Potassium channel mutants of the yeast Saccharomyces cerevisiae and their use for screening eukaryotic potassium channels

Abstract

The invention relates to processes for identifying inhibitors and activators of eukaryotic potassium channels, in which a mutated S. cerevisiae cell is used whose endogenous potassium channels TRK1, TRK2 and TOK1 are not expressed functionally, but which expresses heterologously a eukaryotic potassium channel to be studied. Other subject matters of the invention are mutated S. cerevisiae cells which do not express TRK1, TRK2 and TOK1, and the preparation and use of these mutated S. cerevisiae cells.

1 - 24 . (canceled) 
     
     
         25 . A process for identifying inhibitors of a human potassium channel,
 a) providing a mutated  S. cerevisiae  cell which does not express the three endogenous potassium channels TRK1, TRK2 and TOK1;   b) treating said mutated cell with a human potassium channel wherein said human potassium channel is expressed heterologously in this mutated  S. cerevisiae  cell;   c) incubating the mutated  S. cerevisiae  cell expressing the human potassium channel together with a substance to be tested; and   d) determining the effect of the substance to be tested on the human potassium channel, wherein a decrease in the transport of potassium across the human potassium channel indicates that the substance is an inhibitor of the human potassium channel.   
     
     
         26 . The process as claimed in  claim 25 , wherein the genes TRK1, TRK2 and TOK1 are switched off in the mutated  S. cerevisiae  cell (Δtrk1, Δtrk2, Δtok1). 
     
     
         27 . The process as claimed in  claim 25 , wherein the human potassium channel is a HERG1, Kv1.5 or IRK1. 
     
     
         28 . The process as claimed in  claim 25 , wherein the human potassium channel is mutated. 
     
     
         29 . The process as claimed in  claim 25 , wherein the human potassium channel is present in a yeast expression plasmid. 
     
     
         30 . The process as claimed in  claim 25 , wherein the mutated  S. cerevisiae  cell expresses constitutively a growth reporter. 
     
     
         31 . The process as claimed in  claim 25 , wherein the substance to be tested, which has an effect on the human potassium channel, inhibits the growth of the mutated  S. cerevisiae  cell. 
     
     
         32 . The process as claimed in  claim 31 , wherein the effect of the substance to be tested on the human potassium channel is determined by measuring the cell count of the mutated  S. cerevisiae  cells. 
     
     
         33 . The process as claimed in  claim 30 , wherein the cell count is determined via fluorescence or luminescence of the constitutively expressed growth reporter. 
     
     
         34 . A mutated  S. cerevisiae  cell in which TRK1, TRK2 and TOK1 are not expressed, and which expresses heterologously a human potassium channel. 
     
     
         35 . The mutated  S. cerevisiae  cell as claimed in  claim 34 , wherein the human potassium channel is HERG1, Kv1.5 or IRK1. 
     
     
         36 . The mutated  S. cerevisiae  cell as claimed in  claim 34 , wherein the human potassium channel is mutated. 
     
     
         37 . A process of identifying activators of a human potassium channel,
 a) providing a mutated  S. cerevisiae  cell which does not express the three endogenous potassium channels TRK1, TRK2 and TOK1;   b) reacting said mutated cell with a human potassium channel, wherein said human potassium channel is expressed heterologously in this mutated  S. cerevisiae  cell;   c) incubating the mutated  S. cerevisiae  cell expressing the human potassium channel together with a substance to be tested; and   d) determining the effect of the substance to be tested on the human potassium channel, wherein an increase in the transport of potassium across the human potassium channel indicates that the substance is an activator of the human potassium channel.   
     
     
         38 . A process of identifying activators of a human potassium channel,
 a) providing a mutated  S. cerevisiae  cell which does not express the three endogenous potassium channels TRK1, TRK2 and TOK1;   b) reacting said mutated cell with a human potassium channel wherein said human potassium channel is expressed heterologously in this mutated  S. cerevisiae  cell;   c) incubating the mutated  S. cerevisiae  cell expressing the human potassium channel together with a substance to be tested in the presence of an inhibitor of the human potassium channel; and   d) determining the effect of the substance to be tested on the human potassium channel, wherein an increase in the transport of potassium across the human potassium channel indicates that the substance is an activator of the human potassium channel.   
     
     
         39 . A test kit comprising the mutated  S. cerevisiae  cell as claimed in  claim 34 .