Prolactin receptors in cultured rat mammary tumor cells. Energy-dependent uptake and degradation of hormone receptors.

After energy depletion by uncouplers of oxidative phosphorylation or inhibitors of electron transport, primary cultures of carcinogen-induced rat mammary tumors have a 2- to 20-fold increase in the number of cell surface prolactin receptors. When energy-depleted cells were treated with 0.15 M NaCl plus 50 mM glycine pH 3, for 1 min at 4 degrees C, 75% of the specific surface-bound 125I-labeled ovine prolactin was removed, but prolactin and its receptor were not destroyed. Using this technique, we found that receptor-bound prolactin can be internalized (becomes resistant to pH 3.0 treatment) and then degraded. The internalization of occupied receptors required energy, was completed 30-60 min before degradation, and was independent of protein synthesis. Hormone degradation (t1/2, 42 min) but not uptake was prevented by NH4Cl or lysosomotropic amines. In the presence of cycloheximide, receptors were lost (t1/2, 62 min) unless such loss was prevented by KCN. After unoccupied receptors were activated by energy depletion, surface receptors were lost when inhibitor was removed and glucose was added. Thus, both occupied and unoccupied prolactin receptors are constantly removed from the cell surface via an energy-dependent uptake mechanism. If the receptor levels are first increased by energy depletion (with or without bound ligand) or if protein synthesis is inhibited, there is a net loss of surface binding sites. Since the receptors reappeared with 15 h after cycloheximide removal, some of the receptors probably are recycled under normal steady state conditions.