Abstract
Aim:
ATP-sensitive potassium (KATP) channels formed by a combination of SUR/Kir6.x subunits play a crucial role in protection against hypoxic or ischemic injuries resulting from cell metabolic disorders. In this study we investigated the effects of Na-azide, a metabolic inhibitor, on KATP channels expressed in Xenopus oocytes, and explored the structure basis for their sensitivity to cell metabolic disorders.
Methods:
Six subtypes of KATP channels (wild SUR1/Kir6.2, SUR2B/Kir6.2, SUR1/Kir6.1, SUR2B/Kir6.1, SUR2A/Kir6.2 and SUR2A/Kir6.1), as well as eleven subtypes of KATP channels with mutant subunits were expressed in Xenopus oocytes. KATP currents were recorded using a two-electrode voltage clamp recording technique. The drugs were applied through bath.
Results:
Except SUR2A/Kir6.1, five subtypes of KATP channels were activated by Na-azide (3 mmol/L) with an order of the responses: SUR1/Kir6.2>SUR2B/Kir6.2>SUR1/Kir6.1>SUR2B/Kir6.1>SUR2A/Kir6.2, and the opening rate (t1/2) was SUR1/Kir6.x>SUR2B/Kir6.x>SUR2A/Kir6.2. Furthermore, Kir6.2, rather than Kir6.1, had intrinsic sensitivity to Na-azide, and the residues involved in ATP-binding (R50 and K185) or pH-sensing (H175) were associated with the sensitivity of the Kir6.2 subunit to Na-azide. Moreover, the residues (K707 and K1348) within the Walker A (WA) motifs of two nucleotide-binding domains (NBDs) were essential for SUR2B/Kir6.x (especially SUR2B/Kir6.1) channel activation by Na-azide, suggesting a key role for Mg-adenine nucleotide binding and/or hydrolysis in the SUR2B subunit.
Conclusion:
Among the six subtypes of KATP channels, SUR1/Kir6.2 is the most sensitive, whereas SUR2A/Kir6.1 is insensitive, to cell metabolic disorders. The Kir6.2 subunit, rather than the Kir6.1 subunit, has intrinsic sensitivity to cell metabolic disorders. The residues (K707 and K1348) within the WA motifs of SUR2B are important for the sensitivity of SUR2B/Kir6.x channels to cell metabolic disorders.
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Acknowledgements
We thank Prof LY JAN (Howard Hughes Medical Institute) for kindly providing cDNAs for the KATP channels used for expression in Xenopus oocytes.
This work was supported by grants from the State Key Research Project of China (No AWS11J003) and the National Basic Research “973” Program (No 2012CB518200 and JCKY2013000B001).
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Supplementary Figure S1 is available in the Acta Pharmacologica Sinica's website.
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Supplementary Information, Figure S1
Representative time courses of currents recorded at -80 mV from water-injected Xenopus oocytes (B1), oocytes expressing SUR1/Kir6.1 (B2), SUR2A/Kir6.1 (B3), SUR2B/Kir6.1 (B4), SUR1/Kir6.2 (B5), SUR2A/Kir6.2 (B6) and SUR2B/Kir6.2 (B7). (DOC 24 kb)
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Li, Cg., Cui, Wy. & Wang, H. Sensitivity of KATP channels to cellular metabolic disorders and the underlying structural basis. Acta Pharmacol Sin 37, 134–142 (2016). https://doi.org/10.1038/aps.2015.134
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DOI: https://doi.org/10.1038/aps.2015.134
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