Mechanisms that regulate androgen receptor transcriptional activity

Testosterone (T) and dihydrotestosterone (DHT) are natural ligands for the androgen receptor (AR), an intracellular transcription factor and nuclear receptor. DHT is a more potent androgen than T in vivo. In this study, the mechanistic basis for the differential effects of T and DHT on AR activity was investigated. Dissociation kinetics, motif binding affinity and activation function 2 (AF2) transactivation measurements reveal that the slow dissociation of DHT relative to T from AR results from weaker T-induced AR FXXLF motif binding to the AF2 site. T acquires DHT-like activity when the AR ligand binding domain (LBD) contains the H874Y somatic prostate cancer mutation that results in the formation of direct hydrogen bonds between external and core helices 4 and 5, improving AF2 motif binding. The studies reveal that DHT better stabilizes the AR LBD core from the ligand binding pocket to the AF2 surface for maximal AR transactivation.;To further define the mechanisms whereby the AR specific coregulator melanoma antigen gene protein-A11 (MAGE-11) modulates AR activity, we pursued observations that MAGE-11 increases AR transcriptional activity independent of AF2. We sought to characterize the effects of MAGE-11 and the coactivators transcription intermediary factor 2 (TIF2) and p300 on AR transcriptional activity. The site of interaction in MAGE-11 that binds the AR FXX LF motif is an F-box within the MAGE homology domain. MAGE-11 Ser-174 is phosphorylated by MAP kinase which influences the interaction of the MAGE-11 F-box with AR. MAGE-11 forms a complex with TIF2 and p300 to modulate AR transactivation independent of AF2. This research provides evidence for a novel function for an F-box protein in which F-box/FXXLF like motif interactions modulate AR transcriptional activity in the absence and presence of ligand.