ABCMdb

ABCC8 p.Gly70Glu

Predicted by SNAP2:	A: D (53%), C: D (66%), D: D (80%), E: D (66%), F: D (63%), H: D (71%), I: N (66%), K: D (71%), L: N (78%), M: D (53%), N: N (61%), P: D (75%), Q: D (63%), R: D (66%), S: N (61%), T: D (53%), V: N (53%), W: D (85%), Y: D (66%),
Predicted by PROVEAN:	A: D, C: D, D: D, E: D, F: D, H: D, I: D, K: D, L: D, M: D, N: D, P: D, Q: D, R: D, S: D, T: D, V: D, W: D, Y: D,

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[hide] Hyperinsulinism of infancy: novel ABCC8 and KCNJ11... J Clin Endocrinol Metab. 2004 Dec;89(12):6224-34. Tornovsky S, Crane A, Cosgrove KE, Hussain K, Lavie J, Heyman M, Nesher Y, Kuchinski N, Ben-Shushan E, Shatz O, Nahari E, Potikha T, Zangen D, Tenenbaum-Rakover Y, de Vries L, Argente J, Gracia R, Landau H, Eliakim A, Lindley K, Dunne MJ, Aguilar-Bryan L, Glaser B
Hyperinsulinism of infancy: novel ABCC8 and KCNJ11 mutations and evidence for additional locus heterogeneity.
J Clin Endocrinol Metab. 2004 Dec;89(12):6224-34., [PMID:15579781]

Abstract [show]
Hyperinsulinism of infancy is a genetically heterogeneous disease characterized by dysregulation of insulin secretion resulting in severe hypoglycemia. To date, mutations in five different genes, the sulfonylurea receptor (SUR1, ABCC8), the inward rectifying potassium channel (K(IR)6.2, KCNJ11), glucokinase (GCK), glutamate dehydrogenase (GLUD1), and short-chain 3-hydroxyacyl-coenzyme A dehydrogenase (SCHAD), have been implicated. Previous reports suggest that, in 40% of patients, no mutation can be identified in any of these genes, suggesting additional locus heterogeneity. However, previous studies did not screen all five genes using direct sequencing, the most sensitive technique available for mutation detection. We selected 15 hyperinsulinism of infancy patients and systematically sequenced the promoter and all coding exons and intron/exon boundaries of ABCC8 and KCNJ11. If no mutation was identified, the coding sequence and intron/exon boundaries of GCK, GLUD1, and SCHAD were sequenced. Seven novel mutations were found in the ABCC8 coding region, one mutation was found in the KCNJ11 coding region, and one novel mutation was found in each of the two promoter regions screened. Functional studies on beta-cells from six patients showed abnormal ATP-sensitive K+ channel function in five of the patients; the sixth had normal channel activity, and no mutations were found. Photolabeling studies using a reconstituted system showed that all missense mutations altered intracellular trafficking. Each of the promoter mutations decreased expression of a reporter gene by about 60% in a heterologous expression system. In four patients (27%), no mutations were identified. Thus, further genetic heterogeneity is suggested in this disorder. These patients represent a cohort that can be used for searching for mutations in other candidate genes.

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101 ABCC8 -64 c3g Promoter gcc gcc ccc Promoter 11 gGc G70E 2 ccc ggg cac Missense 5 gAg G111R 3 gcc ggg atg Missense 3 Agg 2154 ϩ 3a3g Intron 15 agg tat ggc Splice-site 6, 10 tGt R836X 21 cag cga atc Nonsense 2 Tga 1113 ins T 27 ttt ttt gag Single-base insertion 7 ttt ttt Tga 3992-9 g3a Intron 32 cgc aag cgt Splice-site 1 aaA G1342E 33 caa ggg aag Missense 6 gAg R1419H 35 ctg cgc tca Missense 5 cAc R1494W 37 gcc cgg gcc Missense 4 Tgg KCBJ11 ϩ88 g3t Promoter gaa gtg agg Promoter 9 Ttg P254L Exon 1 gcc cCg ctg Missense 8 cTg a For each mutation, the upper line indicates the wt sequence, and the lower line indicates the mutant sequence. Login to comment
164 When the mutations in ABCC8 (G70E, G111R, R836X, G1343E, R1419H, and R1494W) were expressed alone, all except R836X were photolabeled with the high-affinity sulfonylurea ligand [125 I]- azido-glibenclamide (data not shown). Login to comment
180 In contrast, the mature form, which is indicative of channels trafficking at least to the Golgi, was present only in the G70E homozygote, in the G70E/R1419H compound heterozygote, and to a lesser extent, in G111R. Login to comment
185 The first three SUR1 mutant channels, G70E homozygote, G70E/ R1419H compound heterozygote, and G111R, were expressed at the plasma membrane, albeit at much lower levels than the wild-type channel. Login to comment
188 We confirmed that channel activity of the G70E and G111R mutants was decreased when compared with wild-type controls. Login to comment
190 Patient 5 was a compound heterozygote for mutations G70E and R1419H. Login to comment
236 COS cells coexpress wild-type (wt) KIR6.2 with SUR1 mutations (G70E, G70E/R1419H, G111R, G1343E, R1419H, and R1494W) or coexpress wt SUR1 with KIR6.2 mutation (P254L). Login to comment
242 Homozygous expression of G70E and G111R had reduced surface expression, whereas mutations R836X, G1343E, R1419H, R1494W, and P254L did not reach the plasma membrane at all, although they did associate with their respective wt partner as shown in Fig. 5A (lower panel). Login to comment
243 Compound heterozygous expression for mutation G70E/R1419H (4 ␮g/4 ␮g) showed reduced surface expression when compared with mutation G70E (8 ␮g) expressed alone. Login to comment
250 Only G70E (8 ␮g), G111R (8 ␮g), and compound heterozygote G70E/R1419H (4 ␮g/4 ␮g) reconstituted with wt KIR6.2 (1 ␮g) show reduced efflux. Login to comment
280 In the current study, we describe two different missense mutations in the same domain, G70E and G111R. Login to comment

[hide] In vitro recovery of ATP-sensitive potassium chann... Diabetes. 2011 Apr;60(4):1223-8. Epub 2011 Mar 16. Powell PD, Bellanne-Chantelot C, Flanagan SE, Ellard S, Rooman R, Hussain K, Skae M, Clayton P, de Lonlay P, Dunne MJ, Cosgrove KE
In vitro recovery of ATP-sensitive potassium channels in beta-cells from patients with congenital hyperinsulinism of infancy.
Diabetes. 2011 Apr;60(4):1223-8. Epub 2011 Mar 16., [PMID:21411514]

Abstract [show]
OBJECTIVE: Congenital hyperinsulinism in infancy (CHI) is characterized by unregulated insulin secretion from pancreatic beta-cells; severe forms are associated with defects in ABCC8 and KCNJ11 genes encoding sulfonylurea receptor 1 (SUR1) and Kir6.2 subunits, which form ATP-sensitive K(+) (K(ATP)) channels in beta-cells. Diazoxide therapy often fails in the treatment of CHI and may be a result of reduced cell surface expression of K(ATP) channels. We hypothesized that conditions known to facilitate trafficking of cystic fibrosis transmembrane regulator (CFTR) and other proteins in recombinant expression systems might increase surface expression of K(ATP) channels in native CHI beta-cells. RESEARCH DESIGN AND METHODS: Tissue was isolated during pancreatectomy from eight patients with CHI and from adult cadaver organ donors. Patients were screened for mutations in ABCC8 and KCNJ11. Isolated beta-cells were maintained at 37 degrees C or 25 degrees C and in the presence of 1) phorbol myristic acid, forskolin and 3-isobutyl-1-methylxanthine, 2) BPDZ 154, or 3) 4-phenylbutyrate. Surface expression of functional channels was assessed by patch-clamp electrophysiology. RESULTS: Mutations in ABCC8 were detected for all patients tested (n = 7/8) and included three novel mutations. In five of eight patients, no changes in K(ATP) channel activity were observed under different cell culture conditions. However, in three patients, in vitro recovery of functional K(ATP) channels occurred. Here, we report the first cases of recovery of defective K(ATP) channels in human beta-cells using modified cell culture conditions. CONCLUSIONS: Our study establishes the principle that chemical modification of K(ATP) channel subunit trafficking could be of benefit for the future treatment of CHI.

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64 The TABLE 1 CHI patient tissue details Patient (#) Age at surgery Histology Gene defect Genotype Reference 1 10 weeks Diffuse Presumed Homozygous, ABCC8 Unknown - 2 12 weeks Diffuse Homozygous, ABCC8 c.1467+5G.A Novel mutation 3 12 weeks Diffuse Compound Heterozygous, ABCC8 p.Arg998X/p.Ser1449dup 25 4 12 weeks Diffuse Homozygous, ABCC8 c.3992-9G.A 3 5 7 weeks Diffuse Homozygous, ABCC8 c.3992-9G.A 3 6 3.5 years Diffuse Compound Heterozygous, ABCC8 p.Gly70Glu/p.Arg1419Gly 3 7 12 months Diffuse Compound Heterozygous, ABCC8 p.Lys242fs/p.Arg1437X Novel mutations 8 4 weeks Focal Paternal uniparental isodisomy, ABCC8 p.Arg598X 3 Seven patients were found to have diffuse CHI, and one patient was defined as focal CHI. Login to comment