Mitogen-activated protein kinase kinase MEK6 and methods of use thereof

Abstract

Compositions and methods are provided for potentiating the activity of the mitogen-activated protein kinase p38. In particular the mitogen-activated protein kinase kinase MEK6, and variants thereof that stimulate phosphorylation of p38 are provided. Such compounds may be used, for example, for therapy of diseases associated with the p38 cascade and to identify antibodies and other agents that inhibit or activate signal transduction via p38.

1 . A An isolated polypeptide comprising the amino acid sequence provided in SEQ ID NO:2.  
     
     
         2 . A constitutively active variant of a polypeptide according to  claim 1 .  
     
     
         3 . A polypeptide comprising the amino acid sequence provided in SEQ ID NO:2 modified at no more than 10% of the amino acid residues, such that said polypeptide is rendered constitutively inactive.  
     
     
         4 . An isolated DNA molecule encoding a polypeptide according to any of claims  1 - 3 .  
     
     
         5 . An isolated DNA molecule comprising the nucleotide sequence provided in SEQ ID NO: 1.  
     
     
         6 . A recombinant expression vector comprising a DNA molecule according to  claim 4 .  
     
     
         7 . A host cell transformed or transfected with an expression vector according to  claim 6 .  
     
     
         8 . A method for phosphorylating p38 comprising contacting p38 with a polypeptide according to either of claims  1  or  2 , thereby phosphorylating p38.  
     
     
         9 . A method for activating a member of the p38 cascade in an organism, comprising administering to an organism a polypeptide according to either of claims  1  or  2 , thereby activating a member of the p38 cascade.  
     
     
         10 . The method of  claim 9  wherein the member of the p38 cascade is p38.  
     
     
         11 . A method for screening for an agent that inhibits signal transduction via the p38 cascade, comprising: 
 (a) contacting a candidate agent with a polypeptide according to either of claims  1  or  2 ; and    (b) subsequently measuring the ability of said polypeptide to activate p38, and thereby evaluating the ability of the compound to inhibit signal transduction via the p38 cascade.    
     
     
         12 . A method for screening for an agent that stimulates signal transduction via the p38 cascade, comprising: 
 (a) contacting a candidate agent with a polypeptide according to either of claims  1  or  2 ; and    (b) subsequently measuring the ability of said polypeptide to activate p38, and thereby evaluating the ability of the compound to stimulate signal transduction via the p38 cascade.    
     
     
         13 . A monoclonal antibody that binds to a polypeptide according to either of claims  1  or  2 .  
     
     
         14 . A monoclonal antibody according to  claim 13 , wherein said antibody inhibits the phosphorylation of p38 by said polypeptide.  
     
     
         15 . A method for treating a patient afflicted with a disease associated with the p38 cascade, comprising administering to a patient a compound that inhibits the phosphorylation of p38 by MEK6.  
     
     
         16 . The method of  claim 15  wherein said compound is a monoclonal antibody.  
     
     
         17 . The method of  claim 15  wherein said compound comprises a nucleotide sequence.  
     
     
         18 . A method for detecting MEK6 kinase activity in a sample, comprising evaluating the ability of the sample to phosphorylate p38, thereby detecting MEK6 kinase activity in the sample.  
     
     
         19 . A kit for detecting MEK6 kinase activity in a sample, comprising p38 in combination with MEK6 and a suitable buffer.  
     
     
         20 . A method for identifying a composition which affects MEK6 kinase activity, comprising: 
 (a) incubating the composition and MEK6 kinase or polynucleotide encoding the kinase, wherein the step of incubation is carried out under conditions and for a time sufficient to allow the components to interact; and    (b) measuring the effect of the composition on MEK6 kinase or polynucleotide encoding the kinase.    
     
     
         21 . A method of treating an immunologically related disorder associated with MEK6 kinase activity, comprising administering to a subject having the disorder a therapeutically effective amount of a compound which modulates MEK6 kinase activity.  
     
     
         22 . The isolated polypeptide of  claim 1  wherein said polypeptide also comprises additional amino acid residues at the N-terminus or C-terminus.  
     
     
         23 . An isolated polypeptide comprising a variant of the amino acid sequence provided in SEQ ID NO:2 that differs from SEQ ID NO:2 at 10% or less of the amino acid residues, wherein said polypeptide is capable of phosphorylating a p38 substrate.  
     
     
         24 . The isolated polypeptide of  claim 23  wherein the ability of said polypeptide to stimulate p38 phosphorylation is not substantially diminished as compared to a polypeptide having the amino acid sequence provided in SEQ ID NO:2.  
     
     
         25 . The isolated polypeptide of  claim 23  wherein said polypeptide is constitutively active.  
     
     
         26 . A purified antibody that binds to a polypeptide according to any of claims  1 ,  22 ,  23 ,  24 , or  25 .  
     
     
         27 . An antibody according to  claim 26  which is a monoclonal antibody.  
     
     
         28 . An antibody according to  claim 26  which is a polyclonal antibody.  
     
     
         29 . An antibody according to  claim 26 , wherein said antibody inhibits the phosphorylation of p38 by said polypeptide.  
     
     
         30 . A substantially pure human mitogen-activated protein kinase kinase (MKK) polypeptide having serine, threonine, and tyrosine kinase activity, and human mitogen-activated protein (MAP) kinase p38 phosphorylating activity comprising the amino acid sequence of SEQ ID NO:2.  
     
     
         31 . A substantially pure human mitogen-activated protein kinase kinase (MKK) polypeptide having human mitogen-activated protein (MAP) kinase p38 phosphorylating activity comprising the amino acid sequence of SEQ ID NO:2.  
     
     
         32 . A substantially pure human mitogen-activated protein kinase kinase (MKK) polypeptide comprising the amino acid sequence of SEQ ID NO:2.  
     
     
         33 . A purified antibody which specifically binds to a polypeptide of  claim 30 .  
     
     
         34 . A purified antibody which specifically binds to a polypeptide of  claim 31 .  
     
     
         35 . A purified antibody which specifically binds to a polypeptide of  claim 32 .  
     
     
         36 . A purified monoclonal antibody which specifically binds to a polypeptide of  claim 30 .  
     
     
         37 . A purified monoclonal antibody which specifically binds to a polypeptide of  claim 31 .  
     
     
         38 . A purified monoclonal antibody which specifically binds to a polypeptide of  claim 32 .  
     
     
         39 . A purified polyclonal antibody which specifically binds to a polypeptide of  claim 30 .  
     
     
         40 . A purified polyclonal antibody which specifically binds to a polypeptide of  claim 31 .  
     
     
         41 . A polyclonal antibody which specifically binds to a polypeptide of  claim 32.