Transgenic gpcr expressing animals

Abstract

The invention provides isolated nucleic acids molecules, designated muscarinic acetylcholine receptor 6 (“mACHR-6”) nucleic acid molecules, which encode polypeptides involved in the modulation of acetylcholine responses in acetylcholine responsive cells. The invention also provides antisense nucleic acid molecules, expression vectors containing mACHR-6 nucleic acid molecules, host cells into which the expression vectors have been introduced, and non-human transgenic animals in which an mACHR-6 gene has been introduced or disrupted. The invention still further provides isolated mACHR-6 polypeptides, fusion polypeptides, antigenic peptides, and anti-mACHR-6 antibodies. Diagnostic, screening, and therapeutic methods utilizing compositions of the invention are also provided.

What is claimed is:  
     
         1 . An isolated nucleic acid molecule selected from the group consisting of: 
 a) a nucleic acid molecule which encodes a polypeptide comprising the amino acid sequence of SEQ ID NO:2;    b) a nucleic acid molecule which encodes a polypeptide comprising the amino acid sequence of SEQ ID NO:5;    c) a nucleic acid molecule which encodes a polypeptide comprising the amino acid sequence of SEQ ID NO:32;    d) a nucleic acid molecule which encodes a fragment of a polypeptide comprising the amino acid sequence of SEQ ID NO:2, wherein the fragment comprises at least 15 contiguous amino acids of SEQ ID NO:2;    e) a nucleic acid molecule which encodes a fragment of a polypeptide comprising the amino acid sequence of SEQ ID NO:5, wherein the fragment comprises at least 15 contiguous amino acids of SEQ ID NO:5;    f) a nucleic acid molecule which encodes a fragment of a polypeptide comprising the amino acid sequence of SEQ ID NO:32, wherein the fragment comprises at least 15 contiguous amino acids of SEQ ID NO:32;    g) a nucleic acid molecule which encodes a naturally occurring allelic variant of a polypeptide comprising the amino acid sequence of SEQ ID NO:2, wherein the nucleic acid molecule hybridizes to a nucleic acid molecule comprising SEQ ID NO:1 under stringent conditions; and    h) a nucleic acid molecule which encodes a naturally occurring allelic variant of a polypeptide comprising the amino acid sequence of SEQ ID NO:5, wherein the nucleic acid molecule hybridizes to a nucleic acid molecule comprising SEQ ID NO:4 under stringent conditions; and    i) a nucleic acid molecule which encodes a naturally occurring allelic variant of a polypeptide comprising the amino acid sequence of SEQ ID NO:32, wherein the nucleic acid molecule hybridizes to a nucleic acid molecule comprising SEQ ID NO:31 under stringent conditions.    
     
     
         2 . The nucleic acid molecule of  claim 1  further comprising vector nucleic acid sequences.  
     
     
         3 . The nucleic acid molecule of  claim 1  further comprising nucleic acid sequences encoding a heterologous polypeptide.  
     
     
         4 . A host cell which contains the nucleic acid molecule of  claim 1 .  
     
     
         5 . The host cell of  claim 4  which is a mammalian host cell.  
     
     
         6 . A non-human mammalian host cell containing the nucleic acid molecule of  claim 1 .  
     
     
         7 . The isolated nucleic acid molecule of  claim 1 , which is selected from the group consisting of: 
 a) the coding region of SEQ ID NO:1;    b) the coding region of SEQ ID NO:4;    c) the coding region of SEQ ID NO:31;    d) a nucleic acid molecule which encodes a polypeptide comprising the amino acid sequence of SEQ ID NO:7;    e) a nucleic acid molecule which encodes a polypeptide comprising the amino acid sequence of SEQ ID NO:8;    f) a nucleic acid molecule which encodes a polypeptide comprising the amino acid sequence of SEQ ID NO:9;    g) a nucleic acid molecule which encodes a polypeptide comprising the amino acid sequence of SEQ ID NO:10;    h) a nucleic acid molecule which encodes a polypeptide comprising the amino acid sequence of SEQ ID NO:11;    i) a nucleic acid molecule which encodes a polypeptide comprising the amino acid sequence of SEQ ID NO:12; and    j) a nucleic acid molecule which encodes a polypeptide comprising the amino acid sequence of SEQ ID NO:13.    
     
     
         8 . An isolated polypeptide selected from the group consisting of: 
 a) a polypeptide comprising the amino acid sequence of SEQ ID NO:2;    b) a polypeptide comprising the amino acid sequence of SEQ ID NO:5;    c) a polypeptide comprising the amino acid sequence of SEQ ID NO:32;    d) a fragment of a polypeptide comprising the amino acid sequence of SEQ ID NO:2, wherein the fragment comprises at least 15 contiguous amino acids of SEQ ID NO:2;    e) a fragment of a polypeptide comprising the amino acid sequence of SEQ ID NO:5, wherein the fragment comprises at least 15 contiguous amino acids of SEQ ID NO:5;    f) a fragment of a polypeptide comprising the amino acid sequence of SEQ ID NO:32, wherein the fragment comprises at least 15 contiguous amino acids of SEQ ID NO:32;    g) a naturally occurring allelic variant of a polypeptide comprising the amino acid sequence of SEQ ID NO:2, wherein the polypeptide is encoded by a nucleic acid molecule which hybridizes to a nucleic acid molecule comprising SEQ ID NO:1 under stringent conditions;    h) a naturally occurring allelic variant of a polypeptide comprising the amino acid sequence of SEQ ID NO:5, wherein the polypeptide is encoded by a nucleic acid molecule which hybridizes to a nucleic acid molecule comprising SEQ ID NO:4 under stringent conditions; and    i) a naturally occurring allelic variant of a polypeptide comprising the amino acid sequence of SEQ ID NO:32, wherein the polypeptide is encoded by a nucleic acid molecule which hybridizes to a nucleic acid molecule comprising SEQ ID NO:31 under stringent conditions.    
     
     
         9 . The polypeptide of  claim 8 , further comprising heterologous amino acid sequences.  
     
     
         10 . An antibody which selectively binds to a polypeptide of  claim 8 .  
     
     
         11 . A method for producing a polypeptide selected from the group consisting of: 
 a) a polypeptide comprising the amino acid sequence of SEQ ID NO:2;    b) a polypeptide comprising the amino acid sequence of SEQ ID NO:5;    c) a polypeptide comprising the amino acid sequence of SEQ ID NO:32;    d) a fragment of a polypeptide comprising the amino acid sequence of SEQ ID NO:2, wherein the fragment comprises at least 15 contiguous amino acids of SEQ ID NO:2;    e) a fragment of a polypeptide comprising the amino acid sequence of SEQ ID NO:5, wherein the fragment comprises at least 15 contiguous amino acids of SEQ ID NO:5;    f) a fragment of a polypeptide comprising the amino acid sequence of SEQ ID NO:32, wherein the fragment comprises at least 15 contiguous amino acids of SEQ ID NO:32;    g) a naturally occurring allelic variant of a polypeptide comprising the amino acid sequence of SEQ ID NO:2, wherein the polypeptide is encoded by a nucleic acid molecule which hybridizes to a nucleic acid molecule comprising SEQ ID NO:1 under stringent conditions;    h) a naturally occurring allelic variant of a polypeptide comprising the amino acid sequence of SEQ ID NO:5, wherein the polypeptide is encoded by a nucleic acid molecule which hybridizes to a nucleic acid molecule comprising SEQ ID NO:4 under stringent conditions; and    i) a naturally occurring allelic variant of a polypeptide comprising the amino acid sequence of SEQ ID NO:32, wherein the polypeptide is encoded by a nucleic acid molecule which hybridizes to a nucleic acid molecule comprising SEQ ID NO:31 under stringent conditions;    the method comprising the step of culturing the host cell of  claim 4  under conditions in which the nucleic acid molecule is expressed.    
     
     
         12 . A method for detecting the presence of a polypeptide selected from the group consisting of: 
 a) a polypeptide comprising the amino acid sequence of SEQ ID NO:2;    b) a polypeptide comprising the amino acid sequence of SEQ ID NO:5;    c) a polypeptide comprising the amino acid sequence of SEQ ID NO:32;    d) a fragment of a polypeptide comprising the amino acid sequence of SEQ ID NO:2, wherein the fragment comprises at least 15 contiguous amino acids of SEQ ID NO:2;    e) a fragment of a polypeptide comprising the amino acid sequence of SEQ ID NO:5, wherein the fragment comprises at least 15 contiguous amino acids of SEQ ID NO:5;    f) a fragment of a polypeptide comprising the amino acid sequence of SEQ ID NO:32, wherein the fragment comprises at least 15 contiguous amino acids of SEQ ID NO:32;    g) a naturally occurring allelic variant of a polypeptide comprising the amino acid sequence of SEQ ID NO:2, wherein the polypeptide is encoded by a nucleic acid molecule which hybridizes to a nucleic acid molecule comprising SEQ ID NO:1 under stringent conditions;    h) a naturally occurring allelic variant of a polypeptide comprising the amino acid sequence of SEQ ID NO:5, wherein the polypeptide is encoded by a nucleic acid molecule which hybridizes to a nucleic acid molecule comprising SEQ ID NO:4 under stringent conditions; and    i) a naturally occurring allelic variant of a polypeptide comprising the amino acid sequence of SEQ ID NO:32, wherein the polypeptide is encoded by a nucleic acid molecule which hybridizes to a nucleic acid molecule comprising SEQ ID NO:31 under stringent conditions;    in a sample, the method comprising the steps of: 
 i) contacting the sample with a compound which selectively binds to the polypeptide; and  
 ii) determining whether the compound binds to the polypeptide in the sample.  
   
     
     
         13 . The method of  claim 12 , wherein the compound which binds to the polypeptide is an antibody.  
     
     
         14 . A kit comprising reagents used for the method of  claim 12 , wherein the reagents comprise a compound which selectively binds to a polypeptide selected from the group consisting of: 
 a) a polypeptide comprising the amino acid sequence of SEQ ID NO:2;    b) a polypeptide comprising the amino acid sequence of SEQ ID NO:5;    c) a polypeptide comprising the amino acid sequence of SEQ ID NO:32;    d) a fragment of a polypeptide comprising the amino acid sequence of SEQ ID NO:2, wherein the fragment comprises at least 15 contiguous amino acids of SEQ ID NO:2;    e) a fragment of a polypeptide comprising the amino acid sequence of SEQ ID NO:5, wherein the fragment comprises at least 15 contiguous amino acids of SEQ ID NO:5;    f) a fragment of a polypeptide comprising the amino acid sequence of SEQ ID NO:32, wherein the fragment comprises at least 15 contiguous amino acids of SEQ ID NO:32;    g) a naturally occurring allelic variant of a polypeptide comprising the amino acid sequence of SEQ ID NO:2, wherein the polypeptide is encoded by a nucleic acid molecule which hybridizes to a nucleic acid molecule comprising SEQ ID NO:1 under stringent conditions;    h) a naturally occurring allelic variant of a polypeptide comprising the amino acid sequence of SEQ ID NO:5, wherein the polypeptide is encoded by a nucleic acid molecule which hybridizes to a nucleic acid molecule comprising SEQ ID NO:4 under stringent conditions; and    i) a naturally occurring allelic variant of a polypeptide comprising the amino acid sequence of SEQ ID NO:32, wherein the polypeptide is encoded by a nucleic acid molecule which hybridizes to a nucleic acid molecule comprising SEQ ID NO:31 under stringent conditions.    
     
     
         15 . A method for detecting the presence of a nucleic acid molecule selected from the group consisting of: 
 a) a polypeptide comprising the amino acid sequence of SEQ ID NO:2;    b) a polypeptide comprising the amino acid sequence of SEQ ID NO:5;    c) a polypeptide comprising the amino acid sequence of SEQ ID NO:32;    d) a fragment of a polypeptide comprising the amino acid sequence of SEQ ID NO:2, wherein the fragment comprises at least 15 contiguous amino acids of SEQ ID NO:2;    e) a fragment of a polypeptide comprising the amino acid sequence of SEQ ID NO:5, wherein the fragment comprises at least 15 contiguous amino acids of SEQ ID NO:5;    f) a fragment of a polypeptide comprising the amino-acid sequence of SEQ ID NO:32, wherein the fragment comprises at least 15 contiguous amino acids of SEQ ID NO:32;    g) a naturally occurring allelic variant of a polypeptide comprising the amino acid sequence of SEQ ID NO:2, wherein the polypeptide is encoded by a nucleic acid molecule which hybridizes to a nucleic acid molecule comprising SEQ ID NO:1 under stringent conditions;    h) a naturally occurring allelic variant of a polypeptide comprising the amino acid sequence of SEQ ID NO:5, wherein the polypeptide is encoded by a nucleic acid molecule which hybridizes to a nucleic acid molecule comprising SEQ ID NO:4 under stringent conditions; and    i) a naturally occurring allelic variant of a polypeptide comprising the amino acid sequence of SEQ ID NO:32, wherein the polypeptide is encoded by a nucleic acid molecule which hybridizes to a nucleic acid molecule comprising SEQ ID NO:31 under stringent conditions;    in a sample, the method comprising the steps of: 
 i) contacting the sample with a nucleic acid probe or primer which selectively hybridizes to the nucleic acid molecule; and  
 ii) determining whether the nucleic acid probe or primer binds to a nucleic acid molecule in the sample.  
   
     
     
         16 . The method of  claim 15 , wherein the sample comprises mRNA molecules and is contacted with a nucleic acid probe.  
     
     
         17 . A kit comprising reagents used for the method of  claim 15 , wherein the reagents comprise a compound which selectively hybridizes to a nucleic acid molecule selected from the group consisting of: 
 a) a polypeptide comprising the amino acid sequence of SEQ ID NO:2;    b) a polypeptide comprising the amino acid sequence of SEQ ID NO:5;    c) a polypeptide comprising the amino acid sequence of SEQ ID NO:32;    d) a fragment of a polypeptide comprising the amino acid sequence of SEQ ID NO:2, wherein the fragment comprises at least 15 contiguous amino acids of SEQ ID NO:2;    e) a fragment of a polypeptide comprising the amino acid sequence of SEQ ID NO:5, wherein the fragment comprises at least 15 contiguous amino acids of SEQ ID NO:5;    f) a fragment of a polypeptide comprising the amino acid sequence of SEQ ID NO:32, wherein the fragment comprises at least 15 contiguous amino acids of SEQ ID NO:32;    g) a naturally occurring allelic variant of a polypeptide comprising the amino acid sequence of SEQ ID NO:2, wherein the polypeptide is encoded by a nucleic acid molecule which hybridizes to a nucleic acid molecule comprising SEQ ID NO:1 under stringent conditions;    h) a naturally occurring allelic variant of a polypeptide comprising the amino acid sequence of SEQ ID NO:5, wherein the polypeptide is encoded by a nucleic acid molecule which hybridizes to a nucleic acid molecule comprising SEQ ID NO:4 under stringent conditions; and    i) a naturally occurring allelic variant of a polypeptide comprising the amino acid sequence of SEQ ID NO:32, wherein the polypeptide is encoded by a nucleic acid molecule which hybridizes to a nucleic acid molecule comprising SEQ ID NO:31 under stringent conditions.    
     
     
         18 . A method for identifying a compound which binds to a polypeptide selected from the group consisting of: 
 a) a polypeptide comprising the amino acid sequence of SEQ ID NO:2;    b) a polypeptide comprising the amino acid sequence of SEQ ID NO:5;    c) a polypeptide comprising the amino acid sequence of SEQ ID NO:32;    d) a fragment of a polypeptide comprising the amino acid sequence of SEQ ID NO:2, wherein the fragment comprises at least 15 contiguous amino acids of SEQ ID NO:2;    e) a fragment of a polypeptide comprising the amino acid sequence of SEQ ID NO:5, wherein the fragment comprises at least 15 contiguous amino acids of SEQ ID NO:5;    f) a fragment of a polypeptide comprising the amino acid sequence of SEQ ID NO:32, wherein the fragment comprises at least 15 contiguous amino acids of SEQ ID NO:32;    g) a naturally occurring allelic variant of a polypeptide comprising the amino acid sequence of SEQ ID NO:2, wherein the polypeptide is encoded by a nucleic acid molecule which hybridizes to a nucleic acid molecule comprising SEQ ID NO:1 under stringent conditions;    h) a naturally occurring allelic variant of a polypeptide comprising the amino acid sequence of SEQ ID NO:5, wherein the polypeptide is encoded by a nucleic acid molecule which hybridizes to a nucleic acid molecule comprising SEQ ID NO:4 under stringent conditions; and    i) a naturally occurring allelic variant of a polypeptide comprising the amino acid sequence of SEQ ID NO:32, wherein the polypeptide is encoded by a nucleic acid molecule which hybridizes to a nucleic acid molecule comprising SEQ ID NO:31 under stringent conditions,    the method comprising the steps of: 
 i) contacting the polypeptide, or a cell expressing the polypeptide with a test compound; and  
 ii) determining whether the polypeptide binds to the test compound.  
   
     
     
         19 . The method of  claim 18 , wherein the binding of the test compound to the polypeptide is detected by a method selected from the group consisting of: 
 a) detection of binding by direct detecting of test compound/polypeptide binding;    b) detection of binding using a competition binding assay;    c) detection of binding using an assay for mACHR-6 activity.    
     
     
         20 . A method for modulating the activity of a polypeptide selected from the group consisting of: 
 a) a polypeptide comprising the amino acid sequence of SEQ ID NO:2;    b) a polypeptide comprising the amino acid sequence of SEQ ID NO:5;    c) a polypeptide comprising the amino acid sequence of SEQ ID NO:32;    d) a fragment of a polypeptide comprising the amino acid sequence of SEQ ID NO:2, wherein the fragment comprises at least 15 contiguous amino acids of SEQ ID NO:2;    e) a fragment of a polypeptide comprising the amino acid sequence of SEQ ID NO:5, wherein the fragment comprises at least 15 contiguous amino acids of SEQ ID NO:5;    f) a fragment of a polypeptide comprising the amino acid sequence of SEQ ID NO:32, wherein the fragment comprises at least 15 contiguous amino acids of SEQ ID NO:32;    g) a naturally occurring allelic variant of a polypeptide comprising the amino acid sequence of SEQ ID NO:2, wherein the polypeptide is encoded by a nucleic acid molecule which hybridizes to a nucleic acid molecule comprising SEQ ID NO:1 under stringent conditions;    h) a naturally occurring allelic variant of a polypeptide comprising the amino acid sequence of SEQ ID NO:5, wherein the polypeptide is encoded by a nucleic acid molecule which hybridizes to a nucleic acid molecule comprising SEQ ID NO:4 under stringent conditions; and    i) a naturally occurring allelic variant of a polypeptide comprising the amino acid sequence of SEQ ID NO:32, wherein the polypeptide is encoded by a nucleic acid molecule which hybridizes to a nucleic acid molecule comprising SEQ ID NO:31 under stringent conditions,    the method comprising the steps of: 
 i) contacting a cell expressing the polypeptide with a compound which binds to the polypeptide in a sufficient concentration to modulate the activity of the polypeptide.  
   
     
     
         21 . The method of  claim 20 , wherein the activity is a phosphatidylinositol activity.  
     
     
         22 . The method of  claim 20 , wherein the method results in an increase in phosphatidylinositol metabolism.  
     
     
         23 . The method of  claim 20 , wherein the method results in a decrease in phosphatidylinositol metabolism.