The androgen receptor (AR) is essential for development of the male gender
and in the growth of the majority of prostate cancers. Agonists as well as
most antagonists induce translocation of the receptor to the nucleus,
whereas only agonists can activate AR function. Antagonists are therefore
used in the therapy of metastasized prostate cancer. To obtain insight
into the mechanism by which antagonists block AR function in living cells,
we studied nuclear mobility and localization of green fluorescent protein
(GFP)-tagged AR in the presence of either the agonist R1881 or the
antagonists bicalutamide and hydroxyflutamide. As controls we investigated
a non-DNA-binding AR mutant (A573D) and two mutants (W741C and T877A) with
broadened ligand specificity. We demonstrate that in the presence of
R1881, AR localizes in numerous intranuclear foci and, using complementary
fluorescence recovery after photobleaching (FRAP) approaches and computer
modelling, that a fraction of AR ( approximately 10-15%) is transiently
immobilized in a DNA-binding-dependent manner (individual ARs being
immobile for approximately 45 seconds). By contrast, antagonist-bound
GFP-AR showed no detectable immobile fraction and the mobility was similar
to that of the R1881-liganded non-DNA-binding mutant (A573D), indicating
that antagonists do not induce the relatively stable DNA-binding-dependent
immobilization observed with agonist-bound AR. Moreover, in the presence
of bicalutamide and hydroxyflutamide GFP-AR was homogeneously distributed
in the nucleus. Binding of bicalutamide and hydroxyflutamide to
GFP-AR(W741C) and GFP-AR(T877A), respectively, resulted in similar
mobility and heterogeneous nuclear distribution as observed for
R1881-liganded GFP-AR. The live cell studies indicate that the
investigated antagonists interfere with events early in the
transactivation function of the AR