We have studied various properties of the binding of
5 alpha-dihydrotestosterone (DHT) or the synthetic, nonmetabolizable
androgen methyltrienolone (
R1881; 17 beta-hydroxy-17 alpha-methylestra-4,9,11-
trien-3-one) to the
androgen receptor of genital skin fibroblasts (GSF) from controls and a subject with familial, receptor-positive,
partial androgen insensitivity. The mutant cells form R1881-receptor complexes that dissociate 7 times more rapidly than normal at 30 degrees C, and they do not increase their specific R1881-receptor activity in response to a 72-hr period of incubation with
R1881, whereas the cells from normal individuals do so two- to three-fold. As previously reported [Pinsky et al, 1981; Kaufman et al, 1981], the mutant cells have similar abnormalities with DHT as with
R1881. When the patient's cells are incubated for 120 min with varying concentrations (0.05-3 nM) of
R1881 or DHT, Scatchard analysis shows that their
androgen-receptor activity has an apparent equilibrium dissociation constant (Kd) for each
ligand that is six-fold greater than that of normal cells (approximately 0.2 nM). Normal GSF have higher, more variable values of Kd (0.3-1.8 nM) for either
ligand after 30 compared to 120 min of incubation, and the 60-min values are intermediate. This explains why we previously reported that the patient's cells had a 30-min Kd for DHT in the normal range [Pinsky et al, 1981].
Sucrose gradient centrifugation of mutant GSF cytosol incubated with DHT in the presence of 10 mM
sodium molybdate yields a normal 6.5-8S peak of DHT-receptor complexes. From these data we conclude that normal GSF form initial, low-affinity
androgen-receptor complexes that are transformed into one (or more) higher-affinity (? activated) states by a process that depends on time and initial concentration of
androgen; the subject's GSF can form low-affinity
androgen-receptor complexes but cannot generate the normal high-affinity state of the complexes, and this lack precludes augmentation of their
androgen-receptor activity in response to prolonged incubation with either
androgen; and failure of
molybdate to stabilize the
androgen-receptor activity in GSF cytosol is not a more sensitive
indicator of structurally altered
androgen-receptor proteins than are other qualities described heretofore.