ABCMdb

PMID: 18346985

Tarasov AI, Nicolson TJ, Riveline JP, Taneja TK, Baldwin SA, Baldwin JM, Charpentier G, Gautier JF, Froguel P, Vaxillaire M, Rutter GA
A rare mutation in ABCC8/SUR1 leading to altered ATP-sensitive K+ channel activity and beta-cell glucose sensing is associated with type 2 diabetes in adults.
Diabetes. 2008 Jun;57(6):1595-604. Epub 2008 Mar 17., [PubMed]

Sentences

No. Mutations Sentence Comment

6 ABCC8 p.Tyr356Cys RESULTS-A mutation in ABCC8/SUR1, leading to a Y356C substitution in the seventh membrane-spanning ␣-helix, was observed in a patient diagnosed with hyperglycemia at age 39 years and in two adult offspring with impaired insulin secretion. Login to comment 7 ABCC8 p.Tyr356Cys Single KATP channels incorporating SUR1-Y356C displayed lower sensitivity to MgATP (IC50 ϭ 24 and 95 ␮mol/l for wild-type and mutant channels, respectively). Login to comment 9 ABCC8 p.Tyr356Cys Overexpression of SUR1-Y356C in INS1(832/13) cells impaired glucose-induced cell depolarization and increased in intracellular free Ca2ϩ concentration, albeit more weakly than neonatal diabetes-associated SUR1 mutants. Login to comment 23 ABCC8 p.Tyr356Cys Through electrophysiology and Ca2ϩ imaging, we demonstrate that one of the mutations, Y356C, affects the ATP sensitivity of KATP channels and glucose-induced Ca2ϩ influx but to a far smaller extent than TND-associated mutations. Login to comment 83 ABCC8 p.His1023Tyr ABCC8 p.His1023Tyr ABCC8 p.Leu582Val ABCC8 p.Leu582Val ABCC8 p.Tyr356Cys ABCC8 p.Arg1379Cys ABCC8 p.Arg1379Cys ABCC8 p.Arg248Gln ABCC8 p.Arg248Gln ABCC8 p.Lys1521Asn (E), wild type; (F), Y356C; (f), K1521N; (Ⅺ), H1023Y; (Œ), R248Q; (‚), L582V; (ૺ), R1379C. Login to comment 88 ABCC8 p.Leu582Val ABCC8 p.Tyr356Cys A: Currents from inside-out patches excised from HEK293 cells overexpressing recombinant Kir6.2/SUR1-wild type (WT), Kir6.2/SUR1-Y356C, and Kir6.2/SUR1-L582V in Mg2؉ -containing (left) and Mg2؉ -free (right) solution. Login to comment 92 ABCC8 p.Tyr356Cys B: MgATP concentration-inhibition curves for wild-type and Kir6.2/SUR1-Y356C KATP channels. Login to comment 93 ABCC8 p.Tyr356Cys C: ATP (Mg2؉ -free) concentration-inhibition curves for wild-type and Kir6.2/SUR1-Y356C KATP channels. Login to comment 94 ABCC8 p.Leu582Val D: MgATP concentration-inhibition curves for wild-type and Kir6.2/SUR1-L582V KATP channels. Login to comment 95 ABCC8 p.Leu582Val E: ATP (Mg2؉ -free) concentration-inhibition curves for wild-type and Kir6.2/SUR1-L582V KATP channels. Login to comment 113 ABCC8 p.Tyr356Cys ABCC8 p.Tyr356Cys One of the patients with normal BMI, diagnosed with hyperglycemia at age 39 years, having developed overt diabetes at age 45 years, presented an ABCC8 missense mutation causing a substitution of tyrosine 356 with cysteine (Y356C) in the SUR1 subunit of the KATP channel (Fig. 1B and online appendix Fig. 1). Login to comment 116 ABCC8 p.Tyr356Cys The Y356C mutation was not found in 170 unrelated normoglycaemic individuals of European Caucasian origin. Login to comment 117 ABCC8 p.Arg248Gln ABCC8 p.Lys1521Asn The two other ABCC8 mutations that we found to be associated with adult-onset diabetes were R248Q (type 2 diabetic patient diagnosed at age 39 years without familial cosegregation) (online appendix Fig. 1) and K1521N (two type 2 diabetic patients diagnosed at age 37 and 42 years). Login to comment 120 ABCC8 p.His1023Tyr ABCC8 p.Leu582Val ABCC8 p.Tyr356Cys ABCC8 p.Arg1379Cys ABCC8 p.Arg248Gln ABCC8 p.Lys1521Asn To test whether the mutations associated with type 2 diabetes might affect stimulus-secretion coupling in beta-cells, we next measured the sensitivity to ATP of recombinant KATP channels carrying SUR-Y356C, -R248Q, and -K1521N and compared these to the ATP sensitivity of TND-associated mutants (4), L582V, H1023Y, and R1379C. Login to comment 125 ABCC8 p.Arg248Gln ABCC8 p.Lys1521Asn The concentration-inhibition curves for KATP channels carrying SUR1-R248Q and SUR1-K1521N were practically identical to the wild type, suggesting either that these mutations affected other properties of the channel or were not responsible for diabetes (Fig. 1C). Login to comment 126 ABCC8 p.Lys1521Asn KATP channel conductance of the inside-out patches expressing SUR1-K1521N was not different from wild type (11.3 Ϯ 5.6 ns and 12.5 Ϯ 5.9 ns, respectively), as measured in nucleotide-free solution. Login to comment 127 ABCC8 p.Arg248Gln Patches with SUR1-R248Q channels exhibited much smaller conductances of 1.2 Ϯ 0.8 ns. Login to comment 129 ABCC8 p.Tyr356Cys By contrast, KATP channels carrying SUR1-Y356C showed an approximately fourfold decrease in ATP sensitivity (Fig. 1C). Login to comment 130 ABCC8 p.Tyr356Cys This prompted us to investigate in detail how the Y356C mutation affected the ATP sensitivity and/or surface expression of KATP channels. Login to comment 132 ABCC8 p.Leu582Val Given the limited magnitude of the type 2 diabetes-associated mutant`s effects, we used a TND-associated SUR1 mutation, L582V (4), as a positive control. Login to comment 138 ABCC8 p.Tyr356Cys ABCC8 p.Tyr356Cys The ATP sensitivity of heterozygous SUR1-Y356C (hetY356C) (Fig. 2A and B) (Table 1) was higher than that of homozygous SUR1-Y356C (homY356C) channels. Login to comment 140 ABCC8 p.Leu582Val ABCC8 p.Leu582Val Heterozygous channels expressing SUR1-L582V (hetL582V) were also more ATP sensitive than homozygous SUR1-L582V channels (homL582V): IC50 ϭ 869 ␮mol/l and IC50 ϭ 1,140 ␮mol/l for het582V and hom582V, respectively (Fig. 2A and D) (Table 1). Login to comment 143 ABCC8 p.Tyr356Cys Gain-of-function mutations in either subunit frequently act by enhancing the Mg2ϩ -dependent activation (4,27), so we tested if this was the case for SUR1-Y356C. Login to comment 147 ABCC8 p.Leu582Val ABCC8 p.Tyr356Cys Thus, the gain-of-function effect of L582V mutation was mediated via Mg2ϩ -dependent activation, while the effect of Y356C apparently occurred through a different mechanism. Login to comment 148 ABCC8 p.Tyr356Cys Y356C does not alter surface expression of KATP channels. Login to comment 151 ABCC8 p.Tyr356Cys Introduction of the Y356C mutation into SUR1 did not affect cytoplasmic (Fig. 3C) or membrane (Fig. 3D) localization. Login to comment 152 ABCC8 p.Leu582Val Similarly, the cytoplasmic disposition of SUR1-L582V was not different from that of the wild-type SUR1 (Fig. 3E). Login to comment 153 ABCC8 p.Leu582Val Interestingly, we did note a tendency toward lower cell surface expression of SUR1-L582V (Fig. 3F) versus wild type. Login to comment 158 ABCC8 p.Leu582Val ABCC8 p.Tyr356Cys We therefore studied the effect on these two phenomena of the Y356C and L582V mutations in SUR1. Login to comment 171 ABCC8 p.Tyr356Cys ABCC8 p.Tyr356Cys ABCC8 p.Tyr356Cys This effect is not typical for native beta-cells, where the firing frequency increases monotonously, with the KATP TABLE 1 Clinical characteristics of nondiabetic carriers of the SUR1-Y356C mutation compared with normoglycemic control subjects Daughter* Son* Control subjects (n ϭ 18) Age at examination (years) 35 33 26.79 Ϯ 6.56 BMI (kg/m2 ) 19.37 22.22 22.9 Ϯ 3.3 Fasting blood concentrations Glucose (mmol/l) 4.9 5.0 4.6 Ϯ 0.3 Insulin (␮U/ml) 1.33 4.11 5.5 Ϯ 3.8 Insulinogenic index (␮UI/␮mol)† 2.58 7.95 16.9 Ϯ 10.7 Insulin sensitivity (mg ⅐ kg-1 ⅐ min-1 )‡ 12.32 6.71 10.88 Ϯ 2.36 Disposition index (␮UI/mol)§ 31.78 53.34 184.21 Ϯ 25.16 Glycemic status Nondiabetic Nondiabetic Nondiabetic Data are means Ϯ SD. *Daughter and son of the diabetic proband identified with the Y356C mutation. Login to comment 177 ABCC8 p.Leu582Val ABCC8 p.Tyr356Cys TABLE 2 Parameters of ATP inhibition for KATP channels with mutant SUR1 subunit IC50 WT Y356C L582V ЉheteroЉ ЉhomoЉ ЉheteroЉ ЉhomoЉ 2 Mg2ϩ mmol/l 24 Ϯ 3 (5) 61 Ϯ 11 (10)* 95 Ϯ 9 (10)* 869 Ϯ 48 (6)* 1140 Ϯ 346 (6)* 0 Mg2ϩ mmol/l 8 Ϯ 1 (6) 25 Ϯ 5 (7)* 38 Ϯ 8 (8)* 17 Ϯ 3 (5)† 17 Ϯ 3 (6)† Data are means Ϯ SE (number of experiments). Login to comment 182 ABCC8 p.Tyr356Cys Thus, despite a relatively small shift in the ATP sensitivity of KATP channels, beta-cell lines expressing SUR1-Y356C demonstrated impaired coupling between nutrient stimulation and electrical activity. Login to comment 186 ABCC8 p.Tyr356Cys The addition of 20 mmol/l glucose had different effects on the two groups: in the majority of wild-type cells we observed oscillations of [Ca2ϩ ]cyt, which were not detected in Y356C cells. Login to comment 187 ABCC8 p.Lys1521Asn INS1(832/13) cells overexpressing SUR1-K1521N channels showed an unchanged [Ca2ϩ ]cyt response to glucose compared with wild-type cells (Fig. 5B and C). Login to comment 190 ABCC8 p.Tyr356Cys In the case of SUR1-Y356C this was indeed observed. Login to comment 195 ABCC8 p.Tyr356Cys However, the Y356C mutation that we describe appears to cause diabetes outside the range described (34) (maximum age recorded A C E D F B FIG. 3. Login to comment 198 ABCC8 p.Leu582Val ABCC8 p.Tyr356Cys A, C, and E: Representative confocal images of cells expressing the SUR1 (wild-type [A], Y356C [C], and L582V [E]) subunit alone. Login to comment 200 ABCC8 p.Leu582Val ABCC8 p.Tyr356Cys B, D, and F: Representative images of cells expressing SUR1 (wild-type [B], Y356C [D], and L582V [F]) subunits together with Kir6.2. Login to comment 205 ABCC8 p.Tyr356Cys ABCC8 p.Tyr356Cys The relatively small shift in ATP sensitivity (from 24 to 95 ␮mol/l as measured in inside-out patches) caused by the Y356C mutation in ABCC8/SUR1, clearly affected glucose-induced changes in beta-cell electrical activity. Login to comment 206 ABCC8 p.Tyr356Cys This result strongly suggests that the Y356C mutation may lead to a diabetic phenotype. Login to comment 207 ABCC8 p.Tyr356Cys Indeed, the oral glucose tolerance test and euglycemic- hyperinsulinic clamp performed in the two nondiabetic carriers of the Y356C mutation showed a mild decrease of insulinogenic and disposition indexes (Table 1). Login to comment 210 ABCC8 p.Leu582Val ABCC8 p.Tyr356Cys We observed that two mutations (Y356C and L582V) that are associated with phenotypes of different severity in heterozygous patients cause different shifts in the ATP sensitivity of the KATP channel (Fig. 2B and D). Login to comment 221 ABCC8 p.Leu582Val ABCC8 p.Tyr356Cys Thus, the Y356C mutant lead to a substantially less marked inhibition of glucose-induced [Ca2ϩ ]cyt increase than L582V (Fig. 5B). Login to comment 224 ABCC8 p.Tyr356Cys Molecular mechanism of Y356C effect on ATP sensitivity. Login to comment 245 ABCC8 p.Tyr356Cys involving the SUR1 NBDs, would be consistent with the observation that the removal of Mg2ϩ (which abolishes the activatory effect of adenine-nucleotides on NBDs) (24) did not abolish the activatory effect of Y356C (Fig. 3C). Login to comment 248 ABCC8 p.His1023Tyr ABCC8 p.Leu582Val ABCC8 p.Tyr356Cys In contrast to Y356C, the activatory effect of mutations L582V (Fig. 3D) and H1023Y (4) was not observed under Mg2ϩ -free conditions, suggesting that these mutations exert their effects via the SUR1 NBDs. Login to comment