Functional expression of the voltage-gated neuronal mammalian potassium channel rat ether à go-go1 in yeast.
It has been shown previously that heterologous expression of inwardly rectifying potassium channels (K+-channels) from plants and mammals in K+-transport defective yeast mutants can restore the ability of growth in media with low [K+]. In this study, the functional expression of an outward rectifying mammalian K+-channel in yeast is presented for the first time. The outward-rectifying mammalian neuronal K+-channel rat ether à go-go channel 1 (rEAG1, Kv 10.1) was expressed in yeast (Saccharomyces cerevisiae) strains lacking the endogenous K+-uptake systems and/or alkali-metal-cation efflux systems. It was found that a truncated channel version, lacking almost the complete intracellular N-terminus (rEAG1 Delta 190) but not the full-length rEAG1, partially complemented the growth defect of K+-uptake mutant cells (trk1,2 Delta tok1 Delta) in media containing low K+ concentrations. The expression of rEAG1 Delta 190 in a strain lacking the cation efflux systems (nha1 Delta ena1-4 Delta) increased the sensitivity to high monovalent cation concentrations. Both phenotypes were observed, when rEAG1 Delta 190 was expressed in a trk1,2 Delta and nha1, ena1-4 Delta mutant strain. In the presence of K+-channel blockers (Cs+, Ba2+ and quinidine), the growth advantage of rEAG1 Delta 190 expressing trk1,2 tok1 Delta cells disappeared, indicating its dependence on functional rEAG1 channels. The results demonstrate that S. cerevisiae is a suitable expression system even for voltage-gated outward-rectifying mammalian K+-channels.
Schwarzer
Sarah
S
Molekulare Bioenergetik, IZMB, Universität Bonn, Bonn, Germany.
Kolacna
Lucie
L
Lichtenberg-Fraté
Hella
H
Sychrova
Hana
H
Ludwig
Jost
J
D000818:Animals; D017609:Barium Compounds / Q000494:pharmacology; D002712:Chlorides / Q000494:pharmacology; D051638:Ether-A-Go-Go Potassium Channels / Q000235:genetics / Q000502:physiology*; D006863:Hydrogen-Ion Concentration; D011188:Potassium / Q000378:metabolism; D026902:Potassium Channel Blockers / Q000494:pharmacology; D051381:Rats; D011994:Recombinant Proteins / Q000378:metabolism; D012413:Rubidium / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*
D017609:Barium Compounds; D002712:Chlorides; D051638:Ether-A-Go-Go Potassium Channels; D026902:Potassium Channel Blockers; D011994:Recombinant Proteins; C024986:barium chloride; D012413:Rubidium; D011188:Potassium