SEQUENCES IN THE LIGAND-BINDING DOMAINS OF THE HUMAN ANDROGEN AND PROGESTERONE RECEPTORS WHICH DETERMINE THEIR DISTINCT LIGAND IDENTITIES

The natural ligands of the progesterone (PR) and androgen (AR) recepto rs, progesterone and testosterone, differ only by their 17 beta-substi tution. To identify within the AR and PR ligand-binding domains (LBDs) the sequences responsible for the differential recognition of these l igands, chimeric LBDs assembled from five homologous AR/PR 'cassettes' linked to the GAL4-DNA binding domain were constructed, and their lig and binding and transactivation characteristics were determined. Repla cing the central cassette 3 of PR by that of AR generated a progestero ne- and testosterone-responsive PR LED with the AR residues 788-RHLS-7 91 being specifically involved in testosterone recognition, while the introduction of the C-terminal PR cassette 5 into AR conferred progest in responsiveness onto the AR LED. These results suggest that residues within AR 788-RHLS-791 interact with the testosterone 17 beta-OH, whi le PR cassette 5 apparently contains the amino acid(s) specifically in volved in the recognition of the progesterone 17 beta-acetyl group. Ho wever, ligand binding and transactivation by these chimeras were signi ficantly decreased compared with those of the parental LBDs, indicatin g that residues located outside of these cassettes contribute to the p roper positioning of the steroids in the AR and PR ligand-binding Dock ets (LBPs). Indeed, certain AR/PR chimeras acquired efficient ligand b inding, but were unable to transactivate, indicating that the ligand w as improperly bound in the chimeric LBP and could not induce the confo rmational changes leading to a transcriptionally competent activation function (AF-2) within the LED. The properties of the various LED chim eras are discussed in view of the recently solved three-dimensional st ructures of the retinoid X receptor alpha apo- and retinoic acid recep tor gamma holo-LBDs.