Identification of a K+ channel from potato leaves by functional expression in Xenopus oocytes.
Poly(A)+ mRNA was isolated from leaves of potato plants (Solanum tuberosum L. cv. Desiree) according to standard protocols. This poly(A)+ mRNA was injected via glass microcapillaries into oocytes that were surgically removed from the African clawed toad Xenopus laevis. As a control, oocytes were either injected with H2O or remained untreated. Three days after injection the oocytes were analyzed by two electrode voltage clamping. Current voltage analysis revealed that a K+ channel from potato was functionally expressed in injected oocytes. The identity of this K+ channel was confirmed by its substrate specificity and a shift in the reversal potential. In particular, when the outside K+ concentration was increased the reversal potential of poly(A)+ injected oocytes shifted to more positive values. Furthermore, K+ outward currents declined when the outside K+ concentration was raised from 0.1 to 100 mM. Inward currents increased with an elevation of the K+ concentration. Several pharmaceuticals were tested for their potential to block this K+ channel. As a result, the channel was completely blocked by BaCl2. A three state reaction kinetic model was used to simulate the currents through the K+ transport protein as function of the extracellular K+ concentration. In particular, the simulation revealed current voltage relations that exactly matched the measured ones. Saturation of current voltage curves emerged from the simulation as a consequence of high extracellular potassium concentration.
Max Planck Institut für molekulare Pflanzenphysiologie, Golm, Germany.
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Brandt
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Fisahn
D000818:Animals; D005260:Female; D017930:Genes, Reporter; D005966:Glucuronidase / Q000096:biosynthesis; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D009865:Oocytes; D018408:Patch-Clamp Techniques; D018515:Plant Leaves; D011188:Potassium / Q000494:pharmacology*; D015221:Potassium Channels / Q000096:biosynthesis / Q000235:genetics / Q000502:physiology*; D012333:RNA, Messenger / Q000302:isolation & purification / Q000378:metabolism; D018749:RNA, Plant / Q000302:isolation & purification / Q000378:metabolism; D011994:Recombinant Proteins / Q000096:biosynthesis; D011198:Solanum tuberosum / Q000502:physiology*; D014982:Xenopus laevis
D015221:Potassium Channels; D012333:RNA, Messenger; D018749:RNA, Plant; D011994:Recombinant Proteins; D005966:Glucuronidase; D011188:Potassium