The Journal of Membrane Biology

, Volume 245, Issue 11, pp 667–674 | Cite as

Expression, Purification and Functional Reconstitution of Slack Sodium-Activated Potassium Channels

  • Yangyang Yan
  • Youshan Yang
  • Shumin Bian
  • Fred J. Sigworth
Article
First Online:
Received:
Accepted:

Abstract

The slack (slo2.2) gene codes for a potassium-channel α-subunit of the 6TM voltage-gated channel family. Expression of slack results in Na+-activated potassium channel activity in various cell types. We describe the purification and reconstitution of Slack protein and show that the Slack α-subunit alone is sufficient for potassium channel activity activated by sodium ions as assayed in planar bilayer membranes and in membrane vesicles.

Keywords

Artificial planar membrane Ion channel expression and reconstitution Patch clamp Potassium ion channel 
This is a preview of subscription content, log in to check access.

Notes

Acknowledgments

We are grateful to Drs. David Chester and Yufeng Zhou for their advice and patient instruction in cryo-EM and reconstitution techniques. This work was supported by NIH grant NS21501 to F.S.

References

  1. Bhattacharjee A, Kaczmarek LK (2005) For K+ channels, Na+ is the new Ca2+. Trends Neurosci 28:422–428PubMedCrossRefGoogle Scholar
  2. Bhattacharjee A, Gan L, Kaczmarek LK (2002) Localization of the Slack potassium channel in the rat central nervous system. J Comp Neurol 454:241–254PubMedCrossRefGoogle Scholar
  3. Bhattacharjee A, Joiner WJ, Wu M, Yang Y, Sigworth FJ, Kaczmarek LK (2003) Slick (Slo2.1), a rapidly-gating sodium-activated potassium channel inhibited by ATP. J Neurosci 23:11681–11691PubMedGoogle Scholar
  4. Bhattacharjee A, von Hehn CAA, Mei X, Kaczmarek LK (2005) Localization of the Na+-activated K+ channel Slick in the rat central nervous system. J Comp Neurol 484:80–92PubMedCrossRefGoogle Scholar
  5. Brown MR, Kronengold J, Gazula VR, Spilianakis CG, Flavell RA, von Hehn CA, Bhattacharjee A, Kaczmarek LK (2008) Amino-termini isoforms of the Slack K+ channel, regulated by alternative promoters, differentially modulate rhythmic firing and adaptation. J Physiol 586:5161–5179PubMedCrossRefGoogle Scholar
  6. Brown MR, Kronengold J, Gazula V-R, Chen Y, Strumbos JG, Sigworth FJ, Navaratnam D, Kaczmarek LK (2010) Fragile X mental retardation protein controls gating of the sodium-activated potassium channel Slack. Nat Neurosci 13:819–821PubMedCrossRefGoogle Scholar
  7. Budelli G, Hage TA, Wei A, Rojas P, Jong Y-JI, Oandapos-Malley K, Salkoff L (2009) Na+-activated K+ channels express a large delayed outward current in neurons during normal physiology. Nat Neurosci 12:745–750PubMedCrossRefGoogle Scholar
  8. Chen H, Kronengold J, Yan Y, Gazula V-R, Brown MR, Ma L, Ferreira G, Yang Y, Bhattacharjee A, Sigworth FJ, Salkoff L, Kaczmarek LK (2009) The N-terminal domain of Slack determines the formation and trafficking of Slick/Slack heteromeric sodium-activated potassium channels. J Neurosci 29:5654–5665PubMedCrossRefGoogle Scholar
  9. Chester DW, Klemic JF, Stern E, Sigworth FJ, Klemic KG (2007) Holey carbon micro-arrays for transmission electron microscopy: a microcontact printing approach. Ultramicroscopy 107:685–691PubMedCrossRefGoogle Scholar
  10. Cong Y, Ludtke SJ (2010) Single particle analysis at high resolution. Methods Enzymol 482:211–235PubMedCrossRefGoogle Scholar
  11. Fleming MR, Kaczmarek LK (2009) Use of optical biosensors to detect modulation of Slack potassium channels by G protein-coupled receptors. J Recept Signal Transduct Res 29:173–181PubMedCrossRefGoogle Scholar
  12. Joiner WJ, Tang MD, Wang LY, Dworetzky SI, Boissard CG, Gan L, Gribkoff VK, Kaczmarek LK (1998) Formation of intermediate-conductance calcium-activated potassium channels by interaction of Slack and Slo subunits. Nat Neurosci 1:462–469PubMedCrossRefGoogle Scholar
  13. Kameyama M, Kakei M, Sato R, Shibasaki T, Matsuda H, Irisawa H (1984) Intracellular Na+ activates a K+ channel in mammalian cardiac cells. Nature 309:354–356PubMedCrossRefGoogle Scholar
  14. Kim YC, Sim JH, Kang TM, Suzuki H, Kim SR, Kwon SC, Xu WX, Lee SJ, Kim KW (2007) Sodium-activated potassium current in guinea pig gastric myocytes. J Korean Med Sci 22:57–62PubMedCrossRefGoogle Scholar
  15. Paulais M, Lachheb S, Teulon J (2006) A Na+- and Cl−-activated K+ channel in the thick ascending limb of mouse kidney. J Gen Physiol 127:205–215PubMedCrossRefGoogle Scholar
  16. Ruffin VA, Gu XQ, Zhou D, Douglas RM, Sun X, Trouth CO, Haddad GG (2008) The sodium-activated potassium channel Slack is modulated by hypercapnia and acidosis. Neuroscience 151:410–418PubMedCrossRefGoogle Scholar
  17. Santi CM, Ferreira G, Yang B, Gazula V-R, Butler A, Wei A, Kaczmarek LK, Salkoff L (2006) Opposite regulation of Slick and Slack K+ channels by neuromodulators. J Neurosci 26:5059–5068PubMedCrossRefGoogle Scholar
  18. Uchino S, Wada H, Honda S, Hirasawa T, Yanai S, Nakamura Y, Ondo Y, Kohsaka S (2003) Slo2 sodium-activated K+ channels bind to the PDZ domain of PSD-95. Biochem Biophys Res Commun 310:1140–1147PubMedCrossRefGoogle Scholar
  19. Wang L, Sigworth FJ (2009) Structure of the BK potassium channel in a lipid membrane from electron cryomicroscopy. Nature 461(7261):292–295PubMedCrossRefGoogle Scholar
  20. Yang B, Desai R, Kaczmarek LK (2007) Slack and Slick K(Na) channels regulate the accuracy of timing of auditory neurons. J Neurosci 27:2617–2627PubMedCrossRefGoogle Scholar
  21. Yuan A, Santi CM, Wei A, Wang ZW, Pollak K, Nonet M, Kaczmarek L, Crowder CM, Salkoff L (2003) The sodium-activated potassium channel is encoded by a member of the Slo gene family. Neuron 37:765–773PubMedCrossRefGoogle Scholar
  22. Yuan P, Leonetti MD, Pico AR, Hsiung Y, MacKinnon R (2010) Structure of the human BK channel Ca2+-activation apparatus at 3.0 Å resolution. Science 329:182–186PubMedCrossRefGoogle Scholar
  23. Zhang Y, Paterson WG (2007) Functional evidence for Na+-activated K+ channels in circular smooth muscle of the opossum lower esophageal sphincter. Am J Physiol Gastrointest Liver Physiol 292:G1600–G1606PubMedCrossRefGoogle Scholar
  24. Zhang Z, Rosenhouse-Dantsker A, Tang Q-Y, Noskov S, Logothetis DE (2010) The RCK2 domain uses a coordination site present in Kir channels to confer sodium sensitivity to Slo2.2 channels. J Neurosci 30:7554–7562PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2012

Authors and Affiliations

  • Yangyang Yan
    • 1
  • Youshan Yang
    • 1
  • Shumin Bian
    • 1
  • Fred J. Sigworth
    • 1
  1. 1.Department of Cellular and Molecular PhysiologyYale UniversityNew HavenUSA

Personalised recommendations