ATP inhibition of K_ATP channels: control of nucleotide sensitivity by the N‐terminal domain of the Kir6.2 subunit

Abstract

ATP-sensitive potassium channels (KATP) are unique among K¤ channels in being rapidly and reversibly inhibited by the non-hydrolytic binding of cytoplasmic adenine nucleotides (Noma, 1983;Ashcroft, 1988;. The realization that these channels are encoded by an ATP binding cassette (ABC) protein, the sulphonylurea receptor (SUR1 or SUR2), in addition to an inward rectifier K¤ channel subunit (Kir6.1 or Kir6.2; Inagaki et al. 1995Inagaki et al. , 1996 led to the natural presumption that inhibitory ATP binding should occur at the nucleotide binding folds (NBFs) of the SUR subunit. Mutagenic studies have now demonstrated that the NBFs of SUR1 control activation of the channel by Mg¤-bound nucleotides and diazoxide, probably as a consequence of these agents mimicking, or enhancing, an effect of nucleotide hydrolysis (Nichols et al. 1996;Gribble et al. 1997;Shyng et al. 1997b). However, none of these studies has demonstrated an effect of SUR1 mutations on the intrinsic sensitivity of expressed channels to ATP inhibition. On the other hand, tandem dimeric constructs of SUR1 and Kir6.2 express channels with reduced ATP sensitivity (Clement et al. 1997;Shyng & Nichols, 1997), and, moreover, mutations of Kir6.2 can alter the inhibitory effect of ATP on channels formed by co-expression with SUR1 (Shyng et al. 1997a;Tucker et al. 1997Tucker et al. , 1998. In the study of Shyng et al. (1997a), mutations of asparagine 160 in the second transmembrane segment alter sensitivity to polyamine-induced rectification, as well as decreasing the apparent ATP sensitivity approximately 5-fold. Tucker et al. (1997) reported that a C-terminally truncated Kir6.2 subunit (Kir6.2ÄC36) could express ATPsensitive channels in the absence of SUR1, and reported a significant reduction of ATP sensitivity for Kir6.2ÄC36 channels with an additional mutation of lysine 185 in the truncated C-terminus. Lorenz et al. (1998) 1. To gain insight into the role of the cytoplasmic regions of the Kir6.2 subunit in regulating channel activity, we have expressed the sulphonylurea receptor SUR1 with Kir6.2 subunits containing systematic truncations of the N-and C-termini. Up to 30 amino acids could be truncated from the N-terminus, and up to 36 amino acids from the C-terminus without loss of functional channels in co-expression with SUR1. Furthermore, Kir6.2ÄC25 and Kir6.2ÄC36 subunits expressed functional channels in the absence of SUR1. 2. In co-expression with SUR1, N-terminal truncations increased Ki,ATP ([ATP] causing halfmaximal inhibition of channel activity) by as much as 10-fold, accompanied by an increase in the ATP-insensitive open probability, whereas the C-terminal truncations did not affect the ATP sensitivity of co-expressed channels. 3. A mutation in the near C-terminal region, K185Q, reduced ATP sensitivity of co-expressed channels by approximately 30-fold, and on the Kir6.2ÄN2-30 background, this mutation decreased ATP sensitivity of co-expressed channels by approximately 400-fold. 4. Each of these mutations also reduced the sensitivity to inhibit...

The Common Single Nucleotide Polymorphism E23K in KIR6.2 Sensitizes Pancreatic β-Cell ATP-Sensitive Potassium Channels Toward Activation Through Nucleoside Diphosphates

“…In particular, E23K markedly affects channel gating, significantly reducing the time spent in long interburst closed states and thereby producing an evident increase of spontaneous open probability (P O ). Consistent with the idea that nucleotide-induced channel inhibition results from interaction with the interburst closed states (12,13), sensitivity toward inhibitory ATP 4Ϫ was found to be decreased (11).…”

Nucleotide Sensitivity of Pancreatic ATP-Sensitive Potassium Channels and Type 2 Diabetes

“…Importantly, an in vitro model for the heterozygous genotype (E/K, with E in position 23 of K IR 6.2 in one allele and K in the other) resulted in intermediate P O values. Consistent with the idea that nucleotide-induced channel inhibition results from interaction with the interburst closed states (39,40), E23K decreased sensitivity toward inhibitory ATP 4Ϫ in the models for the heterozygous and homozygous genotypes (K/K, with K in position 23 of K IR 6.2 in both alleles). Stimulation of insulin secretion requires reduction of the P O of pancreatic ␤-cell K ATP channels to values Ͻ0.02 (25,41).…”

Characterisation of new KATP-channel mutations associated with congenital hyperinsulinism in the Finnish population

“…Kir6.2/SUR1-L1551V currents showed reduced sensitivity to ATPinhibition, loss of activation by MgADP and diazoxide and impaired block by tolbutamide. The reduced ATP sensitivity and tolbutamide block are consistent with the increased channel open probability, as similar effects have been observed when the open probability was increased by certain mutations [25], by N-terminal truncation of Kir6.2 [29,30], or by increasing the concentration of PIP 2 [31]. The inhibitory effect of MgADP on Kir6.2/SUR1-L1551V currents was less than that observed when MgADP activation was abolished by mutation of the Walker A lysines in SUR1 [32,33].…”