Hormone-independent repression of AP-1-inducible collagenase promoter activity by glucocorticoid receptors.

The role of the ligand in glucocorticoid receptor-mediated transactivation and transrepression of gene expression was investigated. Half-maximal transactivation of a mouse mammary tumor virus-chloramphenicol acetyltransferase reporter gene in transfected cells expressing the human glucocorticoid receptor mutant GRL753F, from which the rate of ligand dissociation is four to five times higher than the rate of dissociation from normal receptors, required a 200- to 300-fold-higher concentration of dexamethasone than was required in cells expressing the normal receptor. Immunocytochemical analysis demonstrated that this difference was not the result of a failure of the mutant receptor to accumulate in the nucleus after steroid treatment. In contrast, in cells cotransfected with a reporter gene containing the AP-1-inducible collagenase gene promoter, the concentration of dexamethasone required for 50% transrepression was the same for mutant and normal receptors. Efficient receptor-mediated transrepression was also observed with the double mutant GRL753F/C421Y, in which the first cysteine residue of the proximal zinc finger has been replaced by tyrosine, indicating that neither retention of the ligand nor direct binding of the receptor to DNA is required. RU38486 behaved as a full agonist with respect to transrepression. In addition, receptor-dependent transrepression, but not transactivation, was observed in transfected cells after heat shock in the absence of the ligand. Taken together, these results suggest that unlike transactivation, transrepression of AP-1 activity by the nuclear glucocorticoid receptor is ligand independent.     

Identification of two transactivation domains in the mouse oestrogen receptor.

We have identified two discrete transactivation domains within the mouse oestrogen receptor whose relative activities vary according to the target promoter. One domain lies within the N-terminal region and is active in the absence of oestradiol. The second domain is contained within the C-terminal portion of the protein and depends upon oestrogen binding for its activity. The location and oestrogen dependence of this domain has been confirmed using chimaeric receptors containing the Lex A DNA binding domain. Although transactivation by the C-terminal domain is dependent upon ligand binding the analysis of receptor deletion mutants has demonstrated that these two functions are not entirely coincident.     

Identification of constitutive and steroid-dependent transactivation domains in the mouse oestrogen receptor

We have identified two transactivation domains in the mouse oestrogen receptor whose activities depend on the target promoter. The major domain is contained within the C-terminal portion of the protein and depends upon oestrogen binding for its activity. The location and oestrogen dependence of this domain has been confirmed using chimaeric receptors containing the Lex A DNA binding domain. Although transactivation by the C-terminal domain is dependent upon ligand binding the analysis of receptor deletion mutants has demonstrated that these two functions are not entirely coincident. The second transactivation domain lies within the N-terminal region and is active in the absence of oestradiol. The differences in oestrogen requirement for the activity of the two transactivation domains may account for the partial agonist activity of certain antihormones.