Polynucleotides encoding acetylcholine-gated chloride channel subunits of caenorhabditis elegans

Abstract

The invention relates to novel polynucleotides which encode novel polypeptides that are acetylcholine-gated chloride channel subunits and immunogenic or acetylcholine-binding fragments thereof. The novel polypeptides may be used for identifying compounds that modulate the acetylcholine-gated chloride channels, e.g. for use as pesticides and antiparasitic agents. Methods for identifying compounds that modulate the acetylcholine-gated chloride channels are provided.

1 - 48 . (canceled) 
     
     
         49 . A method of identifying a modulator of an acetylcholine-gated chloride channel, comprising:
 (a) providing a test compound;   (b) providing a polypeptide comprising an acetylcholine-gated chloride channel (ACC) subunit polypeptide or a biologically active fragment thereof; and   (c) detecting whether the test compound modulates the activity of said polypeptide or biologically active fragment thereof;   
       wherein said polypeptide has at least 60% amino acid identity to the mature extracellular ligand-binding domain of an ACC channel subunit selected from the group consisting of ACC-1 (SEQ ID NO: 2), ACC-2 (SEQ ID NO: 4), ACC-3 (SEQ ID NO: 8), and ACC-4 (SEQ ID NO: 10). 
     
     
         50 . The method according to  claim 49 , wherein said polypeptide, when aligned with ACC-1 (SEQ ID NO:2), comprises the following amino acid residues:
 Y at the position corresponding to Y81 in ACC-1;   N or T or a conservative substitution thereof at the position corresponding to N83 in ACC-1;   Y at the position corresponding to Y100 in ACC-1;   I at the position corresponding to 1121 in ACC-1;   M at the position corresponding to M 139 in ACC-1;   F at the position corresponding to F141 in ACC-1;   W or M or a conservative substitution thereof at the position corresponding to W147 in ACC-1;   N at the position corresponding to N149 in ACC-1;   F at the position corresponding to F156 in ACC-1;   M at the position corresponding to M158 in ACC-1;   F at the position corresponding to F179 in ACC-1;   F at the position corresponding to F198 in ACC-1.   Y at the position corresponding to Y221 in ACC-1; and   W at the position corresponding to W226 in ACC-1.   
     
     
         51 . The method according to  claim 49  wherein said polypeptide, when aligned with ACC-2 (SEQ ID NO: 4), comprises the following amino acid residues:
 Y at the position corresponding to Y81 in ACC-2;   Y at the position corresponding to Y 100 in ACC-2;   N or T or a conservative substitution thereof at the position corresponding to T102 in ACC-2;   F at the position corresponding to F137 in ACC-2;   M at the position corresponding to M139 in ACC-2;   I at the position corresponding to I140 in ACC-2;   M at the position corresponding to M158 in ACC-2;   F at the position corresponding to F160 in ACC-2;   W or M or a conservative substitution thereof at the position corresponding to W166 in ACC-2;   N at the position corresponding to N168 in ACC-2;   F at the position corresponding to F179 in ACC-2   F at the position corresponding to F198 in ACC-2;   Y at the position corresponding to Y24I in ACC-2; and   W at the position corresponding to W246 in ACC-2.   
     
     
         52 . The method according to  claim 49 , wherein said polypeptide comprises an anion-selective PART motif or variant thereof in its M2 transmembrane domain. 
     
     
         53 . The method according to  claim 52  wherein the variant of the PART motif consists of conserved proline and alanine residues and conservative amino acid substitutions at the arginine and/or threonine positions of the PART motif. 
     
     
         54 . The method according to  claim 49  wherein said polypeptide or biologically active fragment thereof is fused to a second polypeptide. 
     
     
         55 . A method of identifying a modulator of an acetylcholine-gated chloride channel, comprising the steps of:
 (a) providing a test compound;   (b) providing a polypeptide comprising an acetylcholine-gated chloride channel (ACC) subunit polypeptide or a biologically active fragment thereof;   (c) providing a binding partner for the polypeptide of step b); and   (d) detecting whether the test compound modulates binding of the binding partner to the polypeptide;   
       wherein said polypeptide has at least 60% amino acid identity to the mature extracellular ligand-binding domain of an ACC channel subunit selected from the group consisting of ACC-1 (SEQ ID NO: 2), ACC-2 (SEQ ID NO: 4), ACC-3 (SEQ ID NO: 8), and ACC-4 (SEQ ID NO: 10). 
     
     
         56 . The method according to  claim 55 , wherein said polypeptide, when aligned with ACC-1 (SEQ ID NO:2), comprises the following amino acid residues:
 Y at the position corresponding to Y81 in ACC-1;   N or T or a conservative substitution thereof at the position corresponding to N83 in ACC-1;   Y at the position corresponding to Y100 in ACC-1;   I at the position corresponding to I121 in ACC-1;   M at the position corresponding to M139 in ACC-1;   F at the position corresponding to F141 in ACC-1;   W or M or a conservative substitution thereof at the position corresponding to W147 in ACC-1;   N at the position corresponding to N149 in ACC-1;   F at the position corresponding to F156 in ACC-1;   M at the position corresponding to M158 in ACC-1;   F at the position corresponding to F179 in ACC-1;   F at the position corresponding to F198 in ACC-1.   Y at the position corresponding to Y221 in ACC-1; and   W at the position corresponding to W226 in ACC-1.   
     
     
         57 . The method according to  claim 55  wherein said polypeptide, when aligned with ACC-2 (SEQ ID NO: 4), comprises the following amino acid residues:
 Y at the position corresponding to Y81 in ACC-2;   Y at the position corresponding to Y100 in ACC-2;   N or T or a conservative substitution thereof at the position corresponding to T102 in ACC-2;   F at the position corresponding to F137 in ACC-2;   M at the position corresponding to M139 in ACC-2;   I at the position corresponding to I140 in ACC-2;   M at the position corresponding to M158 in ACC-2;   F at the position corresponding to F160 in ACC-2;   W or M or a conservative substitution thereof at the position corresponding to W166 in ACC-2;   N at the position corresponding to N168 in ACC-2;   F at the position corresponding to F179 in ACC-2   F at the position corresponding to F198 in ACC-2;   Y at the position corresponding to Y241 in ACC-2; and   W at the position corresponding to W246 in ACC-2.   
     
     
         58 . The method according to  claim 55 , wherein said polypeptide comprises an anion-selective PART motif or variant thereof in its M2 transmembrane domain. 
     
     
         59 . The method according to  claim 58  wherein the variant of the PART motif consists of conserved proline and alanine residues and conservative amino acid substitutions at the arginine and/or threonine positions of the PART motif. 
     
     
         60 . The method according to  claim 55 , wherein said polypeptide or biologically active fragment thereof is fused to a second polypeptide.