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Differential mechanisms of Cantú syndrome-associated gain of function mutations in the ABCC9 (SUR2) subunit of the KATP channel.

Cooper PE, Sala-Rabanal M, Lee SJ, Nichols CG - J. Gen. Physiol. (2015)

Bottom Line: For P429L and A475V mutants, sensitivity to ATP inhibition was comparable to WT channels, but activation by MgADP was significantly greater.C1039Y-dependent channels were significantly less sensitive to inhibition by ATP or by glibenclamide, but MgADP activation was comparable to WT.The results indicate that these three CS mutations all lead to overactive K(ATP) channels, but at least two mechanisms underlie the observed gain of function: decreased ATP inhibition and enhanced MgADP activation.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Cell Biology and Physiology, and Center for the Investigation of Membrane Excitability Diseases, Washington University School of Medicine, Saint Louis, MO 63110 Department of Cell Biology and Physiology, and Center for the Investigation of Membrane Excitability Diseases, Washington University School of Medicine, Saint Louis, MO 63110.

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P429L and A475V show enhanced MgADP activation. (A) Representative KATP current traces after excision and in the presence of nucleotides as indicated. (B) MgADP activation, as a ratio between the steady-state current in the presence of MgADP and the maximum current measured in the patch upon excision, in the absence of nucleotides. Individual patch data represented by symbols (n = 11–24); error bars are the means ± SEM, respectively 26.8 ± 0.03% (WT), 56.9 ± 0.06% (P429L), 59.7 ± 0.1% (A475V), and 38.7 ± 0.04% (C1039Y). *, P < 0.05 as compared with WT (unpaired Student’s t test). Inset shows the mean current measured in patches in the presence of 0.1 mM of Mg-free ATP (from Fig. 5 B) subtracted from the mean value of steady-state current measured in patches in the presence of MgADP (from Fig. 7 A). Error bars represent the propagated error from both experiments. (C) Half-time of activation by MgADP.
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fig7: P429L and A475V show enhanced MgADP activation. (A) Representative KATP current traces after excision and in the presence of nucleotides as indicated. (B) MgADP activation, as a ratio between the steady-state current in the presence of MgADP and the maximum current measured in the patch upon excision, in the absence of nucleotides. Individual patch data represented by symbols (n = 11–24); error bars are the means ± SEM, respectively 26.8 ± 0.03% (WT), 56.9 ± 0.06% (P429L), 59.7 ± 0.1% (A475V), and 38.7 ± 0.04% (C1039Y). *, P < 0.05 as compared with WT (unpaired Student’s t test). Inset shows the mean current measured in patches in the presence of 0.1 mM of Mg-free ATP (from Fig. 5 B) subtracted from the mean value of steady-state current measured in patches in the presence of MgADP (from Fig. 7 A). Error bars represent the propagated error from both experiments. (C) Half-time of activation by MgADP.

Mentions: We investigated the current response to intracellular MgADP in the presence of 0.1 mM ATP (Fig. 7 A). MgADP-dependent activation was estimated as the ratio between the steady-state activated current in MgADP+ATP and the maximal current in zero ATP, immediately after excision (Fig. 7 B). The relative current in MgADP was markedly higher than WT for P429L and A475V channels. Using this analysis, the current was also statistically higher for C1039Y channels. However, this analysis ignores the intrinsically lower sensitivity of C1039Y channels to ATP inhibition (Fig. 5 C). An alternative estimation for the stimulatory action of Mg nucleotides is to calculate the ratio of current in MgADP+ATP to that in Mg-free ATP alone (Fig. 7 B, inset). This analysis implies no difference in the MgADP activation of C1039Y and WT channels. In addition, although the MgADP activation for P429L and A475V channels is more rapid than WT, activation of the C1039Y channel is even slower than WT (Fig. 7 C).


Differential mechanisms of Cantú syndrome-associated gain of function mutations in the ABCC9 (SUR2) subunit of the KATP channel.

Cooper PE, Sala-Rabanal M, Lee SJ, Nichols CG - J. Gen. Physiol. (2015)

P429L and A475V show enhanced MgADP activation. (A) Representative KATP current traces after excision and in the presence of nucleotides as indicated. (B) MgADP activation, as a ratio between the steady-state current in the presence of MgADP and the maximum current measured in the patch upon excision, in the absence of nucleotides. Individual patch data represented by symbols (n = 11–24); error bars are the means ± SEM, respectively 26.8 ± 0.03% (WT), 56.9 ± 0.06% (P429L), 59.7 ± 0.1% (A475V), and 38.7 ± 0.04% (C1039Y). *, P < 0.05 as compared with WT (unpaired Student’s t test). Inset shows the mean current measured in patches in the presence of 0.1 mM of Mg-free ATP (from Fig. 5 B) subtracted from the mean value of steady-state current measured in patches in the presence of MgADP (from Fig. 7 A). Error bars represent the propagated error from both experiments. (C) Half-time of activation by MgADP.
© Copyright Policy - openaccess
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4664827&req=5

fig7: P429L and A475V show enhanced MgADP activation. (A) Representative KATP current traces after excision and in the presence of nucleotides as indicated. (B) MgADP activation, as a ratio between the steady-state current in the presence of MgADP and the maximum current measured in the patch upon excision, in the absence of nucleotides. Individual patch data represented by symbols (n = 11–24); error bars are the means ± SEM, respectively 26.8 ± 0.03% (WT), 56.9 ± 0.06% (P429L), 59.7 ± 0.1% (A475V), and 38.7 ± 0.04% (C1039Y). *, P < 0.05 as compared with WT (unpaired Student’s t test). Inset shows the mean current measured in patches in the presence of 0.1 mM of Mg-free ATP (from Fig. 5 B) subtracted from the mean value of steady-state current measured in patches in the presence of MgADP (from Fig. 7 A). Error bars represent the propagated error from both experiments. (C) Half-time of activation by MgADP.
Mentions: We investigated the current response to intracellular MgADP in the presence of 0.1 mM ATP (Fig. 7 A). MgADP-dependent activation was estimated as the ratio between the steady-state activated current in MgADP+ATP and the maximal current in zero ATP, immediately after excision (Fig. 7 B). The relative current in MgADP was markedly higher than WT for P429L and A475V channels. Using this analysis, the current was also statistically higher for C1039Y channels. However, this analysis ignores the intrinsically lower sensitivity of C1039Y channels to ATP inhibition (Fig. 5 C). An alternative estimation for the stimulatory action of Mg nucleotides is to calculate the ratio of current in MgADP+ATP to that in Mg-free ATP alone (Fig. 7 B, inset). This analysis implies no difference in the MgADP activation of C1039Y and WT channels. In addition, although the MgADP activation for P429L and A475V channels is more rapid than WT, activation of the C1039Y channel is even slower than WT (Fig. 7 C).

Bottom Line: For P429L and A475V mutants, sensitivity to ATP inhibition was comparable to WT channels, but activation by MgADP was significantly greater.C1039Y-dependent channels were significantly less sensitive to inhibition by ATP or by glibenclamide, but MgADP activation was comparable to WT.The results indicate that these three CS mutations all lead to overactive K(ATP) channels, but at least two mechanisms underlie the observed gain of function: decreased ATP inhibition and enhanced MgADP activation.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Cell Biology and Physiology, and Center for the Investigation of Membrane Excitability Diseases, Washington University School of Medicine, Saint Louis, MO 63110 Department of Cell Biology and Physiology, and Center for the Investigation of Membrane Excitability Diseases, Washington University School of Medicine, Saint Louis, MO 63110.

No MeSH data available.


Related in: MedlinePlus