Allosteric control of nuclear hormone receptors

Abstract

Heterodimerization is a common paradigm among eucaryotic transcription factors, though it remains unclear how individual monomers contribute to the overall transcriptional activities of the complex. The 9-cis retinoic acid receptor (RXR) serves as a common heterodimerization partner for several nuclear receptors including the thyroid hormone (T 3 R), retinoic acid (RAR) and vitamin D receptors. A strategy has been devised to examine the transcriptional properties of each receptor individually or when tethered to a heterodimeric partner. It has been found that the intrinsic activity of RXR is masked in RXR-T 3 R and RXR-RAR heterodimers. In contrast, a novel RXR-Nurrl heterodimer described herein is highly responsive to RXR ligands, suggesting that different partners exert unique allosteric control over the RXR response. These findings establish a novel 9-cis retinoic acid response pathway and resolve the paradox as to how T 3 R, RAR and VDR contribute to distinct physiologic pathways while sharing a common RXR subunit.

That which is claimed is:  
     
         1 . A method to suppress the constitutive activity of Nurrl, said method comprising contacting Nurrl with at least the ligand binding domain of RXR.  
     
     
         2 . A method according to  claim 1  wherein the ligand binding domain of RXR is selected from RXRα, RXRβ or RXRγ.  
     
     
         3 . A method to render NurrI-containing cells inducibly responsive to RXR selective ligands, said method comprising contacting said cells with at least the ligand binding domain of RXR.  
     
     
         4 . A method according to  claim 3  wherein the ligand binding domain of RXR is selected from RXRα, RXRβ or RXRγ.  
     
     
         5 . A method to render RXR-containing cells responsive to RXR selective ligands, said method comprising contacting said cells with a silent partner therefor.  
     
     
         6 . A method according to  claim 5 , wherein said silent partner is an isoform of Nurrl.  
     
     
         7 . A method for the identification of nuclear receptor(s) which participate as silent partner(s) in the formation of a heterodimer with RXR, said method comprising introducing into a cell: 
 at least the ligand binding domain of a putative silent partner for RXR,    a chimeric construct containing a GAL4 DNA binding domain and at least the ligand binding domain of RXR, and    a reporter construct, wherein said reporter construct comprises: 
 (a) a promoter that is operable in said cell,  
 (b) a GAL4 response element, and  
 (c) DNA encoding a reporter protein, 
 wherein said reporter protein-encoding DNA is operatively linked to said promoter for transcription of said DNA, and  
 wherein said GAL4 response element is operatively linked to said promoter for activation thereof, and thereafter  
 monitoring expression of reporter upon exposure of the above-described cell to RXR selective ligand(s).  
 
   
     
     
         8 . A method for the identification of nuclear receptor(s) which participate as silent partner(s) in the formation of heterodimer(s) with RXR, said method comprising introducing into a cell: 
 a putative silent partner for RXR,    at least the ligand binding domain of RXR, and    a reporter construct, wherein said reporter construct comprises: 
 (a) a promoter that is operable in said cell,  
 (b) a response element for said putative silent partner, and  
 (c) DNA encoding a reporter protein, 
 wherein said reporter protein-encoding DNA is operatively linked to said promoter for transcription of said DNA, and  
 wherein said response element for said putative silent partner is operatively linked to said promoter for activation thereof, and thereafter  
 monitoring expression of reporter upon exposure of the above-described cell to RXR selective ligand(s).  
 
   
     
     
         9 . A method according to  claim 8  wherein the response element for the putative silent partner has the sequence AAAGGTCA.  
     
     
         10 . A method for identifying ligands selective for heterodimers comprising RXR and a silent partner therefor, said method comprising comparing the level of expression of reporter when cells containing a reporter construct, RXR and silent partner therefor are exposed to test compound, relative to the level of expression of reporter when cells containing a reporter construct, RXR and a member of the steroid/thyroid superfamily which is not a silent partner therefor are exposed to test compound, and selecting those compounds which activate only the combination of RXR and silent partner therefor.