Transformation of human progesterone receptor in the presence of the progestin (R5020) and the antiprogestin (RU486).

We have examined and compared the binding characteristics and transformation in vitro of human uterine cytosolic progesterone receptor (PR) bound to either the progestin agonist, R5020 or the antiprogestin, RU486. Incubation of cytosol with 5-20 nM [3H]R5020 and [3H]RU486 yielded macromolecular complexes that sedimented in the 4S and 8S regions in 10-35% glycerol gradients. The 8S peaks of radioactivity due to macromolecular-bound [3H]R5020 or [3H]RU486 could be eliminated by a preincubation of the uterine cytosol with 1-2 microM progesterone or RU486. The [3H]R5020 binding in the 4S peak was not competable with either steroid. In contrast, the binding of [3H]RU486 in the 4S peak could be abolished by a pretreatment of uterine cytosol with excess RU486, but not progesterone. Selective fractionation of the cytosol with ammonium sulfate, in the presence of sodium molybdate, eliminated the non-specific 4S [3H]R5020 binder. The thermal (23 degrees C) transformation of the [3H]RU486-receptor complex, as a function of the loss of the area under the 8S peak, appeared to be comparable to that achieved with [3H]R5020-receptor complex. The 8S [3H]RU486 peak was reduced by only 46% compared to the [3H]R5020 peak, which was reduced by 60%. These results demonstrate that in human uterine cytosol, R5020 and RU486 bind in a specific and saturable manner to an 8S PR, which is susceptible to thermal 8S to 4S transformation. In addition, [3H]R5020 also interacts with a nonsaturable 4S macromolecule, whereas the 4S [3H]RU486 binder is saturable and specific for RU486. The above observations indicate the heterogeneity of the steroid binding components present in the human uterine cytosol, and suggest that caution should be taken when interpreting data which shows the presence of different molecular forms of the steroid receptors.