Pacemaker ion channel proteins and uses thereof

Abstract

The present invention provides an isolated nucleic acid molecule encoding a brain cyclic nucleotide gated ion channel (BCNG) protein. An isolated BCNG protein is also provided as is a composition comprising a BCNG encoding nucleic acid or protein or a portion thereof. The present invention also provides a method of identifying an ion channel subunit related protein encoding nucleic acid molecule in a sample. The invention further provides a method for evaluating the ability of a compound to modulate an ion channel associated neurological, cardia, or renal condition. The invention also provides a method for evaluating the ability of a compound to interact with a BCNG-related ion channel subunit protein. Additionally, the present invention provides a method for identifying a compound capable of modulating the activity of a BCNG-related protein. The present invention additionally provides a method of treating a cardiac, renal or neurological condition. Finally, the present invention provides an antibody which specifically reacts with BCNG protein.

1 . An isolated nucleic acid molecule having sequence encoding a BCNG ion channel protein or a portion thereof.  
     
     
         2 . The nucleic acid molecule of  claim 1 , wherein the BCNG ion channel protein is mBCNG-1, mBCNG-2, mBCNG-3, mBCNG-4, hBCNG-1 or hBCNG-2.  
     
     
         3 . The nucleic acid molecule of  claim 1 , wherein the nucleic acid is DNA or RNA.  
     
     
         4 . The nucleic acid molecule of  claim 3 , wherein the DNA is cDNA.  
     
     
         5 . The nucleic acid molecule of  claim 4 , wherein the cDNA has substantially the same coding sequence as the coding sequence shown in SEQ. ID. No.: 1, SEQ. ID. No.: 3, SEQ. ID. No.: 5, SEQ. ID. No.: 7, SEQ. ID. No.: 9, or SEQ. ID. No.:11.  
     
     
         6 . A vector comprising the nucleic acid molecule of  claim 4 .  
     
     
         7 . The vector of  claim 6 , wherein the vector is a virus or a plasmid.  
     
     
         8 . A host vector system for the production of a mammalian BCNG molecule which comprises the vector of  claim 6  in a suitable host.  
     
     
         9 . The host vector system of  claim 8 , wherein the suitable host is a bacterial cell, a eukaryotic cell, a mammalian cell or an insect cell.  
     
     
         10 . A nucleic acid molecule capable of specifically hybridizing with the nucleic acid molecule of  claim 1 .  
     
     
         11 . An isolated BCNG protein.  
     
     
         12 . The protein of  claim 11 , wherein the BCNG protein is mBCNG-1, mBCNG-2, mBCNG-3, mBCNG-4, hBCNG-1 or hBCNG-2.  
     
     
         13 . The protein of  claim 11 , wherein the BCNG protein has substantially the same amino acid sequence as the amino acid sequence shown in SEQUENCE ID NO.: 2, SEQ. ID. No.: 4, SEQ. ID. No.: 6, SEQ. ID. No.: 8, SEQ. ID. No.: 10, or SEQ. ID. No.: 12.  
     
     
         14 . A composition comprising a nucleic acid, having sequence encoding a BCNG protein or a portion thereof and a carrier.  
     
     
         15 . The composition of  claim 14 , wherein the nucleic acid comprises substantially the same coding sequence as the coding sequence shown in SEQ. ID. No.: 1, SEQ. ID. No.: 3, SEQ. ID. No.: 5, SEQ. ID. No.: 7, SEQ. ID. No.: 9, SEQ. ID. No.:11 or portion thereof.  
     
     
         16 . A composition comprising a BCNG protein or portion thereof and a carrier.  
     
     
         17 . The composition of  claim 16 , wherein the BCNG protein comprises substantially the same amino acid sequence as the amino acid sequence shown in SEQ. ID. No.: 2, SEQ. ID. No.: 4, SEQ. ID. No.: 6, SEQ. ID. No.: 8, SEQ. ID. No.: 10, SEQ. ID. No.:12 or portion thereof.  
     
     
         18 . A method of identifying a nucleic acid encoding an ion channel subunit-related protein in a sample which comprises detecting in the sample a nucleic acid molecule encoding a BCNG-related protein, positive detection indicating the presence of an ion channel subunit related protein encoding nucleic acid molecule, thereby identifying a nucleic acid encoding an ion channel subunit-related protein in the sample.  
     
     
         19 . The method of  claim 18 , wherein the nucleic acid molecule encoding the ion channel subunit related protein is detected by size fractionation.  
     
     
         20 . The method of  claim 19 , wherein the size fractionation is effected by a polyacrylamide or an agarose gel.  
     
     
         21 . The method of  claim 20 , wherein the detection of the nucleic acid molecule encoding the BCNG-related protein comprises contacting the sample with a second nucleic acid molecule capable of hybridizing with the nucleic acid molecule encoding the BCNG-related protein, wherein the second nucleic acid molecule is labeled with a detectable marker under conditions permitting the second nucleic acid molecule to hybridize with the nucleic acid molecule encoding the BCNG-related protein, thereby identifying the nucleic acid molecule in the sample.  
     
     
         22 . The method of  claim 21 , wherein the second nucleic acid molecule is capable of hybridizing to mBCNG-1, mBCNG-2, mBCNG-3, mBCNG-4, hBCNG-1 or hBCNG-2.  
     
     
         23 . The method of  claim 21 , wherein the detectable marker is a radiolabeled molecule, a fluorescent molecule, an enzyme, a ligand, or a magnetic bead.  
     
     
         24 . The method of  claim 18 , wherein the detection of the nucleic acid molecule encoding the BCNG-related protein comprises amplifying the nucleic acid molecule encoding the BCNG-related protein, thereby identifying the nucleic acid molecule encoding the BCNG-related protein in the sample.  
     
     
         25 . The method of  claim 24 , wherein the amplification of the nucleic acid molecule encoding the BCNG-related protein comprises contacting the sample with at least one primer capable of hybridizing to mBCNG-1, mBCNG-2, mBCNG-3, mBCNG-4, hBCNG-1 or hBCNG-2 under conditions suitable for polymerase chain reaction.  
     
     
         26 . The method of  claim 25 , wherein the amplified nucleic acid molecule encoding the BCNG-related protein is detected by size fractionation.  
     
     
         27 . The method of  claim 26 , wherein the size fractionation is via a polyacrylamide gel or an agarose gel.  
     
     
         28 . A method for evaluating the ability of a compound to modulate an ion channel associated with a condition in a subject which comprises: 
 (a) contacting a cell which expresses a BCNG-related protein in a cell culture with a compound;    (b) determining the amount of BCNG-related protein expression in the cell culture; and    (c) comparing the amount of BCNG-related protein expression determined in step (b) with the amount determined in the absence of the compound so as to evaluate the ability of the compound to modulate the ion channel associated with the condition in the subject.    
     
     
         29 . The method of  claim 28 , wherein the condition is a neurological condition, cardiovascular condition, renal condition, pulmonary condition, or hepatic condition.  
     
     
         30 . The method of  claim 28 , wherein the condition is epilepsy, Alzheimer's Disease, Parkinson's Disease, long QT syndrome, sick sinus syndrome, age-related memory loss, cystic fibrosis, sudden death syndrome or a pacemaker rhythm dysfunction.  
     
     
         31 . The method of  claim 28 , wherein the BCNG-related protein comprises mBCNG-1, mBCNG-2, mBCNG-3, MBCNG-4, hBCNG-1 or hBCNG-2 or a portion thereof.  
     
     
         32 . The method of  claim 28 , wherein the cell is a cardiac cell, a kidney cell, a hepatic cell, an airway epithelial cell, a muscle cell, a neuronal cell, a glial cell, a microglial cell, an endothelial cell, a mononuclear cell, a tumor cell, a mammalian cell, an insect cell, or a  Xenopus  oocyte.  
     
     
         33 . The method of  claim 28 , wherein the compound is a peptide, a peptidomimetic, a nucleic acid, a polymer, or a small molecule.  
     
     
         34 . The method of  claim 28 , wherein the compound is bound to a solid support.  
     
     
         35 . A method for evaluating the ability of a compound to interact with a BCNG-related ion channel subunit protein which comprises: 
 (a) contacting a cell which expresses a BCNG-related protein in a cell culture with a compound under conditions permissive to formation of a complex between the compound and the BCNG-related protein;    (b) determining the amount of complex formed between the compound and the BCNG-related protein in the cell culture; and    (c) comparing the amount of complex formed in step (b) with the amount formed in the absence of the compound so as to evaluate the ability of the compound to interact with a BCNG-related ion channel subunit protein.    
     
     
         36 . The method of  claim 35 , wherein the BCNG-related protein comprises mBCNG-1, mBCNG-2, mBCNG-3, mBCNG-4, hBCNG-1 or hBCNG-2 or a portion thereof.  
     
     
         37 . The method of  claim 35 , wherein the cell is a cardiac cell, a kidney cell, a hepatic cell, an airway epithelial cell, a muscle cell, a neuronal cell, a glial cell, a microglial cell, an endothelial cell, a mononuclear cell, a tumor cell, a mammalian cell, an insect cell, or a  Xenopus  oocyte.  
     
     
         38 . The method of  claim 35 , wherein the compound is a peptide, a peptidomimetic, a nucleic acid, a polymer, or a small molecule.  
     
     
         39 . The method of  claim 35 , wherein the compound is bound to a solid support.  
     
     
         40 . A method for identifying whether a compound is capable of modulating a dysfunction in an animal comprising: 
 (a) administering the compound to the animal under conditions permissive to formation of a complex between the compound and the BCNG-related protein;    (b) determining the amount of complex formed between the compound and the BCNG-related protein in the animal; and    (c) comparing the amount of complex formed in step (b) with the amount of complex formed in the animal in the absence of the compound so as to identify whether the compound is capable of modulating the dysfunction in the animal, a change in the amount of complex formed in the presence of the compound indicating that the compound is capable of modulating dysfunction in the animal.    
     
     
         41 . The method of  claim 40 , wherein the condition is a neurological condition, cardiovascular condition, renal condition, pulmonary condition, or hepatic condition.  
     
     
         42 . The method of  claim 41 , wherein the condition is epilepsy, Alzheimer's Disease, Parkinson's Disease, long QT syndrome, sick sinus syndrome, age-related memory loss, cystic fibrosis, sudden death syndrome or a pacemaker rhythm dysfunction.  
     
     
         43 . A method of identifying a compound which modulates the activity of BCNG-related protein which comprises: 
 (a) introducing the nucleic acid of  claim 1  into an expression system and causing the expression system to express the nucleic acid under conditions whereby an ion channel subunit protein is produced;    (b) contacting the channel subunit protein with the compound;    (c) determining the activity of the ion channel subunit protein; and    (d) comparing the activity determined in step (c) with the activity determined in the absence of the compound, an increase or decrease in activity in the presence of the compound indicating identification of a compound which modulates the activity of BCNG-related protein.    
     
     
         44 . The method of  claim 43 , wherein the activity determination step (c) comprises measuring the channel electrical current or intracellular calcium level in the presence of the compound.  
     
     
         45 . The method of  claim 43 , wherein the expression system comprises a cultured host cell.  
     
     
         46 . The method of  claim 45 , wherein the cultured host cell is a cardiac cell, a kidney cell, a hepatic cell, an airway epithelial cell, a muscle cell, a neuronal cell, a glial cell, a microglial cell, an endothelial cell, a mononuclear cell, a tumor cell, a mammalian cell, an insect cell, or a  Xenopus  oocyte.  
     
     
         47 . A method of treating a condition in a subject which comprises administering to the subject an effective amount of a compound which modulates the activity of BCNG-related protein identified by the method of  claim 43 , so as to treat the condition.  
     
     
         48 . The method of  claim 47 , wherein the condition is a neurological condition, cardiovascular condition, renal condition, pulmonary condition, or hepatic condition.  
     
     
         49 . The method of  claim 48 , wherein the condition is epilepsy, Alzheimer's Disease, Parkinson's Disease, long QT syndrome, sick sinus syndrome, age-related memory loss, cystic fibrosis, sudden death syndrome or a pacemaker rhythm dysfunction.  
     
     
         50 . A pharmaceutical composition which comprises a compound capable of modulating BCNG-related protein activity identified by the method of  claim 43 , and a pharmaceutically acceptable carrier.  
     
     
         51 . The pharmaceutical composition of  claim 50 , wherein the carrier is a diluent, an aerosol, a topical carrier, an aqueous solution, a nonaqueous solution or a solid carrier.  
     
     
         52 . A method for treating a condition in a subject which comprises administering to the subject an amount of the pharmaceutical composition of  claim 51 , effective to treat the condition in the subject.  
     
     
         53 . The method of  claim 52 , wherein the condition is a neurological condition, renal condition, pulmonary condition, hepatic condition, or cardiovascular condition.  
     
     
         54 . The method of  claim 53 , wherein the condition is epilepsy, Alzheimer's Disease, Parkinson's Disease, long QT syndrome, sick sinus syndrome, age-related memory loss, cystic fibrosis, sudden death syndrome or a pacemaker rhythm dysfunction.  
     
     
         55 . The method of  claim 52 , wherein the subject is a human.  
     
     
         56 . An antibody which binds specifically to the protein of  claim 11 .  
     
     
         57 . A cell line producing the antibody of  claim 56 .  
     
     
         58 . A method of identifying the protein of  claim 11  in a sample comprising: 
 (a) contacting the sample with the antibody of  claim 56  under conditions permissive to the formation of a complex between the antibody and the protein;  
 (b) determining the amount of complex formed; and  
 (c) comparing the amount of complex formed in step (b) with the amount of complex formed in the absence of the antibody, the presence of an increased amount of complex formed in the presence of the antibody indicating identification of the protein in the sample.