ABCMdb

PMID: 23739550

Smith KJ, Chadburn AJ, Adomaviciene A, Minoretti P, Vignali L, Emanuele E, Tammaro P
Coronary spasm and acute myocardial infarction due to a mutation (V734I) in the nucleotide binding domain 1 of ABCC9.
Int J Cardiol. 2013 Oct 9;168(4):3506-13. doi: 10.1016/j.ijcard.2013.04.210. Epub 2013 Jun 3., [PubMed]

Sentences

No. Mutations Sentence Comment

2 ABCC9 p.Val734Ile Here we explore the pathophysiological mechanism of a rare mutation (V734I) found in exon 17 of the ABCC9 gene, estimated to cause a 6.4-fold higher risk of AMI before the age of 60. Methods and results: Eleven patients carrying the mutation were identified; they presented AMI of vasospastic origin associated with increased plasma levels of endothelin-1 and increased leukocyte ROCK activity. Login to comment 7 ABCC9 p.Val734Ile ABCC9 p.Val734Ile The antianginal drug nicorandil activated Kir6.2/SUR2B-V734I channels, thus substituting for the loss of MgNDP stimulation, suggesting that this drug could be of therapeutic use in the treatment of AMI associated with V734I. Login to comment 38 ABCC9 p.Val734Ile ABCC9 p.Val734Ile The mutation is a non-synonymous substitution of valine 734 with isoleucine (V734I). Login to comment 42 ABCC9 p.Val734Ile In this study, we identified a total of 11 patients with early-onset AMI who carried the V734I mutation. Login to comment 49 ABCC9 p.Val734Ile In contrast, mutant Kir6.2/SUR2B channels (Kir6.2/SUR2B-V734I) presented reduced sensitivity to i) MgATP inhibition and ii) MgNDP activation assessed in the presence of MgATP. Login to comment 50 ABCC9 p.Val734Ile We investigated the molecular basis of the impaired regulation by nucleotides of Kir6.2/SUR2B-V734I channels and propose that this may involve a change in the stimulatory effect of MgATP at the NBD1 of SUR2B. Login to comment 57 ABCC9 p.Val734Ile Carriers of the V734I mutation in ABCC9 identified by genetic screening (n = 11) were enrolled in this study as cases. Login to comment 58 ABCC9 p.Val734Ile For control purposes, AMI patients without the V734I mutation were considered. Login to comment 65 ABCC9 p.Val734Ile Molecular genetic analysis All 1123 patients considered in this study were screened for the V734I mutation. Login to comment 79 ABCC9 p.Val734Ile Patient characteristics We identified a heterozygous mutation in ABCC9 (V734I) in 11 of the 1123 patients (0.98%) with precocious AMI who underwent genetic screening. Login to comment 80 ABCC9 p.Val734Ile The characteristics of the carriers of the mutation were compared with those of 33 age-and gender-matched AMI individuals who did not possess the V734I variant. Login to comment 81 ABCC9 p.Val734Ile Patients were aged between 40 and 59 years (V734I carriers) and between 41 and 59 years (control group). Login to comment 82 ABCC9 p.Val734Ile As Table 1 shows, heterozygous carriers of the V734I mutation did not differ from non-carriers in terms of body mass index, smoking, diabetes mellitus, hypertension, family history of ischemic heart disease, lipid parameters, and left ventricular ejection fraction. Login to comment 84 ABCC9 p.Val734Ile However, carriers of the V734I mutation showed increased surrogate markers of endothelial dysfunction such as raised plasma endothelin levels and increased leukocyte ROCK activity. Login to comment 85 ABCC9 p.Val734Ile Interestingly, the number of stenosed coronary arteries was significantly lower in the 11 patients presenting the V734I mutation: 7 of them demonstrated unstenosed coronary arteries while the remaining 4 presented only one stenosed artery (Table 1). Login to comment 87 ABCC9 p.Val734Ile However, none of the carriers of the V734I mutation had evidence of left ventricular apical aneurysm, left ventricular ballooning syndrome, or hypertrophic cardiomyopathy. Login to comment 93 ABCC9 p.Val734Ile ABCC9 p.Val734Ile ABCC9 p.Val734Ile Functional analysis of the V734I mutation To gain insights into the molecular mechanisms by which the V734I mutation causes AMI, we began by constructing a structural model of the human NBD1 of SUR2B harbouring the V734I mutation (Fig. 2). Login to comment 98 ABCC9 p.Val734Ile To examine the possibility that the V734I mutation affects the channel's regulation by nucleotides, we assessed the sensitivity of wild-type and mutant channels to MgATP in inside-out membrane patches. Login to comment 99 ABCC9 p.Val734Ile Fig. 3A shows that Kir6.2/SUR2A-V734I presented unaltered sensitivity to MgATP compared to Kir6.2/ SUR2A (Suppl. Table 1A). Login to comment 100 ABCC9 p.Val734Ile In contrast, Kir6.2/SUR2B-V734I channels were less sensitive to MgATP inhibition than Kir6.2/SUR2B channels: the concentration causing half-maximal block (IC50) was 214 bc;M and 96 bc;M, for mutant and wild-type channels, respectively (Fig. 3B, Suppl. Table 1B). Login to comment 106 ABCC9 p.Val734Ile Coronary angiogram carried out on a male AMI patient with the V734I mutation in the ABCC9 gene showing a focal narrowing of the right coronary artery (panel A, red arrow). Login to comment 112 ABCC9 p.Val734Ile The V734I mutation within NBD1 is represented as a magenta star. Login to comment 119 ABCC9 p.Val734Ile The V734I mutation is shown in magenta. Login to comment 122 ABCC9 p.Val734Ile A and B. i, Currents recorded in inside-out patches excised from HEK-293T cells expressing Kir6.2 and either SUR2x or SUR2x-V734I as indicated. The membrane potential was held at -80 mV. Login to comment 124 ABCC9 p.Val734Ile ABCC9 p.Val734Ile The dashed lines indicate the zero current level. ii, Mean relationship between [MgATP] and KATP current (I), expressed relative to the current in the absence of nucleotide (I0) for KATP channels composed of Kir6.2 and either SUR2x or SUR2x-V734I subunits as indicated. The continuous lines are the best fit of suppl. Eq. (1) to the data. The number of experiments was 8-11 (A) and 18 (B) in each case. C. i Currents recorded in inside-out patches excised from HEK-293T cells expressing Kir6.1 and either SUR2B or SUR2B-V734I as indicated. The membrane potential was held at -80 mV. Login to comment 127 ABCC9 p.Val734Ile The dashed lines indicate the zero current level. ii, Mean relationship between [MgATP] and KATP current (I), expressed relative to the current in the absence of nucleotide (I0) for KATP channels composed of Kir6.1 and either SUR2B or SUR2B-V734I. Login to comment 131 ABCC9 p.Val734Ile In the presence of concentrations of MgATP lower than ~30 bc;M, MgATP had a stimulatory effect on Kir6.1/SUR2B and Kir6.1/ SUR2B-V734I channels (EC50 values were 1.9 &#b1; 0.7 bc;M (n = 7) and 2.2 &#b1; 0.8 bc;M (n = 10), respectively) while at higher doses, MgATP inhibited the activity of these channels (Fig. 3C). Login to comment 132 ABCC9 p.Val734Ile Importantly, the response of wild-type (Kir6.1/SUR2B) and mutant (Kir6.1/SUR2B-V734I) channels to MgATP were unaltered (Suppl. Table 1C). Login to comment 135 ABCC9 p.Val734Ile We therefore assessed the sensitivity of Kir6.2/SUR2B and Kir6.2/SUR2B-V734I channels to ATP in the absence of Mg2+ . Login to comment 136 ABCC9 p.Val734Ile Under these conditions, ATP does not interact with SUR2 to stimulate channel activity [43]; this enables the effects of the V734I mutation on ATP inhibition at Kir6.2 to be analysed in isolation from MgATP activation at SUR2. Login to comment 138 ABCC9 p.Val734Ile ABCC9 p.Val734Ile Suppl. Fig. 1 shows that V734I does not affect the sensitivity of Kir6.2/ SUR2B channels to ATP inhibition; the IC50 and h being 13 &#b1; 3 bc;M and 1.1 &#b1; 0.1 (n = 13) (Kir6.2/SUR2B) and 10 &#b1; 2 bc;M and 1.3 &#b1; 0.1 (n = 9) (Kir6.2/SUR2B-V734I). Login to comment 139 ABCC9 p.Val734Ile Furthermore, single-channel recordings demonstrated that the mutation did not affect either the single channel current (i) or the open probability in the absence of ligands (Po), which were 4.9 &#b1; 0.1 pA and 0.34 &#b1; 0.04 (n = 11) and 4.9 &#b1; 0.2 pA and 0.42 &#b1; 0.05 (n = 11) for Kir6.2/SUR2B and Kir6.2/SUR2B-V734I, respectively (Fig. 4). Login to comment 141 ABCC9 p.Val734Ile We therefore investigated if V734I enhances the Kir6.2/ SUR2B response to MgADP, as well as the response to other MgNDPs (MgUDP and MgGDP), studied in the absence of MgATP. Login to comment 145 ABCC9 p.Val734Ile These analyses indicate that the V734I mutation had no or only a small effect on the time course of the channel response to MgNDPs. Login to comment 146 ABCC9 p.Val734Ile Another possible explanation for the rightward shift of the ATP dose-response curve for Kir6.2/SUR2B-V734I channels is that the mutation results in a conformational change in the NBD1 of SUR2B that allows MgATP to act directly as a channel activator (rather than via hydrolysis to MgADP). Login to comment 148 ABCC9 p.Val734Ile Suppl. Fig. 3 shows that Kir6.2/SUR2B and Kir6.2/SUR2B-V734I channels were inhibited to the same extent by this compound, the IC50 being 56.3 &#b1; 14.6 bc;M (n = 6) and 42.2 &#b1; 2.3 bc;M (n = 7) and h being 1.29 &#b1; 0.09 (n = 6) and 1.84 &#b1; 0.17 (n = 7), respectively. Login to comment 149 ABCC9 p.Val734Ile This supports the idea that impaired MgATP hydrolysis underlies the stimulatory effects of the V734I mutation. Login to comment 153 ABCC9 p.Val734Ile In these experiments, for both Kir6.2/SUR2A and Kir6.2/SUR2A-V734I channels, IC50 was ~140 bc;M, ~170 bc;M and ~250 bc;M, in the presence of MgADP, MgUDP and MgGDP, respectively (Suppl. Table 1A). Login to comment 154 ABCC9 p.Val734Ile However, Kir6.2/ SUR2B-V734I channels were almost insensitive to MgNDP activation, the IC50 for MgATP inhibition being ~200-400 bc;M when measured in either the absence or in the presence of intracellular MgNDPs (Suppl. Table 1B). Login to comment 156 ABCC9 p.Val734Ile Finally, for Kir6.1/SUR2B and Kir6.1/SUR2B-V734I channels, the sensitivity to MgATP was assessed in the presence of 500 bc;M MgGDP and 300 bc;M pinacidil. Login to comment 159 ABCC9 p.Val734Ile ABCC9 p.Val734Ile Response of Kir6.2/SUR2B-V734I channels to nicorandil Finally, we asked if nicorandil, an efficacious antianginal drug that acts on KATP channels [45,46], could be used to augment the sensitivity of Kir6.2/SUR2B-V734I channels to stimulatory MgNDPs in the presence of MgATP. Login to comment 160 ABCC9 p.Val734Ile The response of Kir6.2/SUR2B-V734I channels to MgATP was tested in the presence of 500 bc;M MgGDP and 30 bc;M nicorandil. Login to comment 161 ABCC9 p.Val734Ile Suppl. Fig. 4 shows that for Kir6.2/SUR2B-V734I channels IC50 values were 1364 &#b1; 350 bc;M (n = 6) and h was 1.2 &#b1; 0.1 (n = 5), respectively. Login to comment 162 ABCC9 p.Val734Ile Thus, low doses of nicorandil potently activate Kir6.2/SUR2B-V734I channels. Login to comment 164 ABCC9 p.Val734Ile Discussion The key finding of this paper is the observation that a variant of human ABCC9 (V734I), which influences susceptibility to AMI during adulthood [32], results in an altered responsiveness of KATP channels to intracellular nucleotides. Login to comment 166 ABCC9 p.Val734Ile We found that KATP channels composed of Kir6.2 and SUR2B-V734I presented a reduced sensitivity to MgATP inhibition compared to wild-type. Login to comment 169 ABCC9 p.Val734Ile ABCC9 p.Val734Ile Nicorandil activated Kir6.2/SUR2B-V734I channels, effectively substituting for the loss of MgNDP stimulation, suggesting that this drug could be efficacious in the treatment of AMI associated with V734I. Login to comment 173 ABCC9 p.Val734Ile Representative single KATP channel currents recorded at -80 mV from inside-out patches excised from HEK-293 T cells expressing Kir6.2 and either SUR2B or SUR2B-V734I, as indicated. The dashed line indicates the zero current level. Login to comment 176 ABCC9 p.Val734Ile All carriers of the V734I mutation presented with levels of inflammatory markers, such as interleukin-6, hs-CRP, TNF-b1; and adiponectin, which did not differ from those of the control group. Login to comment 178 ABCC9 p.Val734Ile Four out of the 11 carriers of the V734I mutation presented with one stenosed coronary artery. Login to comment 186 ABCC9 p.Val734Ile However, none of the carriers of the V734I mutation in our study presented with ventricular ballooning. Login to comment 189 ABCC9 p.Val734Ile ABCC9 p.Val734Ile Functional implication of mutation V734I The IC50 for MgATP inhibition of Kir6.2/SUR2B-V734I was decreased relative to that of wild-type channels. Login to comment 190 ABCC9 p.Val734Ile In the physiological range of cytoplasmic MgATP concentrations (i.e. >~0.5 mM) [53,54], however, there was only a small difference between the residual Kir6.2/SUR2B and Kir6.2/SUR2B-V734I channel currents. Login to comment 191 ABCC9 p.Val734Ile The addition of physiological MgNDPs altered the MgATP response of wild-type, but not that of Kir6.2/SUR2B-V734I channels. Login to comment 192 ABCC9 p.Val734Ile For instance, in the presence of 1 mM MgATP, the addition of 0.5 mM MgADP did not produce a significant change in Kir6.2/SUR2B-V734I current whereas the Kir6.2/SUR2B current showed a two-fold increase in amplitude. Login to comment 193 ABCC9 p.Val734Ile ABCC9 p.Val734Ile In the absence of human or animal samples of coronary arteries carrying the SUR2B-V734I mutation, it is only possible to speculate on the exact cellular consequences of the V734I mutation. Login to comment 194 ABCC9 p.Val734Ile It could be envisaged that in intact endothelial cells, where Kir6.2/SUR2B channels are expressed, mutant Kir6.2/SUR2B-V734I channels would fail to respond in the normal way to MgNDP activation under metabolic stress (Fig. 6). Login to comment 199 ABCC9 p.Val734Ile Mean relationship between [MgATP] and KATP current (I), expressed relative to the current in the absence of nucleotide (I0) for experiments conducted in the presence of 500 bc;M of individual MgNDP, as indicated, for Kir6.2/SUR2x (i) and Kir6.2/SUR2x-V734I channels (ii). Login to comment 202 ABCC9 p.Val734Ile Mean relationship between [MgATP] and KATP current (I), expressed relative to the current in the absence of nucleotide (I0) for experiments conducted in the presence of 500 bc;M MgGDP, as indicated, for Kir6.1/SUR2B (i) and Kir6.1/SUR2B-V734I channels (ii). Login to comment 209 ABCC9 p.Val734Ile The increase in circulating levels of endothelin-1 in patients carrying V734I seen here is consistent with the observation that blockage of KATP channels increased endothelin-1 secretion from isolated human coronary endothelial cells [31]. Login to comment 210 ABCC9 p.Val734Ile In contrast, under conditions of high metabolism, when the ratio between MgATP and MgADP is high, the small decrease in MgATP inhibition presented by Kir6.2/SUR2B-V734I channels might have a protective effect. Login to comment 211 ABCC9 p.Val734Ile This is consistent with the fact that patients carrying the V734I mutation suffer AMI only in adulthood. Login to comment 212 ABCC9 p.Val734Ile Another possible mechanism by which the V734I mutation may promote AMI is via a loss of Kir6.2/SUR2B current in neuronal cells. Login to comment 218 ABCC9 p.Val734Ile The observation that the responses of Kir6.2/SUR2A to MgATP and MgNDP are unaltered by the mutation excludes the possibility that V734I causes AMI by altering the protective role of cardiac KATP channels in ischemia/reperfusion injury. Login to comment 219 ABCC9 p.Val734Ile The exact molecular mechanism of the SUR2B-V734I mutation in vivo will be more thoroughly understood when animal models carrying this mutation become available, thus enabling cardiovascular changes in conditions of high and low metabolism to be assessed. Login to comment 221 ABCC9 p.Val734Ile Molecular mechanism The V734I mutation caused a decrease in sensitivity to MgATP inhibition. Login to comment 222 ABCC9 p.Val734Ile This effect appeared to be Mg2+ -dependent as Kir6.2/SUR2B and Kir6.2/SUR2B-V734I channels presented identical sensitivities to ATP in the absence of Mg2+ . Login to comment 224 ABCC9 p.Val734Ile Thus, it can be inferred that the V734I mutation did not alter the affinity/efficacy of ATP binding at the inhibitory site of Kir6.2. Login to comment 225 ABCC9 p.Val734Ile Furthermore, the reduction in MgATP sensitivity of Kir6.2/SUR2B-V734I channels was not secondary to a change in the intrinsic Po of the channel [61]. Login to comment 227 ABCC9 p.Val734Ile However, the observation that the non-hydrolysable ATP analogue AMP-PNP was equally potent on Kir6.2/SUR2B and Kir6.2/SUR2B-V734I channels argues against this hypothesis. Login to comment 229 ABCC9 p.Val734Ile ABCC9 p.Val734Ile A plausible scenario is that the V734I mutation results in channels with an enhanced rate of MgATP hydrolysis which would account for the greater residual current in the presence of MgATP for Kir6.2/SUR2B-V734I channels. Login to comment 233 ABCC9 p.Val734Ile Structural considerations A recent study indicated that the region to which V734I maps is a site potentially critical for the interaction between NBD1 of adjacent SUR2 subunits [62]. Login to comment 235 ABCC9 p.Val734Ile The mutation acts only on SUR2B- and not SUR2A-containing KATP channels: our findings indicate a possible functional interaction between the region where V734I lies and the 42 amino-acid C-terminal region that differs between SUR2A and SUR2B. Login to comment 238 ABCC9 p.Val734Ile We have shown that low doses (30 bc;M) of nicorandil promoted activation of Kir6.2/SUR2B-V734I channels. Login to comment 239 ABCC9 p.Val734Ile It can be extrapolated that the circulating plasma levels of nicorandil, which are likely to be in the nanomolar range [64], may be sufficient to activate Kir6.2/SUR2B-V734I channels to near normal levels. Login to comment 240 ABCC9 p.Val734Ile These observations point out nicorandil as a possible therapeutic drug in the treatment of AMI associated with the V734I mutation. Login to comment