RESUMEN
Voltage-gated potassium (K(v)) channels work in concert with other ion channels to determine the frequency and duration of action potentials in excitable cells. Little is known about K(v)3 channels from invertebrates, but those that have been characterized generally display slow kinetics. Here, we report the cloning and characterization of jShaw1, the first K(v)3 isolated from a cnidarian, the jellyfish Polyorchis penicillatus, in comparison with mouse K(v)3.1 and K(v)3.2. Using a two-electrode voltage clamp on Xenopus laevis oocytes expressing the channels, we compared steady-state and kinetic properties of macroscopic currents. jShaw1 is fast activating, and opens at potentials approximately 40 mV more hyperpolarized than the mouse K(v)3 channels. There is an inverse relationship between the number of positive charges on the voltage sensor and the half-activation voltage of the channel, contrary to what would be expected with the simplest model of voltage sensitivity. jShaw1 has kinetic characteristics that are substantially different from the mammalian K(v)3 channels, including a much lower sensitivity of early activation rates to incremental voltage changes, and a much faster voltage-dependent transition in the last stages of opening. jShaw1 opening kinetics were affected little by pre-depolarization voltage, in contrast to both mouse channels. Similar to the mouse channels, jShaw1 was half-blocked by 0.7 mmol l(-1) tetraethyl ammonium and 5 mmol l(-1) 4-aminopyridine. Comparison of sequence and functional properties of jShaw1 with the mouse and other reported K(v)3 channels helps to illuminate the general relationship between amino acid sequence and electrophysiological activity in this channel family.
Asunto(s)
Hidrozoos/metabolismo , Activación del Canal Iónico/fisiología , Canales de Potasio/metabolismo , 4-Aminopiridina/farmacología , Secuencia de Aminoácidos , Animales , Hidrozoos/efectos de los fármacos , Activación del Canal Iónico/efectos de los fármacos , Cinética , Ratones , Modelos Moleculares , Datos de Secuencia Molecular , Oocitos/efectos de los fármacos , Oocitos/metabolismo , Filogenia , Canales de Potasio/química , Análisis de Secuencia de ADN , Tetraetilamonio/farmacología , Xenopus laevisRESUMEN
Divergence of the Shaker superfamily of voltage-gated (Kv) ion channels early in metazoan evolution created numerous electrical phenotypes that were presumably selected to produce a wide range of excitability characteristics in neurons, myocytes, and other cells. A comparative approach that emphasizes this early radiation provides a comprehensive sampling of sequence space that is necessary to develop generally applicable models of the structure-function relationship in the Kv potassium channel family. We have cloned and characterized two Shaw-type potassium channels from a flatworm (Notoplana atomata) that is arguably a representative of early diverging bilaterians. When expressed in Xenopus oocytes, one of these cloned channels, N.at-Kv3.1, exhibits a noninactivating, outward current with slow opening kinetics that are dependent on both the holding potential and the activating potential. A second Shaw-type channel, N.at-Kv3.2, has very different properties, showing weak inward rectification. These results demonstrate that broad phylogenetic sampling of proteins of a single family will reveal unexpected properties that lead to new interpretations of structure-function relationships.