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Zinc pyrithione-mediated activation of voltage-gated KCNQ potassium channels rescues epileptogenic mutants

Nature Chemical Biology volume 3, pages 287–296 (2007)Cite this article

Abstract

KCNQ potassium channels are activated by changes in transmembrane voltage and play an important role in controlling electrical excitability. Human mutations of KCNQ2 and KCNQ3 potassium channel genes result in reduction or loss of channel activity and cause benign familial neonatal convulsions (BFNCs). Thus, small molecules capable of augmenting KCNQ currents are essential both for understanding the mechanism of channel activity and for developing therapeutics. We performed a high-throughput screen in search for agonistic compounds potentiating KCNQ potassium channels. Here we report identification of a new opener, zinc pyrithione (1), which activates both recombinant and native KCNQ M currents. Interactions with the channel protein cause an increase of single-channel open probability that could fully account for the overall conductance increase. Separate point mutations have been identified that either shift the concentration dependence or affect potentiation efficacy, thereby providing evidence for residues influencing ligand binding and downstream events. Furthermore, zinc pyrithione is capable of rescuing the mutant channels causal to BFNCs.

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Figure 1: ZnPy potentiated potassium currents induced by KCNQ homomultimers.
Figure 2: Combinatorial effects on KCNQ2 current by potassium channel blockers and ZnPy.
Figure 3: Biophysical effects of ZnPy on KCNQ channels.
Figure 4: ZnPy increases the Po of KCNQ2 channel.
Figure 5: Molecular determinants for ZnPy sensitivity.
Figure 6: ZnPy potentiation of native M current.
Figure 7: ZnPy effects on BFNC mutant channels expressed in CHO cells.

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References

  1. Ashcroft, F.M. From molecule to malady. Nature 440, 440–447 (2006).

    Article  CAS  Google Scholar 

  2. Jan, L.Y. & Jan, Y.N. Cloned potassium channels from eukaryotes and prokaryotes. Annu. Rev. Neurosci. 20, 91–123 (1997).

    Article  CAS  Google Scholar 

  3. Tombola, F., Pathak, M.M. & Isacoff, E.Y. How far will you go to sense voltage? Neuron 48, 719–725 (2005).

    Article  CAS  Google Scholar 

  4. Brown, D.A. & Adams, P.R. Muscarinic suppression of a novel voltage-sensitive K+ current in a vertebrate neurone. Nature 283, 673–676 (1980).

    Article  CAS  Google Scholar 

  5. Marrion, N.V. Control of M-current. Annu. Rev. Physiol. 59, 483–504 (1997).

    Article  CAS  Google Scholar 

  6. Cooper, E.C., Harrington, E., Jan, Y.N. & Jan, L.Y. M channel KCNQ2 subunits are localized to key sites for control of neuronal network oscillations and synchronization in mouse brain. J. Neurosci. 21, 9529–9540 (2001).

    Article  CAS  Google Scholar 

  7. Schroeder, B.C., Kubisch, C., Stein, V. & Jentsch, T.J. Moderate loss of function of cyclic-AMP-modulated KCNQ2/KCNQ3 K+ channels causes epilepsy. Nature 396, 687–690 (1998).

    Article  CAS  Google Scholar 

  8. Hille, B. Ion Channels of Excitable Membranes 220–238 (Sinauer, Sunderland, Massachusetts, USA, 2001).

    Google Scholar 

  9. Jentsch, T.J. Neuronal KCNQ potassium channels: physiology and role in disease. Nat. Rev. Neurosci. 1, 21–30 (2000).

    Article  CAS  Google Scholar 

  10. Rogawski, M.A. KCNQ2/KCNQ3 K+ channels and the molecular pathogenesis of epilepsy: implications for therapy. Trends Neurosci. 23, 393–398 (2000).

    Article  CAS  Google Scholar 

  11. Charlier, C. et al. A pore mutation in a novel KQT-like potassium channel gene in an idiopathic epilepsy family. Nat. Genet. 18, 53–55 (1998).

    Article  CAS  Google Scholar 

  12. Gutman, G.A. et al. International Union of Pharmacology. XLI. Compendium of voltage-gated ion channels: potassium channels. Pharmacol. Rev. 55, 583–586 (2003).

    Article  CAS  Google Scholar 

  13. Kubisch, C. et al. KCNQ4, a novel potassium channel expressed in sensory outer hair cells, is mutated in dominant deafness. Cell 96, 437–446 (1999).

    Article  CAS  Google Scholar 

  14. Schroeder, B.C., Hechenberger, M., Weinreich, F., Kubisch, C. & Jentsch, T.J. KCNQ5, a novel potassium channel broadly expressed in brain, mediates M-type currents. J. Biol. Chem. 275, 24089–24095 (2000).

    Article  CAS  Google Scholar 

  15. Singh, N.A. et al. A novel potassium channel gene, KCNQ2, is mutated in an inherited epilepsy of newborns. Nat. Genet. 18, 25–29 (1998).

    Article  CAS  Google Scholar 

  16. Wang, Q. et al. Positional cloning of a novel potassium channel gene: KVLQT1 mutations cause cardiac arrhythmias. Nat. Genet. 12, 17–23 (1996).

    Article  Google Scholar 

  17. Shapiro, M.S. et al. Reconstitution of muscarinic modulation of the KCNQ2/KCNQ3 K(+) channels that underlie the neuronal M current. J. Neurosci. 20, 1710–1721 (2000).

    Article  CAS  Google Scholar 

  18. Wang, H.S. et al. KCNQ2 and KCNQ3 potassium channel subunits: molecular correlates of the M-channel. Science 282, 1890–1893 (1998).

    Article  CAS  Google Scholar 

  19. Cooper, E.C. & Jan, L.Y. Ion channel genes and human neurological disease: recent progress, prospects, and challenges. Proc. Natl. Acad. Sci. USA 96, 4759–4766 (1999).

    Article  CAS  Google Scholar 

  20. Armstrong, C.M. Time course of TEA(+)-induced anomalous rectification in squid giant axons. J. Gen. Physiol. 50, 491–503 (1966).

    Article  CAS  Google Scholar 

  21. Armstrong, C.M. Inactivation of the potassium conductance and related phenomena caused by quaternary ammonium ion injection in squid axons. J. Gen. Physiol. 54, 553–575 (1969).

    Article  CAS  Google Scholar 

  22. Armstrong, C.M. Interaction of tetraethylammonium ion derivatives with the potassium channels of giant axons. J. Gen. Physiol. 58, 413–437 (1971).

    Article  CAS  Google Scholar 

  23. Holmgren, M., Smith, P.L. & Yellen, G. Trapping of organic blockers by closing of voltage-dependent K+ channels: evidence for a trap door mechanism of activation gating. J. Gen. Physiol. 109, 527–535 (1997).

    Article  CAS  Google Scholar 

  24. Xu, J., Chen, Y. & Li, M. High-throughput technologies for studying potassium channels - progresses and challenges. Targets 3, 32–38 (2004).

    CAS  Google Scholar 

  25. Sanguinetti, M.C. & Tristani-Firouzi, M. hERG potassium channels and cardiac arrhythmia. Nature 440, 463–469 (2006).

    Article  CAS  Google Scholar 

  26. Rostock, A. et al. D-23129: a new anticonvulsant with a broad spectrum activity in animal models of epileptic seizures. Epilepsy Res. 23, 211–223 (1996).

    Article  CAS  Google Scholar 

  27. Tober, C., Rostock, A., Rundfeldt, C. & Bartsch, R. D-23129: a potent anticonvulsant in the amygdala kindling model of complex partial seizures. Eur. J. Pharmacol. 303, 163–169 (1996).

    Article  CAS  Google Scholar 

  28. Rundfeldt, C. The new anticonvulsant retigabine (D-23129) acts as an opener of K+ channels in neuronal cells. Eur. J. Pharmacol. 336, 243–249 (1997).

    Article  CAS  Google Scholar 

  29. Tatulian, L., Delmas, P., Abogadie, F.C. & Brown, D.A. Activation of expressed KCNQ potassium currents and native neuronal M-type potassium currents by the anti-convulsant drug retigabine. J. Neurosci. 21, 5535–5545 (2001).

    Article  CAS  Google Scholar 

  30. Ramu, Y., Xu, Y. & Lu, Z. Enzymatic activation of voltage-gated potassium channels. Nature 442, 696–699 (2006).

    Article  CAS  Google Scholar 

  31. Sun, H., Shikano, S., Xiong, Q. & Li, M. Function recovery after chemobleaching (FRAC): evidence for activity silent membrane receptors on cell surface. Proc. Natl. Acad. Sci. USA 101, 16964–16969 (2004).

    Article  CAS  Google Scholar 

  32. Sun, H., Liu, X., Xiong, Q., Shikano, S. & Li, M. Chronic inhibition of cardiac kir2.1 and HERG potassium channels by celastrol with dual effects on both ion conductivity and protein trafficking. J. Biol. Chem. 281, 5877–5884 (2006).

    Article  CAS  Google Scholar 

  33. Terstappen, G.C. Functional analysis of native and recombinant ion channels using a high-capacity nonradioactive rubidium efflux assay. Anal. Biochem. 272, 149–155 (1999).

    Article  CAS  Google Scholar 

  34. Wickenden, A.D., Yu, W., Zou, A., Jegla, T. & Wagoner, P.K. Retigabine, a novel anti-convulsant, enhances activation of KCNQ2/Q3 potassium channels. Mol. Pharmacol. 58, 591–600 (2000).

    Article  CAS  Google Scholar 

  35. Bond, A.D.J. Synthesis and characterization of a novel zinc pyrithione hydrate. Mol. Cryst. Liq. Cryst. Sci. Technol. 356, 305–313 (2001).

    Article  CAS  Google Scholar 

  36. Wang, D.W. Synthesis of N-oxide-2-mercaptopyridine zinc salt. Riyong Huaxue Gongye 33, 340–342 (2003).

    CAS  Google Scholar 

  37. Kimura, E., Takasawa, R., Tanuma, S. & Aoki, S. Monitoring apoptosis with fluorescent Zn2+-indicators. Sci. STKE 2004, PL7 (2004).

    PubMed  Google Scholar 

  38. Barnett, B.L., Kretschmar, H.C. & Hartman, F.A. Structural characterization of bis(N-oxopyridine-2-thionato)zinc (II). Inorg. Chem. 16, 1834–1838 (1977).

    Article  CAS  Google Scholar 

  39. Main, M.J. et al. Modulation of KCNQ2/3 potassium channels by the novel anticonvulsant retigabine. Mol. Pharmacol. 58, 253–262 (2000).

    Article  CAS  Google Scholar 

  40. Schenzer, A. et al. Molecular determinants of KCNQ (Kv7) K+ channel sensitivity to the anticonvulsant retigabine. J. Neurosci. 25, 5051–5060 (2005).

    Article  CAS  Google Scholar 

  41. Wuttke, T.V., Seebohm, G., Bail, S., Maljevic, S. & Lerche, H. The new anticonvulsant retigabine favors voltage-dependent opening of the Kv7.2 (KCNQ2) channel by binding to its activation gate. Mol. Pharmacol. 67, 1009–1017 (2005).

    Article  CAS  Google Scholar 

  42. Long, S.B., Campbell, E.B. & Mackinnon, R. Voltage sensor of Kv1.2: structural basis of electromechanical coupling. Science 309, 903–908 (2005).

    Article  CAS  Google Scholar 

  43. Seebohm, G. et al. Differential roles of S6 domain hinges in the gating of KCNQ potassium channels. Biophys. J. 90, 2235–2244 (2006).

    Article  CAS  Google Scholar 

  44. Biervert, C. et al. A potassium channel mutation in neonatal human epilepsy. Science 279, 403–406 (1998).

    Article  CAS  Google Scholar 

  45. Dedek, K. et al. Myokymia and neonatal epilepsy caused by a mutation in the voltage sensor of the KCNQ2 K+ channel. Proc. Natl. Acad. Sci. USA 98, 12272–12277 (2001).

    Article  CAS  Google Scholar 

  46. Horrigan, F.T. & Aldrich, R.W. Coupling between voltage sensor activation, Ca2+ binding and channel opening in large conductance (BK) potassium channels. J. Gen. Physiol. 120, 267–305 (2002).

    Article  CAS  Google Scholar 

  47. Bandell, M. et al. High-throughput random mutagenesis screen reveals TRPM8 residues specifically required for activation by menthol. Nat. Neurosci. 9, 493–500 (2006).

    Article  CAS  Google Scholar 

  48. Li, Y., Gamper, N. & Shapiro, M.S. Single-channel analysis of KCNQ K+ channels reveals the mechanism of augmentation by a cysteine-modifying reagent. J. Neurosci. 24, 5079–5090 (2004).

    Article  CAS  Google Scholar 

  49. Huang, C.C., Lesburg, C.A., Kiefer, L.L., Fierke, C.A. & Christianson, D.W. Reversal of the hydrogen bond to zinc ligand histidine-119 dramatically diminishes catalysis and enhances metal equilibration kinetics in carbonic anhydrase II. Biochemistry 35, 3439–3446 (1996).

    Article  CAS  Google Scholar 

  50. Laine, M. et al. Atomic proximity between S4 segment and pore domain in Shaker potassium channels. Neuron 39, 467–481 (2003).

    Article  CAS  Google Scholar 

  51. Shealy, R.T., Murphy, A.D., Ramarathnam, R., Jakobsson, E. & Subramaniam, S. Sequence-function analysis of the K+-selective family of ion channels using a comprehensive alignment and the KcsA channel structure. Biophys. J. 84, 2929–2942 (2003).

    Article  CAS  Google Scholar 

  52. Marks, R., Pearse, A.D. & Walker, A.P. The effects of a shampoo containing zinc pyrithione on the control of dandruff. Br. J. Dermatol. 112, 415–422 (1985).

    Article  CAS  Google Scholar 

  53. Fliss, H. Zinc ionophores as therapeutic agents. US patent 6,495,538 (2002).

  54. Mathie, A., Sutton, G.L., Clarke, C.E. & Veale, E.L. Zinc and copper: pharmacological probes and endogenous modulators of neuronal excitability. Pharmacol. Ther. 111, 567–583 (2006).

    Article  CAS  Google Scholar 

  55. Guex, N. & Peitsch, M.C. SWISS-MODEL and the Swiss-PdbViewer: an environment for comparative protein modeling. Electrophoresis 18, 2714–2723 (1997).

    Article  CAS  Google Scholar 

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Acknowledgements

We thank T. Jentsch (Zentrum für Molekulare Neurobiologie, Hamburg), D. Makinnon (State University of New York, Stony Brook), M. Sanguinetti (University of Utah), M. Shapiro (University of Texas Health Science Center, San Antonio) and V. Vardanyan (Universität Hamburg) for gifts of cDNAs. We thank M. Fratine, A. Ince, S. Long, M. Spieker and K. Xie for technical assistance. We also thank B. Coblitz for help with structural modeling and our colleagues and members of the Li laboratory for valuable discussions and comments on the manuscript. The N-type calcium channel stable line was a kind gift from D. Lipscombe and D. Yue. This work is supported by a grant from the US National Institutes of Health to M.L. (GM70959).

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  1. Department of Neuroscience, High Throughput Biology Center and Department of Physiology, School of Medicine, Johns Hopkins University, 733 North Broadway, Baltimore, 21205, Maryland, USA

    Qiaojie Xiong, Haiyan Sun & Min Li

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  1. Qiaojie Xiong
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Contributions

Q.X. performed all electrophysiological recordings and analyses. H.S. performed high-throughput compound screens and constructed site-directed mutants. Q.X. and H.S. constructed vectors and established cell lines expressing KCNQ channels. M.L. helped conceive experiments.

Note: Supplementary information and chemical compound information is available on the Nature Chemical Biology website.

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Correspondence to Min Li.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Fig. 1

Zinc pyrithione effects on other potassium channels and N-type calcium channel. (PDF 15 kb)

Supplementary Fig. 2

Complexed form of ZnPy is required for potentiation. (PDF 30 kb)

Supplementary Fig. 3

Other divalent ion effects on KCNQ2 current. (PDF 26 kb)

Supplementary Fig. 4

Synthesis of zinc pyrithione and sodium pyrithione. (PDF 15 kb)

Supplementary Fig. 5

Stoichiometric preference of Zn ion and pyrithione. (PDF 17 kb)

Supplementary Fig. 6

ZnPy effects on KCNQ2 and KCNQ2/3 deactivation. (PDF 102 kb)

Supplementary Fig. 7

Differential modulatory sites on KCNQ channel and retigabine. (PDF 31 kb)

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Xiong, Q., Sun, H. & Li, M. Zinc pyrithione-mediated activation of voltage-gated KCNQ potassium channels rescues epileptogenic mutants. Nat Chem Biol 3, 287–296 (2007). https://doi.org/10.1038/nchembio874

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