Novel manganese superoxide dismutase regulatory elements and uses therefore

Abstract

A novel transcriptional regulatory element which was isolated from the MnSOD gene and which exhibits promoter-enhancer activity is disclosed. The promoter-enhancer activity of the element is further modulated by inflammatory mediators to regulate transcription. Methods of using the promoter-enhancer element to regulate gene expression, and therapeutic uses involving the promoter-enhancer element are also described.

1 . An isolated manganese superoxide dismutase regulatory element consisting of the nucleotide sequence of SEQ ID NO:1 or SEQ NO:5, the regulatory element being capable of causing inducible transcription or expression of an operably linked heterologous polynucleotide.  
     
     
         2 . The isolated regulatory element of  claim 1  operably linked to a heterologous polynucleotide so that, upon activation of the regulatory element, transcription or expression of the heterologous polynucleotide is induced.  
     
     
         3 . An isolated polynucleotide comprising a manganese superoxide dismutase regulatory element operably linked to a heterologous polynucleotide, wherein the regulatory element comprises the nucleotide sequence of SEQ ID NO:5, the regulatory element being capable of causing inducible transcription or expression of the operably linked heterologous polynucleotide.  
     
     
         4 . The isolatead polynucleotide of  claim 3 , wherein the regulatory element comprises the nucleotide sequence of SEQ ID NO:1.  
     
     
         5 . The isolated polynucleotide of  claim 3 , further comprising a heterologous polynucleotide operably linked to the regulatory element so that, upon activation of the regulatory element, transcription or expression of the heterologous polynucleotide is induced.  
     
     
         6 . The isolated polynucleotide of  claim 1 , wherein the heterologous polynucleotide encodes a cytoprotectant.  
     
     
         7 . The isolated polynucleotide of  claim 1  which induces transcription or expression of an operatively linked heterologous polynucleotide in the presence of an inflammatory stimulus.  
     
     
         8 . The isolated polynucleotide of  claim 9 , wherein the inflammatory stimulus is selected from the group consisting of TNF-α, IL-1β, and LPS.  
     
     
         9 . The isolated polynucleotide of  claim 1  which induces transcription or expression of an operatively linked heterologous polynucleotide in the presence of 5-aminosalicylic acid.  
     
     
         10 . The isolated polynucleotide of  claim 1 , wherein the regulatory sequence is operatively linked to a promoter sequence.  
     
     
         11 . The isolated polynucleotide of  claim 10 , wherein the promoter is the Herpes simplex thymidine kinase promoter.  
     
     
         12 . A cell transformed with the isolated polynucleotide of  claim 1 .  
     
     
         13 . An inducible expression system comprising: 
 a) an isolated polynucleotide comprising a regulatory element, wherein the regulatory element comprises a nucleotide sequence selected from the group consisting of SEQ ID NO:5, SEQ NO:1 and nucleotide sequences having at least about 90% identity to SEQ ID NO:1, wherein the regulatory element induces transcription or expression of an operably linked heterologous polynucleotide upon activation; and    b) a compound which activates the regulatory element, or a polynucleotide encoding a compound which activates the regulatory element.    
     
     
         14 . The expression system of  claim 13  wherein the regulatory element is a human regulatory element  
     
     
         15 . The expression system of  claim 13  wherein the regulatory element is a rat regulatory element  
     
     
         16 . The expression system of  claim 13  wherein the compound which activates the regulatory element is an inflammatory stimulus.  
     
     
         17 . The expression system of  claim 16  wherein the compound which activates the regulatory element is selected from the group consisting of TNF-α, IL-1β, and LPS.  
     
     
         18 . The expression system of  claim 13  wherein the compound which activates the regulatory element is 5-aminosalicylic acid.  
     
     
         19 . The expression system of  claim 13  further comprising a heterologous polynucleotide operably linked to the regulatory element.  
     
     
         20 . The expression system of  claim 13  further comprising a promoter operably linked to the regulatory element.  
     
     
         21 . A method of producing a polypeptide comprising introducing the expression system of  claim 19  into a cell under conditions suitable for expression of the heterologous polypeptide.  
     
     
         22 . A method of achieving inducible transcription or expression of a heterologous polynucleotide in a cell, the method comprising introducing into a cell an isolated polynucleotide comprising a manganese superoxide dismutase regulatory element, wherein the regulatory element comprises a nucleotide sequence selected from the group consisting of SEQ ID NO:5, SEQ NO:1 and nucleotide sequences having at least about 90% identity to SEQ ID NO:1, the regulatory element being capable of causing inducible transcription or expression of an operably linked heterologous polynucleotide.  
     
     
         23 . The method of  claim 22  further comprising introducing into the cell an effective amount of a compound which activates the regulatory element to induce transcription or expression of an operatively linked polynucleotide, or a polynucleotide encoding the compound.  
     
     
         24 . The method of  claim 23  wherein the compound is an inflammatory mediator.  
     
     
         25 . The method of  claim 24  wherein the compound is selected from the group consisting of TNF-α, IL-1β, and LPS.  
     
     
         26 . The method of  claim 23  wherein the compound is 5-aminosalicylic acid.  
     
     
         27 . The method of  claim 22  wherein the regulatory element is operatively linked to a heterologous polynucleotide.