187D Mutations in SUR1 Prevent Normal Cell Surface Expression of KATP Channels-Several recent studies have shown that defective KATP channel trafficking is an underlying mechanism of congenital hyperinsulinism. Login to comment
92 To examine whether mutations located in other parts of the molecule also affect channel trafficking, we focused our attention to two mutations, A116P and V187D, that are located in the first transmembrane domain, or TM0, of SUR1 (Fig. 1), and that have previously been reported to not form functional channels when co-expressed with Kir6.2 (6, 21, 37). Login to comment
98 Fig. 2A shows that, although both the immature and mature forms were seen with cells co-expressing WT-fSUR1 and Kir6.2, only the immature form was evident in cells co-expressing Kir6.2 and the A116P- or the V187D-fSUR1 mutants. Login to comment
103 These results led us to conclude that the A116P and V187D mutations cause loss of functional KATP channels by preventing channels from trafficking to the cell surface. Login to comment
108 Although WT fSUR1/Kir6.2 fusion protein was expressed at a level comparable with that observed in cells transfected with WT-fSUR1 and Kir6.2 as individual subunits, fusion proteins carrying the A116P- or V187D-SUR1 mutation had poor surface expression, b03;10 and 20% that of WT fSUR1/ Kir6.2 fusion, respectively (Fig. 3A). Login to comment
115 The total steady-state protein level of A116P- and V187D-fSUR1 also appears less than that of WT-fSUR1. Login to comment
119 Whereas cells expressing WT-fSUR1 channels have abundant surface staining, those expressing A116P- or V187D-fSUR1 channels have barely detectable staining. Login to comment
123 fWT, WT-fSUR1; fA116P, A116P-fSUR1; fV187D, V187D-fSUR1. Login to comment
130 The surface expression levels of A116P-fSUR1AAA and V187D-fSUR1AAA are 10 and 20% that of WT-fSUR1AAA, respectively (Fig. 3B). Login to comment
132 In addition, although the surface expression levels of the A116P and V187D mutant channels were very low (7 and 19% of WT; see Figs. Login to comment
136 Taken together, the results led us to propose that the A116P and V187D mutations cause trafficking defects in SUR1, possibly by promoting protein misfolding, but that the mutant proteins retain the ability to associate with Kir6.2 to form functional channels. Login to comment
137 Another potential mechanism for the deficient surface expression of KATP channels is that the A116P and V187D mutations interfere with proper shielding of the RKR signals in the channel complex. Login to comment
141 Glibenclamide Corrects the Channel Trafficking Defects Caused by the A116P and V187D Mutations in SUR1-The data presented so far suggest that the two mutations likely cause defective channel trafficking by promoting misfolding of SUR1. Login to comment
148 We found that treating cells with 5% glycerol for 24 h slightly improved surface expression of both the A116P and V187D mutants as well as the WT channel (Fig. 4A). Login to comment
149 Treating cells with 5 òe;M glibenclamide, however, dramatically increased surface expression of A116P-fSUR1, from 5 to 55%, and of V187D-fSUR1, from 19% to 70% of normal WT channel expression level (Fig. 4A). Login to comment
153 A, obligatory association between SUR1 and Kir6.2 does not overcome trafficking defects caused by A116P or V187D. Login to comment
159 B, the A116P and V187D mutations cause trafficking defects in SUR1. Login to comment
161 However, introducing A116P or V187D to fSUR1AAA (fA116PAAA and fV187DAAA) abolishes the ability of the proteins to traffic to the cell surface. Login to comment
163 Inactivation of the RKR signal in SUR1 (SUR1RKR/AAA) only slightly improved surface expression of A116P but not V187D, whereas removal of RKR in Kir6.2 (Kir6.2èc;C25) slightly improved surface expression of V187D but not A116P. Login to comment
168 Glibenclamide rescues surface expression of the A116P and V187D mutant KATPchannels. A, cells co-expressing Kir6.2 and WT-, A116P-, or V187D-fSUR1 were subjected to the different drug treatments indicated for 24 h, and surface expression of fSUR1 was quantified using the chemiluminescence assay as described in Fig. 3. Login to comment
169 Without any drug treatment, expression levels of the A116P and V187D mutants are 6.4 afe; 1.2 and 19.1 afe; 4.8% that of WT, respectively. Login to comment
173 B, Western blots showing that in cells expressing Kir6.2 and the A116P or V187D mutant fSUR1, treatment with 5 òe;M glibenclamide for 24 h led to appearance of the mature band, which was not detected in untreated cells (Fig. 2A). Login to comment
175 Top panels, surface staining of COSm6 cells transfected with Kir6.2 and either WT-, A116P-, or V187D-fSUR1, as described for Fig. 2B. Login to comment
177 In contrast to the data shown in Fig. 2B, cells expressing A116P- or V187D-fSUR1 mutant channels had strong surface staining that is nearly comparable with cells expressing WT-fSUR1 channels. Login to comment
182 In fact, diazoxide slightly decreased the surface expression of both A116P and V187D mutants. Login to comment
201 Concentration and time dependence of the effect of glibenclamide on surface expression of A116P- and V187D-fSUR1 mutant KATPchannels. A, cells transfected with Kir6.2 and WT-, A116P-, or V187D-fSUR1 were treated with different concentrations of glibenclamide for 24 h, and the surface expression of fSUR1 was quantified by the chemiluminescence assay. Login to comment
207 The cells expressing A116P- or V187D-fSUR1 mutant channels were treated with 5 òe;M for different periods of time as indicated, and surface expression of the mutant channel was quantified by chemiluminescence assays. Login to comment
209 Each data point represents the average from 2-3 experiments, and the error bar is the deviation from the average or the S.E. fWT, WT-fSUR1; fA116P, A116P-fSUR1; fV187D, V187D-fSUR1. Login to comment
213 Fig. 7A shows that in chemiluminescence assays, tolbutamide also increases surface expression of both A116P- and V187D-fSUR1 mutant channels, at concentrations of 100 and 300 òe;M that we tested (only 300 òe;M is shown). Login to comment
214 Consistently, in inside-out patch clamp recording experiments, tolbutamide treatment led to parallel increases in the current size of both mutant channels; the average patch current amplitudes in K-INT for A116P- and V187D-fSUR1 channels are 3.24 afe; 0.75 nA (n afd; 12) and 5.40 afe; 1.19 nA (n afd; 13), respectively, compared with 7.05 afe; 1.02 nA (n afd; 17) for control WT-fSUR1 channels. Login to comment
228 In this study, we show that two SUR1 point mutations, A116P and V187D, identified in patients with congenital hyperinsulinism (2, 37) cause defective trafficking and a lack of cell surface expression of KATP channels. Login to comment
230 Mechanisms of Trafficking Defects Caused by the A116P and V187D Mutations-Multiple steps are involved in the proper expression of KATP channels on the cell surface. Login to comment
254 A, cells transfected with Kir6.2 and A116P- or V187D-fSUR1 were treated with 300 òe;M tolbutamide for 24 h, and surface expression of channels was measured by chemiluminescence assays. Login to comment
264 C, representative KATP current traces recorded from inside-out membrane patches containing WT-fSUR1, A116P-fSUR1, or V187D-fSUR1 channels 2 h after tolbutamide removal. Login to comment
269 fWT, WT-fSUR1; fA116P, A116P-fSUR1; fV187D, V187D-fSUR1. Login to comment
281 However, the first transmembrane domain (TM0) where the A116P and V187D mutations are located has not been implicated in glibenclamide binding. Login to comment
289 By contrast, diazoxide, which too binds SUR1 but is structurally quite different from sulfonylureas and results in channel stimulation, does not correct the trafficking defect of either A116P- or V187D-fSUR1. Login to comment
297 Although sulfonylureas rescue the surface expression of mutant channels bearing the A116P or the V187D mutation, they do not rescue surface expression of either èc;F1388 or L1544P mutant channels (36). Login to comment
304 Although genetic and clinical data on the A116P mutation have not been published, the V187D mutation has been shown to account for the majority of PHHI cases in Finland (37). Login to comment
306 The results presented in this study show that sulfonylureas have rapid, potent, and long lasting (for at least 12 h after drug removal) effects on rescuing the A116P and V187D mutant channels to the cell surface. Login to comment

PMID: 12881418 [PubMed] Chan KW et al: "N-terminal transmembrane domain of the SUR controls trafficking and gating of Kir6 channel subunits."

No. Sentence Comment

4 Using Xenopus oocytes to coexpress truncated SUR constructs with Kir6, we demonstrated by immunoprecipitation, single-oocyte chemiluminescence and electrophysiological measurements that the TMD0 of SUR1 strongly associated with Kir6.2 and modulated its traf®cking and gating. Two TMD0 mutations, A116P and V187D, previously correlated with persistent hyperinsulinemic hypoglycemia of infancy, were found to disrupt the association between TMD0 and Kir6.2. Login to comment
147 Two PHHI mutations located in TMD0 abolish its association with 6.2 Two TMD0 mutations, A116P and V187D, have been reported to cause PHHI (Aguilar-Bryan and Bryan, 1999; Otonkoski et al., 1999). Login to comment
215 Two PHHI mutations located in TMD0, A116P and V187D disrupt the association between TMD0 and 6.2. Login to comment
216 (A) A116P and V187D mutations completely abolish the total current expressed from TMD0+6.2HA. Login to comment
218 (B) A116P and V187D mutations completely abolish the ability of TMD0 to enhance the surface expression of 6.2D26. Login to comment
220 (C) A116P and V187D mutations completely abolish the ability of TMD0 to traf®c to the cell surface. Login to comment
222 (D) A116P and V187D mutations disrupt the association between TMD0 and 6.2. Login to comment
232 PHHI mutations affect KATP channels by disrupting the function of TMD0 Two mutations, A116P and V187D, have been reported to correlate with PHHI (Aguilar-Bryan and Bryan, 1999; Otonkoski et al., 1999). Login to comment
234 While detailed analysis of A116P has not been reported, V187D has been shown to abolish the function of the pancreatic KATP channels both in native b cells and when expressed in Xenopus oocytes (Otonkoski et al., 1999). Login to comment
249 Four nanograms of SUR1, 1 ng of TMD0 [including F195, F195(A116P), F195(V187D), TMD0*, 1±195, S2-TMD0 and MRP1-TMD0*], 3 ng of F196-917, 3 ng of 918M and 2 ng of Kir6 (including various 6.2 and 6.1 constructs) RNAs were used in independent or coexpression experiments for TEVC, macropatch recording, western blotting and immunoprecipitation. Login to comment

PMID: 16956886 [PubMed] Yan FF et al: "Sulfonylureas correct trafficking defects of disease-causing ATP-sensitive potassium channels by binding to the channel complex."

No. Sentence Comment

2 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. Login to comment
40 We have previously reported that sulfonylureas rescue surface expression defects of KATP channels caused by two CHI-associated SUR1 mutations, A116P and V187D (23). Login to comment
42 In this work, we investigated the mechanism by which sulfonylureas correct the channel surface expression defects caused by the A116P or V187D mutations and by the PNDM-associated Kir6.2 mutations. Login to comment
44 Interestingly and somewhat unexpectedly, we found that Kir6.2 is required for sulfonylureas to rescue the A116P and V187D mutant SUR1 at the cell surface. Login to comment
59 A, topology model of SUR1 showing the three transmembrane domains, the location of the A116P and V187D trafficking mutations, the location of the -RKR-ER retention motif, and the location of the S1238Y and Y230A mutations that have been proposed to disrupt the Aand B- sulfonyl- urea binding sites (as shown in B), respectively. Login to comment
89 Of these, two mutations, A116P and V187D, were rescued by the pharmacological agent sulfonylurea (23). Login to comment
91 Because both A116P and V187D are located in TMD0, we first tested if sulfonylureas bind directly to TMD0 to facilitate mutant protein biogenesis and trafficking, even though TMD0 has not been implicated in sulfonylurea binding in prior studies (21). Login to comment
92 We compared surface expression of recombinant SUR1-TMD0 (amino acids 1-197) containing either the A116P or the V187D mutation in the presence or absence of 5 ␮M glibenclamide. Login to comment
95 By con- FIGURE2.ThefirsttransmembranedomainofSUR1(TMD0)doesnotconfer the sulfonylurea rescue effect on the A116P or V187D mutations. Login to comment
96 A, schematic showing the fSUR1-TMD0 (amino acids 1-197) and the Kir6.2⌬C25 constructs used in experiments shown in B. B, surface expression of fSUR1-TMD0 harboring mutations A116P or V187D in cells treated with or without glibenclamide (5 ␮M for 24 h). Login to comment
99 Each bar represents the mean Ϯ S.E. of three to four independent experiments. Sulfonylureas and KATP Channel Trafficking trast, when the A116P or V187D mutations were introduced into fSUR1-TMD0, surface expression was greatly reduced (by Ͼ70%), even though the total mutant recombinant proteins were abundantly expressed as assessed by Western blots and immunofluorescent staining of permeabilized cells (not shown). Login to comment
102 Restoration of A116P- and V187D-mutant Channel Expression by Sulfonylureas Is Dependent on Intact Sulfonylurea Binding Sites in SUR1-Several studies indicate that the high affinity tolbutamide binding site in SUR1 resides in transmembrane segments 13-16 (19, 20). Login to comment
105 If sulfonylureas rescue the A116P and V187D trafficking mutants by binding to the channel protein, then introducing the S1237Y mutation should also reduce or abolish the ability of sulfonylureas to correct the trafficking defect. Login to comment
106 We made the equivalent sulfonylurea binding site mutation (S1238Y) in hamster SUR1 (16) and examined how it affects the response of the A116P- or V187D-mutant channels to sulfonylureas. Login to comment
107 Initial assessment by immunofluorescent staining indicates that the S1238Y mutation by itself does not affect fSUR1 surface expression when coexpressed with Kir6.2; however, when combined with the A116P or V187D mutation, it indeed reduced or prevented the ability of glibenclamide to rescue the surface expression defect caused by the A116P and V187D mutations (Fig. 3). Login to comment
116 However, when combined with the A116P- or V187D-SUR1 trafficking mutations, S1238Y completely abolished the rescue effect of tolbutamide at 300 (Fig. 4B) and 600 ␮M (not shown). Login to comment
118 In the WT background (no sulfonylurea binding mutation), 24-h treatment with 1 ␮M glibenclamide increased surface expression of the A116P mutant from 3.1 Ϯ 0.7 to 34.5 Ϯ 7.2% and the V187D mutant from 12.8 Ϯ 1.6 to 49.2 Ϯ 6.8% of WT channels. Login to comment
119 But in the S1238Y background, the same treatment only slightly increased surface expression of the A116P and V187D mutants, from 0.7 Ϯ 1.0 to 6.6 Ϯ 2.1% and 9.8 Ϯ 1.4 to 15.6 Ϯ 2.3% of WT, respectively. Login to comment
120 Increasing the concentration of glibenclamide to 5 ␮M led to a much greater effect on surface expression of the mutants (to 21.8 and 23.9% of WT for A116P and V187D, respectively; Fig. 4C). Login to comment
125 Bryan et al. (14) have shown that mutation of Tyr230 located in the intracellular loop between TMD0 and TMD1 to alanine (Y230A) results in loss of [125 I]azidoglibenclamide photoaffinity labeling, suggesting that the benzamido group lies in close proximity to Tyr230 during binding, although it is possible that Y230A abolishes binding indirectly by affecting a distant site. Login to comment
126 We therefore tested whether this mutation also interferes with the ability of sulfonylureas to rescue the A116P and V187D trafficking mutants. Login to comment
136 The Y230A or S1238Y mutations reduced the ability of glibenclamide to restore surface expression of the A116P mutant and simultaneous mutation of Y230A and S1238Y completely abolished the ability of glibenclamide to rescue A116P to the cell surface. Login to comment
137 C, same as B except that the V187D trafficking mutation was introduced into WT or sulfonylurea binding mutation fSUR1 background. Login to comment
138 Note as previously reported, the V187D mutation showed slightly higher surface expression compared with the A116P mutation, with faint surface staining barely visible (23). Login to comment
140 Accordingly, we examined how repaglinide, a glinide that contains the benzamido moiety but not the sulfonylurea moiety, affects surface expression of A116P and V187D channels in the presence or absence of the Y230A mutation. Login to comment
141 Fig. 5B shows that repaglinide at 10 ␮M effectively rescued surface expression of the A116P and V187D mutants, whereas the Tyr230 mutation abolished this rescue effect; in contrast, the S1238Y mutation had little effect on the ability of repaglinide to rescue the trafficking mutants (not shown). Login to comment
142 Surprisingly, we found that the Y230A mutation also renders tolbutamide unable to rescue the A116P and V187D trafficking mutants (Fig. 5C), suggesting a role of Tyr230 in tolbutamide binding or in conferring tolbutamide sensitivity toward trafficking rescue through an allosteric effect. Login to comment
153 Surface expression of tolbutamide-treated cells was significantly higher than untreated for A116P and V187D (p Ͻ 0.001) but not A116P/S1238Y and V187D/S1238Y. Login to comment
155 Without the S1238Y mutation, both 1 and 5 ␮M glibenclamide significantly increased surface expression of the A116P and V187D trafficking mutants (p Ͻ 0.001). Login to comment
156 However, for the A116P/ S1238Y and V187D/S1238Y mutants, 1 ␮M glibenclamide did not lead to a statistically significant increase in surface expression, whereas 5 ␮M glibenclamide did (p Ͻ 0.01). Login to comment
158 FIGURE 5. Impact of the Y230A mutation on the effectiveness of sulfonylureas and glinides to rescue KATP channel trafficking defects in the presence of Kir6.2. Login to comment
159 The A116P or V187D trafficking mutation was introduced onto the WTor Y230A-fSUR1 background. Login to comment
165 Each bar is the mean Ϯ S.E. of three to five independent experiments. Sulfonylureas and KATP Channel Trafficking 33408 the trafficking defects of the A116P and V187D mutants via direct interactions with the mutant channel proteins. Login to comment
175 The Role of Kir6.2 in Sulfonylurea Rescue of Channel Trafficking Defects-The data we presented so far demonstrate that intact sulfonylurea binding sites in SUR1 are necessary for effective rescue of the A116P- or V187D-SUR1 trafficking mutants by sulfonylureas. Login to comment
177 To investigate the role of Kir6.2 in sulfonylurea rescue of the A116P- or V187D-SUR1 trafficking mutants, we took advantage of the fact that inactivation of the -RKR-ER retention/retrieval motif by mutation to AAA (referred to as WTAAA in Fig. 8A) in SUR1 allows SUR1 to traffic to the cell surface without co-expression of Kir6.2 (29). Login to comment
178 If sulfonylurea binding to SUR1 is sufficient to correct the defect caused by the A116P or V187D mutations, we expect sulfonylureas to improve the surface expression of A116P- or V187D-SUR1 in which the -RKR- motif has been mutated to -AAA- (referred to as A116PAAA and V187DAAA) in the absence of Kir6.2. Login to comment
180 These results are consistent with the A116P and V187D mutations causing defects in the SUR1 protein itself. Login to comment
185 Effect of the Y230A/S1238Y double mutation on sulfonylurea and glinide rescue of KATP channel trafficking mutants in the presence of Kir6.2. Login to comment
186 The experiments were as described in the legend to Fig. 5 except that A116P and V187D were each introduced onto the WTor Y230A/S1238Y-fSUR1 backgrounds. Login to comment
212 Our results demonstrate that mutations in SUR1 previously reported to abolish or reduce sulfonylurea binding also abolish or reduce the ability of sulfonylureas to rescue channel trafficking defects caused by the A116P or V187D SUR1 mutations and that both the sulfonylurea and benzoamido moieties of glibenclamide contribute to the rescue effect. Login to comment
218 A reasonable hypothesis for how sulfonylureas improve surface expression of the A116P and V187D mutants is that A116P or V187D cause SUR1 misfolding and sulfonylureas, upon binding to the mutant SUR1, help it to adopt the correct conformation. Login to comment
219 Our results that the A116P or V187D mutations are sufficient to prevent trafficking of TMD0 as well as SUR1AAA to the cell surface support the idea that the A116P or V187D mutations have an effect on the folding/processing of SUR1 protein itself. Login to comment
227 The A116P or V187D trafficking mutation was engineered into WT-fSUR1 or fSUR1 in which the -RKR-ER retention motif has been inactivated by mutation to AAA. Login to comment
238 It is also worth pointing out that TMD0 harboring the A116P and V187D mutations have been shown to not coimmunoprecipitate with Kir6.2 (27), suggesting weakened association between mutant SUR1 and Kir6.2. Login to comment
254 Although we used the A116P and V187D-SUR1 mutations as two examples for probing the mechanism by which sulfonylureas rescue channel trafficking defect, we have found many more CHI-causing KATP mutants with trafficking defects to respond to sulfonylurea rescue.3 Our findings are therefore applicable to a growing number of naturally occurring channel mutations whose trafficking defects could be targeted for therapy. Login to comment
58 A, topology model of SUR1 showing the three transmembrane domains, the location of the A116P and V187D trafficking mutations, the location of the -RKR-ER retention motif, and the location of the S1238Y and Y230A mutations that have been proposed to disrupt the A- and B- sulfonyl- urea binding sites (as shown in B), respectively. Login to comment
88 Of these, two mutations, A116P and V187D, were rescued by the pharmacological agent sulfonylurea (23). Login to comment
90 Because both A116P and V187D are located in TMD0, we first tested if sulfonylureas bind directly to TMD0 to facilitate mutant protein biogenesis and trafficking, even though TMD0 has not been implicated in sulfonylurea binding in prior studies (21). Login to comment
94 By con- FIGURE2.ThefirsttransmembranedomainofSUR1(TMD0)doesnotconfer the sulfonylurea rescue effect on the A116P or V187D mutations. Login to comment
115 However, when combined with the A116P- or V187D-SUR1 trafficking mutations, S1238Y completely abolished the rescue effect of tolbutamide at 300 (Fig. 4B) and 600 òe;M (not shown). Login to comment
117 In the WT background (no sulfonylurea binding mutation), 24-h treatment with 1 òe;M glibenclamide increased surface expression of the A116P mutant from 3.1 afe; 0.7 to 34.5 afe; 7.2% and the V187D mutant from 12.8 afe; 1.6 to 49.2 afe; 6.8% of WT channels. Login to comment
139 Accordingly, we examined how repaglinide, a glinide that contains the benzamido moiety but not the sulfonylurea moiety, affects surface expression of A116P and V187D channels in the presence or absence of the Y230A mutation. Login to comment
152 Surface expression of tolbutamide-treated cells was significantly higher than untreated for A116P and V187D (p b0d; 0.001) but not A116P/S1238Y and V187D/S1238Y. Login to comment
154 Without the S1238Y mutation, both 1 and 5 òe;M glibenclamide significantly increased surface expression of the A116P and V187D trafficking mutants (p b0d; 0.001). Login to comment
164 Each bar is the mean afe; S.E. of three to five independent experiments. Sulfonylureas and KATP Channel Trafficking 33408 JOURNAL OF BIOLOGICAL CHEMISTRY VOLUME 281ߦNUMBER 44ߦNOVEMBER 3, 2006 the trafficking defects of the A116P and V187D mutants via direct interactions with the mutant channel proteins. Login to comment
174 The Role of Kir6.2 in Sulfonylurea Rescue of Channel Trafficking Defects-The data we presented so far demonstrate that intact sulfonylurea binding sites in SUR1 are necessary for effective rescue of the A116P- or V187D-SUR1 trafficking mutants by sulfonylureas. Login to comment
176 To investigate the role of Kir6.2 in sulfonylurea rescue of the A116P- or V187D-SUR1 trafficking mutants, we took advantage of the fact that inactivation of the -RKR-ER retention/retrieval motif by mutation to AAA (referred to as WTAAA in Fig. 8A) in SUR1 allows SUR1 to traffic to the cell surface without co-expression of Kir6.2 (29). Login to comment
179 These results are consistent with the A116P and V187D mutations causing defects in the SUR1 protein itself. Login to comment
211 Our results demonstrate that mutations in SUR1 previously reported to abolish or reduce sulfonylurea binding also abolish or reduce the ability of sulfonylureas to rescue channel trafficking defects caused by the A116P or V187D SUR1 mutations and that both the sulfonylurea and benzoamido moieties of glibenclamide contribute to the rescue effect. Login to comment
217 A reasonable hypothesis for how sulfonylureas improve surface expression of the A116P and V187D mutants is that A116P or V187D cause SUR1 misfolding and sulfonylureas, upon binding to the mutant SUR1, help it to adopt the correct conformation. Login to comment
226 The A116P or V187D trafficking mutation was engineered into WT-fSUR1 or fSUR1 in which the -RKR-ER retention motif has been inactivated by mutation to AAA. Login to comment
236 It is also worth pointing out that TMD0 harboring the A116P and V187D mutations have been shown to not coimmunoprecipitate with Kir6.2 (27), suggesting weakened association between mutant SUR1 and Kir6.2. Login to comment
252 Although we used the A116P and V187D-SUR1 mutations as two examples for probing the mechanism by which sulfonylureas rescue channel trafficking defect, we have found many more CHI-causing KATP mutants with trafficking defects to respond to sulfonylurea rescue.3 Our findings are therefore applicable to a growing number of naturally occurring channel mutations whose trafficking defects could be targeted for therapy. Login to comment

PMID: 18708750 [PubMed] Masia R et al: "Regulation of KATP channel expression and activity by the SUR1 nucleotide binding fold 1."

No. Sentence Comment

142 The ubiquitination‑proteasomal pathway of degradation may be involved: this is the mechanism that underlies the effects of the PHHI mutants A116P and V187D.26 However, the proteasomal inhibitor MG‑132 failed to rescue total DNBF1 protein levels, and incubation with sulfonylureas, which facilitates SUR1 folding and prevents its degradation, was also without effect. Login to comment

PMID: 18767144 [PubMed] Flanagan SE et al: "Update of mutations in the genes encoding the pancreatic beta-cell K(ATP) channel subunits Kir6.2 (KCNJ11) and sulfonylurea receptor 1 (ABCC8) in diabetes mellitus and hyperinsulinism."

No. Sentence Comment

139 In the Finnish population, two founder mutations have been reported (V187D and E1507 K) [Otonkoski et al., 1999; Huopio et al., 2000]; the V187D mutation is associated with 50% of HI in this population [Otonkoski et al., 1999]. Login to comment

PMID: 12364426 [PubMed] Huopio H et al: "Acute insulin response tests for the differential diagnosis of congenital hyperinsulinism."

No. Sentence Comment

5 The patients with SUR1-V187D mutation showed a reduced response to tolbutamide but unexpectedly did not show any response to calcium stimulation. Login to comment
20 Indeed, the previously reported SUR1 mutations V187D (3) and E1506K (14) are the cause of most genetically characterized CHI cases. Login to comment
21 The mutation SUR1-V187D leads to total loss of function of KATP channels and severe drug-resistant phenotype. Login to comment
39 The fourth group was composed of one homozygous and five compound heterozygote patients with the mutation SUR1-V187D (aged 1-14 yr). Login to comment
41 The patients with paternal SUR1-V187D and maternal SUR1-A1457T (n ϭ 1) or SUR1-V1550D (n ϭ 1) were excluded from AIR tests because of the requirement of insulin more than 0.5 U/kg per day. Login to comment
43 All pancreatectomized patients who were included in AIR tests had the diffuse form of CHI as judged by histopathological examination (no KATP channel mutation, n ϭ 1; Kir6.2-(-54)/K67N, n ϭ 1; SUR1-E1506K, n ϭ 1; SUR1-V187D, n ϭ 5). Login to comment
58 Clinical characteristics of the patients Case Sex Age Cause of hyperinsulinism Previous treatment of hyperinsulinism No KATP channel mutation 1 M 2 Unknown Diazoxide 2 F 3 Unknown Octreotide 3 M 5 Unknown Diazoxide 4 M 20 Unknown Diazoxide, subtotal pancreatectomy 5 F 26 Unknown Diazoxide Kir6.2-(-54)/K67N 6 M 8 Paternal Kir6.2-K67N, maternal Kir6.2-(-54) Octreotide, subtotal pancreatectomy SUR1-E1506K 7 F 6 Dominant maternal SUR1-E1506K Diazoxide 8 F 9 Dominant maternal SUR1-E1506K Diazoxide 9 F 15 Dominant maternal SUR1-E1506K Frequent feeds 10 F 16 Dominant maternal SUR1-E1506K Diazoxide 11 F 19 Dominant maternal SUR1-E1506K Frequent feeds 12 M 27 Dominant maternal SUR1-E1506K Diazoxide, subtotal pancreatectomy SUR1-V187D 13 F 1 Paternal SUR1-V187D, maternal genotype pending Octreotide 14 F 6 Maternal SUR1-V187D, paternal genotype pending Subtotal pancreatectomy 15 M 8 Paternal SUR1-V187D, maternal genotype pending Subtotal pancreatectomy 16 F 8 Homozygous SUR1-V187D Subtotal pancreatectomy 17 F 9 Maternal SUR1-V187D, paternal genotype pending Subtotal pancreatectomy 18 F 14 Maternal SUR1-V187D, paternal genotype pending Subtotal pancreatectomy 19 M 11 Paternal SUR1-V187D, maternal SUR1-A1457T Subtotal pancreatectomy 20 F 13 Paternal SUR1-V187D, maternal SUR1-V1550D Subtotal pancreatectomy SUR1-L1551V 21 M 2 Paternal SUR1-L1551V, maternal genotype pending Diazoxide 22 F 0.2 Paternal SUR1-L1551V, maternal genotype pending Diazoxide Diabetic patients are shown in italics. Login to comment
82 The mutation A1457T in exon 36 was found to be maternally inherited in one compound heterozygote patient with the paternally inherited mutation SUR1-V187D (case 19). Login to comment
83 The mutation V1550D in exon 39 of SUR1 was maternally inherited in one individual who also had paternally inherited SUR1-V187D (case 20). Login to comment
87 Despite two separate screening processes of the KATP channel genes, we were not able to identify another SUR1 mutation in the five SUR1-V187D compound heterozygote patients who were tested with the AIR tests. Login to comment
90 First, three of the patients (cases 14, 17, and 18) had inherited the V187D mutation from their mother, which is inconsistent with focal disease. Login to comment
95 in all study groups: 0.19 mmol/liter in CHI patients without KATP mutations, 0.23 mmol/liter in the patient with both Kir6.2 mutations, 0.17 mmol/liter in SUR1-E1506K patients, and 0.23 mmol/liter in SUR1-V187D patients. Login to comment
96 The acute plasma C-peptide response to calcium was significantly increased in patients with SUR1-E1506K (159 Ϯ 28 pmol/liter), compared with either patients without KATP channel mutations (33 Ϯ 25 pmol/liter) (P Ͻ 0.05) or SUR1-V187D carriers (41 Ϯ 15 pmol/liter) (P Ͻ 0.05). Login to comment
97 The response to calcium was not significantly different between the SUR1-V187D carriers and patients without KATP channel mutations. Login to comment
98 It is obvious that the subjects with SUR1-V187D have very little remaining beta-cell function after the subtotal pancreatectomy and that this is maximally stimulated even under basal conditions. Login to comment
101 The plasma insulin and C-peptide responses to tolbutamide appeared to be lower in subjects with SUR-V187D and SUR-E1506K channel mutations, compared with the subjects without KATP channel mutations, but the differences were not statistically significant because of the small number of observations. Login to comment
105 It was clearly subnormal in the prepubertal SUR1-V187D homozygous patient (case 16) and in the postpubertal SUR1-E1506K heterozygotes. Login to comment
108 The two previously reported founder SUR1 mutations, V187D (3) and E1506K (14), account for 88% of the genetically characterized cases. Login to comment
110 Correlation between genotype and phenotype The verified compound heterozygote subjects (A1457T/ V187D and V1550D/V187D) show a very severe and drug-resistant disease phenotype. Login to comment
111 This is likely to be due to the total loss of channel activity by both of the novel mutations because we have shown that the mutation V187D alone does not cause any impairment of insulin secretion in heterozygous carriers (22). Login to comment
118 AIRs in nondiabetic patients shown as the means of the increments at 1 and 3 min Case Insulin response to calcium C-peptide response to calcium Insulin response to tolbutamide C-peptide response to tolbutamide Insulin response to glucose No KATP channel mutation 1 37 62 175 727 299 2 0 0 392 1099 607 3 20 49 139 5 38 101 906 1413 1438 Median 29 55 450 1132 453 Kir6.2-(-54)/K67N 6 284 652 491 1136 1083 SUR1-E1506K 7 62 147 127 306 197 8 55 171 476 1148 1001 9 36 77 34 36 105 10 83 265 34 93 75 11 42 200 26 33 74 Median 55 171 34 93 105 SUR1-V187D 13 10 80 166 550 216 16 5 30 1 8 42 Median 8 55 84 279 129 Reference values 1 Ϯ 4a 318 Ϯ 72b 252 Ϯ 54b (-12-25) (158-478) The individual results and median values are shown for each group, expressed as picomoles per liter. Login to comment
147 Unexpectedly, subjects with SUR1-V187D did not respond to calcium stimulation. Login to comment
154 The C-peptide response to tolbutamide was severely decreased in SUR1-V187D subjects. Login to comment
155 This finding is in agreement with the previous results of ion channel recordings of beta-cells isolated from a SUR1-V187D homozygous patient. Login to comment
156 Unlike control cells that were activated by diazoxide and inhibited by tolbutamide, no actions of KATP channel agonists diazoxide or octreotide were seen in the cells with SUR1-V187D mutation (3). Login to comment
163 The results show, however, that a negative response to calcium stimulation does not exclude the possibility that a subject has a KATP channel mutation, as clearly demonstrated by the major Finnish SUR1 mutation V187D. Login to comment

PMID: 19254908 [PubMed] James C et al: "The genetic basis of congenital hyperinsulinism."

No. Sentence Comment

38 Review J Med Genet 2009;46:289-299. doi:10.1136/jmg.2008.064337 Homozygous, compound heterozygous and heterozygous recessive inactivating mutations (missense, frameshift, nonsense, insertions/deletions (macrodeletion), splice site and regulatory mutations) have been reported in ABCC8 and KCNJ11.10-17 67 68 So far, more than 150 mutations have been reported in ABCC8 and 25 in KCNJ11.69 In the Ashkenazi Jewish population two common mutations (F1388del and c.3992-9G4A) account for 90% of all cases of CHI9 10 whereas in the Finnish population, two founder mutations have been reported (V187D and E1507 K).14 22 Recessive inactivating mutations in ABCC8 and KCNJ11 usually cause severe CHI which in the vast majority of patients is unresponsive to medical treatment with diazoxide. Login to comment

PMID: 10334322 [PubMed] Otonkoski T et al: "A point mutation inactivating the sulfonylurea receptor causes the severe form of persistent hyperinsulinemic hypoglycemia of infancy in Finland."

No. Sentence Comment

4 Sequence analysis revealed a novel point mutation in exon 4 of SUR1, predicting a valine to aspartic acid change at amino acid 187 (V187D). Login to comment
5 Of the total cases, 15 affected individuals harbored this mutation in heterozygous or homozygous form, and all of these had severe hyperinsulinemia that responded poorly to medical treatment and required subtotal pancreatectomy. No KATP channel activity was observed in -cells isolated from a homozygous patient or after coexpression of recombinant Kir6.2 and SUR1 carrying the V187D mutation. Login to comment
57 To detect the V187D mutation, PCR amplification of genomic DNA was performed using the primers 5 -GTGAGTGTACACATGATG and 5 -CAGAGCCA GAGCCTCTGCTT. Login to comment
85 Oocytes were then co-injected with a mixture of mRNAs encoding Kir6.2 (~0.04 ng) and either wild-type SUR1 or V187D SUR1 (~2 ng) at dilutions yielding a final injection volume of about ~50 nl/oocyte. Login to comment
89 Open circles represent carriers of the V187D mutation of SUR1, detected with the Tth111I restriction endonuclease test. Login to comment
123 Even in this TABLE 1 Major clinical findings in Finnish PHHI patients Gestational Birth weight Blood glucose (mmol/l); Age at Mutation Patient Sex age (weeks) (g) plasma insulin (pmol/l) pancreatectomy (days) V187D 1 M 34 3,920 (+4.6) 0.5; 534 23 +/- 2 M 33 2,350 (+0.2) 5.0; 282 29 - 3 F 35 3,120 (+1.1) 5.0; 342 22 +/- 4 M 36 3,940 (+2.4) 3.0; 564 16 +/+ 5 M 39 5,080 (+3.3) 1.8; 96 57 +/- 6 M 39 3,970 (+0.9) 1.2; 78 - - 7 F 37 3,420 (+0.7) 1.4; 236 38 +/- 8 F 39 4,020 (+1.3) 1.5; 341 161 +/- 9 F 29 2,070 (+4.6) 2.2; 440 31 +/+ 10 M 40 3,710 (0.0) 1.5; 190 38 +/- 11 F 39 3,900 (+1.1) 1.6; 654 11 +/- 12 F 33 3,710 (+5.0) 1.4; 1040 12 +/+ 13 F 37 4,100 (+2.4) 1.7; 149 59 +/+ 14 F 34 3,700 (+4.0) 2.2; 396 - - 15 M 39 3,845 (+0.7) 2.2; 1200 11 +/- 16 M 36 3,150 (+0.4) 1.2; 186 - - 17 M 36 5,250 (+5.6) 2.2; 874 21/196 +/+ 18 M 35 2,910 (+0.3) 2.8; 215 82 +/- 19 M 35 4,565 (+5.2) 4.1; 850 11 - 20 F 38 4,330 (+0.6) 1.5; 138 - - 21 M 28 1,770 (+3.8) 1.8; 615 15 (+/+)* 22 M 39 2,900 (-1.5) 2.1; 294 - - 23 F 37 5,360 (+5.7) 2.4; 984 - - 24 F 37 4,490 (+3.3) 1.2; 257 - - Data are n or n (SD). Login to comment
136 The geographicaldistribution of birthplaces for parents carrying the marker haplotype and V187D supports a single origin for the mutation (Fig. 1). Login to comment
144 This point mutation is predicted to substitute a valine for an aspartic acid residue at site 187 of the SUR1 protein (V187D). Login to comment
150 An additional 20 Finnish PHHI families were analyzed for the presence of the V187D mutation. Login to comment
153 Presence of the V187D mutation was associated with a severeclinical phenotype. Login to comment
155 None of the 23 additional individuals who came from outside Finland, were of diverse ethnic origin, and were affected with classic PHHI possessed the V187D mutation as assessed by the Tth111I restriction assay. Login to comment
156 This demonstrated that the V187D mutation is rare in the PHHI population at large and that its prevalence within the Finnish population may be attributed to a founder effect. Login to comment
160 of control Haplotype with V187D without V187D chromosomes 2-10-5-7-8-8 13 0 0 3-10-5-7-8-8 1 0 1 2-10-5-7-8-7 1 0 0 2-10-5-7-10-7 1 0 0 2-10-5-7-2-9 1 0 0 2-10-5-7-10-10 1 0 0 All other haplotypes 0 20 32 Total 18 24 33 The PHHI-associated chromosomes were divided in two groups based on the occurrence of the V187D mutation. Login to comment
161 All haplotypes associated with V187D were considered derivatives of a single ancestral chromosome with historical recombinations. Login to comment
166 We therefore screened 100 chromosomes from normal individuals living in this area and found the V187D mutation in heterozygous form in one of them. Login to comment
168 These results are consistent with the idea that V187D is not a common polymorphism. Login to comment
170 The functional properties of KATP channels in -cells isolated from a PHHI patient homozygous for the V187D mutation (case 4) were studied using both intact cell recordings and cell-free inside-out patches. Login to comment
180 The relationship between the novel SUR1 gene defect and loss of channel function was therefore investigated in recombinant experiments with SUR1 containing the V187D mutation identified in our PHHI patients. Login to comment
182 Whole-cell currents were recorded from Xenopus oocytes injected with mRNA encoding Kir6.2 and with either wild-type SUR1 (wt-SUR1) or SUR1 engineered to carry the mutation found in the Finnish population of PHHI patients (SUR1-V187D). Login to comment
183 Under basal conditions, currents recorded from oocytes injected with Kir6.2/wt-SUR1 (n = 5) or Kir6.2/SUR1-V187D (n = 5) were no different from those measured in oocytes injected with water (n = 4), entirely consistent with previous observations (24). Login to comment
185 No such increase in current was observed in response to sodium azide in either control oocytes or oocytes injectedwith Kir6.2/SUR1-V187D (Fig. 4A and B). Login to comment
186 This lack of response observed in Kir6.2/SUR1-V187D-injected oocytes may have arisen because the mutant channel is insensitive to metabolic regulation or because it does not form a functional channel in the plasma membrane. Login to comment
187 To determine which of these possibilities is correct, we recordedrecombinant KATP channel activityingiant inside-out membranepatches.Figure4C shows currents elicitedbya voltage-ramp from -100 to +100 mV in the cell-attached condition (c/a) and after excision (i/o) into the ATP-free solution recorded from oocytes injected with wt-SUR1/Kir6.2 or Kir6.2/SUR1-V187D. Login to comment
189 By contrast, there was no increase in KATP channel activity (Fig. 4C [right panel] and D) in patches excised fromoocytes injected with Kir6.2/SUR1-V187D. Login to comment
190 These findings were consistent with those obtained from acutely isolated PHHI -cells (Fig. 3B) and suggested that the presence of the V187D mutation renders the channel completely nonfunctional. Login to comment
192 We have reported here that a single mutation in the SUR1 gene, V187D, detected in 18of42disease-associated chromosomes,revealed FIG. 2. Login to comment
193 Demonstration of the V187D mutation. Login to comment
205 The V187D mutationwas not detected in 9 families, although in 5 of these families, one or both of the disease-associated chromosomes carried partial haplotypes observedin otherpatients heterozygous for V187D. Login to comment
210 The in vitro studies described here have demonstrated clearly that the V187D point mutation leads to loss of functional KATP channelexpression, even in excised patches, and thus falls into the former group. Login to comment
214 The V187D mutation appears to be more severe than N188S, as we could not observe any KATP channel activity in excised patches. Login to comment
222 In contrast, in Finnish patients the V187D mutation was strongly associated with a severe form of the disease. Login to comment
223 It is interesting that the clinical phenotype of PHHI was equally severe, whether the patients were homo- or heterozygous for the V187D mutation. Login to comment
234 presence ofeven a single copy of the V187D mutation will prevent the generation of functionally operating KATP channels with high diazoxide sensitivity, if another, possibly less severe mutation is also present. Login to comment
236 The presence of the V187D mutation can be easily detected. Login to comment
238 Identification of the V187D mutation is prognostically valuable because we observed that this mutation was invariably associated with a severe disease that responded poorly to medication in all cases. Login to comment

PMID: 12627323 [PubMed] Reimann F et al: "Characterisation of new KATP-channel mutations associated with congenital hyperinsulinism in the Finnish population."

No. Sentence Comment

64 The mutations V187D and E1506K have been described previously [3, 5]. Login to comment
186 In both cases they occurred as part of a complex heterozygous genotype, with the SUR1 mutation V187D on the second allele [3]. Login to comment
187 As SUR1-V187D also abolished KATP channel activity [3], these two subjects would be predicted to have no functional KATP channels, consistent with the observed severe, drug-resistant CHI phenotype [23]. Login to comment

PMID: 19475716 [PubMed] Sandal T et al: "The spectrum of ABCC8 mutations in Norwegian patients with congenital hyperinsulinism of infancy."

No. Sentence Comment

109 Clinical characteristics of Norwegian CHI patients carrying mutations in ABCC8 Proband Sex Birth weight (g)/gestation length (weeks)a Treatment Mutationsd Medicalb Surgeryc Maternal chromosome Paternal chromosome Hypo-N3 F 6190/38 Deceased Yes (S) R1493W R1493W Hypo-N6 M 5340/38 Somatostatin, diet (FM, PEG) No V21D V21D Hypo-N8 F 5740/37 Insulin Yes (S) G1400R R1493W Hypo-N9 F 5130/40 Diet (FM) Yes (S) - IVS1011G.T Hypo-N11 M 4000/38 None No - G1478Re Hypo-N14 M 5000/40 Somatostatin, diet (FM, PEG) No - IVS1011G.T Hypo-N16 F 3780/38 Diet (FM) No - C267X Hypo-N19 F 5240/40 Somatostatin, diet (FM, PEG) No IVS1011G.T T1531Af Hypo-N22 M 4500/39 Diazoxide Yes (S) IVS6-3C.G, I462V Q917X Hypo-N23 F 4860/38 Insulin Yes (S) P1413Lg IVS1011G.Tg Hypo-N25 M 3910/34 Insulin Yes (S) V21Dg E490Xg Hypo-N26 M 3790/35 Diet (FM, PEG) Yes (H) V187D R248X Hypo-N29 F 3350/37 None Yes (P) - IVS1011G.T Hypo-N30 F 3800/37 Diazoxide No W231R L503P Hypo-N31 M 4340/40 None Yes (P) - R1493W a All cases had birth weights 12 standard deviation scores except for Hypo-N29 whose score was 11. b Current therapy is given. Login to comment
122 We classified the mutations as either MnMn Hypo-N3 R1493W MMMM Mn Mn Hypo-N6 V21D MM MnMn Hypo-N8 G1400R / R1493W MM nnMn Hypo-N9 IVS10 Mn Hypo-N11 G1478R Mn nnMn Hypo-N16 C267X Mn Mn Hypo-N19 IVS10 / T1531A MM Mn nn Hypo-N29 IVS10 Mn Mn Hypo-N30 W231R / L503P MM MM x Hypo-N23 IVS10 / P1413L MM x Hypo-N14 IVS10 Mn Hypo-N22 IVS6 (I462V) / Q917X MM Hypo-N25 V21D / E490X MM xx Hypo-N26 V187D / R248 X MM x Hypo-N31 R1493W nnMnMnMn nnnnMn MnMn MM MnMn Mn nnnn Fig. 1. Login to comment
133 ABCC8 mutations found in Norwegian CHI patientsa Nucleotide change Location Amino acid change Mutation type PSIC score PD Number of families Reference c.62 T.A Exon 1 V21D Mis 1.96 PoD 2 (24) c.560 T.A Exon 4 V187D Mis 2.01 PrD 1 (2) c.691 T.C Exon 5 W231R Mis 4.03 PrD 1 NR c.742 C.T Exon 5 R248X Non - - 1 (34, 42) c.801 C.A Exon 5 C267X Non - - 1 NR IVS6-3C.G Intron 6 - AS - - 1 NR c.1384 A.G Exon 9 I462V Mis 0.62 PrB 1 NR c.1468 G.T Exon 10 E490X Non - - 1 (43) c.1508 T.C Exon 10 L503P Mis 2.36 PrD 1 (24) IVS1011G.T Intron 10 - AS - - 5 (44) c.2749 C.T Exon 23 Q917X Non - - 1 NR c.4198 G.A Exon 35 G1400R Mis 2.37 PrD 1 (42) c.4238 C.T Exon 35 P1413L Mis 2.76 PrD 1 (25) c.4432 G.A Exon 37 G1478R Mis 2.37 PrD 1 (14, 31) c.4477 C.T Exon 37 R1493W Mis 2.79 PrD 3 (26) c.4591 A.G Exon 38 T1531A Mis 1.93 PoD 1 NR AS, aberrant splicing; Mis, missense; NR, not previously reported; Non, nonsense; PD, pathogenic description; PoD, possibly damaging; PrB, predicted to be benign; PrD, probably damaging; PSIC, position-specific independent counts. Login to comment
168 A largenumber (.150)ofABCC8 alterations have been reported to cause CHI (19) including 10 of the mutations observed in this study (V21D, V187D, R248X, E490X, L503P, IVS1011G.T, G1400R, P1413L, G1478R, and R1493W). Login to comment
187 One example is V187D that is located in TMD0 and was found in family Hypo-N26. Login to comment
189 V187D is a founder mutation in Finland (2), and further investigation revealed that the mother of the proband was of Finnish ancestry. Login to comment

PMID: 23316740 [PubMed] Sampson HM et al: "Compounds that correct F508del-CFTR trafficking can also correct other protein trafficking diseases: an in vitro study using cell lines."

No. Sentence Comment

24 We investigated mutants from a diverse set of well-studied protein trafficking diseases including the nephrogenic diabetes insipidus mutations V206D and L292P in the arginine-vasopressin receptor 2 (AVPR2, also known as V2R) [24,25], the LQTS2 mutations G601S and F805C in the human ether-a-go-go-related gene (KCNH2, also known as hERG) [26,27], and finally the persistent hyperinsulinemic hypoglycemia of infancy (PHHI, also known as congenital hyperinsulinism) mutations A116P and V187D in the sulfonylurea receptor 1 (ABCC8, also known as SUR1) [10,28,29], a component of the KATP channel. Login to comment
34 FLAG-tagged hamster SUR1 WT, A116P and V187D mutants and rat Kir6.2 plasmids have been described previously [32]. Login to comment
120 6 F508del-CFTR correctors correct the trafficking of SUR1 mutants V187D and A116P. Login to comment
121 Representative immunoblots are shown for SUR1 V187D (A-C) and SUR1 A116P (D-F). Login to comment
122 Representative immunoblot for cells expressing SUR1 V187D treated with decreasing concentrations of ouabain (ouab), KM57, and RDR1 (A), KM60, ABT-888, and latonduine (latond) (B), glafenine and carbamazepine (carbam) (C). Login to comment
125 Lanes where Kir6.2 and SUR1 V187D or SUR1 A116P are expressed are indicated by a line above the corresponding lanes. Login to comment
131 We next tested the SUR1 mutants V187D and A116P for correction with F508del-CFTR correctors. Login to comment
158 Interestingly, sulfonylureas correct the trafficking of A116P and V187D mutants by binding to sites outside the affected domain in SUR1, and possibly also with a weak affinity site in Kir6.2 [36]. Login to comment
167 Table 1 F508del-CFTR corrector compounds show distinct profiles of correction for other ER-retained proteins Corrector CFTR F508del hERG G601S hERG F805C SUR1 A116P SUR1 V187D V2R L292P V2R V206D VRT-325 + + - - ND - ND Glycerol + ND ND + ND ND +/- 29&#b0;C ++ + + + + ++ ++ KM60 + + - + + - + KM57 +/- ++ - +/- - - +/- ABT-888 + - ND + + - + Glafenine + + - + - - - RDR1 + - - + + - - Ouabain + +/-* +* + + + + Carbamazepine + - ND - +/- - ND Latonduine + +/- - + ++ +/- + Astemizole ND + - ND ND ND ND Glibenclamide ND ND ND ++ ++ ND ND A qualitative assessment of correction as determined by glycosylation status in immunoblotting is shown for each mutation following treatment with a corrector compound, where "-" indicates no correction observed, "+/-" indicates slight correction, "+" and "++" indicate more and best correction observed, respectively, and "ND" indicates not determined. Login to comment

PMID: 23744072 [PubMed] Chen PC et al: "Carbamazepine as a novel small molecule corrector of trafficking-impaired ATP-sensitive potassium channels identified in congenital hyperinsulinism."

No. Sentence Comment

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
126 Dose-response relationships were further determined for F27S, A116P, and V187D. Login to comment
136 In surface protein biotinylation experiments, there was a significant increase in biotinylated F27S, A116P, or V187D SUR1 in cells treated with carbamazepine or glibenclamide as compared with cells treated with vehicle alone (Fig. 3A). Login to comment
138 Surface staining of FLAG-tagged (N terminus) SUR1 showed a clear increase in surface expression of the F27S mutant upon Carbamazepine as a Novel KATP Channel Corrector JULY 19, 2013ߦVOLUME 288ߦNUMBER 29 JOURNAL OF BIOLOGICAL CHEMISTRY 20945 carbamazepine treatment, resembling that seen in cells treated with the sulfonylurea drug tolbutamide (Fig. 3B). 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
128 Dose-response relationships were further determined for F27S, A116P, and V187D. Login to comment

PMID: 26335336 [PubMed] Zhang Y et al: "5'-adenosine monophosphate mediated cooling treatment enhances DeltaF508-Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) stability in vivo."

No. Sentence Comment

20 Two mutations in the pancreatic ATP-sensitive potassium channels A116P and V187D, located in the SUR1 subunit, reduce channel activity leading to persistent infancy hyperinsulinemic hypoglycemia. Login to comment

PMID: 21185999 [PubMed] Saint-Martin C et al: "KATP channel mutations in congenital hyperinsulinism."

No. Sentence Comment

73 Missense, frameshift, nonsense, insertions/deletions, splice-site mutations, and rare exonic deletions have been reported in CHI.7,15 No "hot spots" for mutations are present; most mutations are unique to the respective family except for 2 mutations common (F1388del and c.3992-9G b0e; A) in the Ashkenazi Jewish population18 and 2 founder mutations (V187D and E1507K) in the Finnish population.19,20 The frequency of KATP mutations identified is different according to the histopathological form and the responsiveness of CHI patients to diazoxide. Login to comment

PMID: 23345197 [PubMed] Kapoor RR et al: "Clinical and molecular characterisation of 300 patients with congenital hyperinsulinism."

No. Sentence Comment

168 Two ABCC8 mutations, c.3992-9GOA and p.F1388del, are associated with CHI in the Ashkenazi Jewish population (11) and the p.V187D mutation has been associated with the Finnish population (46). Login to comment

PMID: 24399968 [PubMed] Martin GM et al: "Pharmacological rescue of trafficking-impaired ATP-sensitive potassium channels."

No. Sentence Comment

214 Among the mutations documented, A116P- and V187D-SUR1, both located in TMD0, exhibited reduced association with Kir6.2 in co-immunoprecipitation experiments (Chan et al., 2003), supporting a role of TMD0 in subunit-subunit interactions. Login to comment
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
248 Subsequent work identified additional SUR1 mutations in CHI patients that impair the proper trafficking of KATP channels, including L1544P, A116P, and V187D (Taschenberger et al., 2002; Yan et al., 2004). Login to comment
250 Yan et al. (2004) demonstrated that two CHI mutations, A116P and V187D, both located in the first transmembrane domain TMD0 of SUR1, could be rescued by sulfonylureas in vitro. Login to comment
254 Previously, Chan et al. showed that TMD0 domain of SUR1 harboring the A116P or V187D mutation, had reduced association with Kir6.2 in co-immunoprecipitation experiments (Chan et al., 2003). Login to comment
256 Yan et al. showed, however, that the trafficking defect in A116P and V187D is intrinsic to SUR1. Login to comment
257 This is based on the observation that in the absence of Kir6.2, A116P and V187D also prevented Kir6.2-independent surface expression of a SUR1 protein in which the RKR ER retention signal is inactivated by mutation to AAA (SUR1AAA). Login to comment
259 Yet mutation of these signals in both subunits also failed to improve surface expression of the A116P or V187D mutants. Login to comment
261 Consistent with this notion, channel trafficking defects caused by A116P and V187D could be overcome by culturing cells at lower temperature (Yang et al., 2005), a condition known to facilitate protein folding. Login to comment
266 Accordingly, mutation of S1238 to tyrosine abolished tolbutamide rescue of SUR1 mutants A116P and V187D. Login to comment
305 A more recent study by Sampson et al. tested the effects of multiple CFTR correctors identified in a chemical library screen (Carlile et al., 2007) on the processing efficiency of two SUR1 trafficking mutants, A116P and V187D (Sampson et al., 2013). Login to comment

PMID: 25637631 [PubMed] Devaraneni PK et al: "Structurally distinct ligands rescue biogenesis defects of the KATP channel complex via a converging mechanism."

No. Sentence Comment

148 TMD0 trafficking mutations F27S, A116P, and V187D used in the study as well as the ER retention motif RKR are also shown. Login to comment
163 For this set of experiments, SUR1-TMD0 trafficking mutations A116P and V187D, which we have shown previously to respond to GBC and CBZ rescue (20, 22, 23, 41, 44), were used as examples. Login to comment
164 In SUR1RKR3AAA bearing A116P or V187D expressed without Kir6.2, both exhibited only the core-glycosylated lower band, in contrast to WT-SUR1RKR3AAA, which showed both lower and upper bands; treatment with CBZ failed to correct the mutant SUR1 processing defects (Fig. 3A). Login to comment
167 Interestingly, we noted that CBZ treatment significantly enhanced the core-glycosylated A116P- and V187D- SUR1RKR3AAA band intensity even in the absence of Kir6.2. Login to comment