A cDNA encoding a potassium channel of the two-pore domain family (K2P, KCNK) of leak channels was cloned from the marine sponge Amphimedon queenslandica. Phylogenetic analysis indicated that AquK2P cannot be placed into any of the established functional groups of mammalian K2P channels. We used the Xenopus oocyte expression system, a two-electrode voltage clamp and inside-out patch clamp electrophysiology to determine the physiological properties of AquK2P. In whole cells, non-inactivating, voltage-independent, outwardly rectifying K+ currents were generated by external application of micromolar concentrations of arachidonic acid (AA; EC50 ∼30 μmol l–1), when applied in an alkaline solution (≥pH 8.0). Prior activation of channels facilitated the pH-regulated, AA-dependent activation of AquK2P but external pH changes alone did not activate the channels. Unlike certain mammalian fatty-acid-activated K2P channels, the sponge K2P channel was not activated by temperature and was insensitive to osmotically induced membrane distortion. In inside-out patch recordings, alkalinization of the internal pH (pKa 8.18) activated the AquK2P channels independently of AA and also facilitated activation by internally applied AA. The gating of the sponge K2P channel suggests that voltage-independent outward rectification and sensitivity to pH and AA are ancient and fundamental properties of animal K2P channels. In addition, the membrane potential of some poriferan cells may be dynamically regulated by pH and AA.
↵* These authors contributed equally to this work
Supplementary material available online at http://jeb.biologists.org/cgi/content/full/215/14/2435/DC1
This research was supported by the National Institutes of Health [GM096142 to L.M.B.; R01 HL59949 to D.E.L.], a Mednick Memorial Grant Fellowship [to L.M.B.], an undergraduate fellowship from the Howard Hughes Medical Institute [to G.W.], the University of Richmond Dickinson Award in Biology [to R.H.], and an Natural Sciences and Engineering Research Council of Canada Discovery grant [to S.P.L.]. Deposited in PMC for release after 6 months.
LIST OF ABBREVIATIONS
- Amphimedon queenslandica K2P channel
- arachidonic acid
- bovine serum albumin
- tandem pore domain potassium channel
- internal/intracellular pH
- external/extracellular pH
- two-electrode voltage clamp
- © 2012.