Signaling Cross-talk from Gβ4 Subunit to Elk-1 in the Rapid Action of Androgens

Androgens act on transcription via intracellular androgen receptors (ARs), but they also have rapid AR-independent effects. We have identified the multistep processes involved in the rapid actions of androgens in male osteoblasts, which also possess the classical AR. Incubating cells with 5α-dihydroxytestosterone (100 p m , DHT) rapidly increased (1 min) the phosphorylation of the transcription factor Elk-1, and this was inhibited by pertussis toxin (PTX). DHT activated ERK1/2, a substrate of Elk-1, within 15 s but had no effect on p38 MAPK or JNK/SAPK. The inhibitors PD98059 (MEK1/2); Gö6976, Gö6983, and chelerythrine (protein kinase C); wortmannin and LY294002 (phosphatidylinositol 3-kinase); PP1 (Src); and PTX all blunted the DHT-stimulated phosphorylation of ERK1/2. DHT increased the phosphorylation of c-Raf-1 within 5 s; this was blocked by conventional protein kinase C and phosphatidylinositol 3-kinase inhibitors. The first activated membrane protein was the PTX-sensitive Gβ 4 subunit coupled to phospholipase C-β2, which triggered a rapid (5 s) increase in intracellular calcium and diacylglycerol formation. The androgen antagonist cyproterone acetate did not modify the responses to DHT. Lastly an anti-AR antibody directed against the ligand binding domain recognized a protein at the plasma membrane. The cascade of rapid effects triggered by androgens may involve the classical AR at the plasma membrane or an uncharacterized form of AR that is insensitive to nuclear antagonists.