Étude fonctionnelle de mutations du gène du récepteur V2 responsables d'un diabète insipide néphrogénique congénital

Abstract

In previous reports the amount of rat uterine progesterone receptor detected in crude nuclear pellets was only 50% of that which was depleted from the cytosol after an injection of progesterone. We have previously shown that this discrepancy is due to a 50% decrease in receptor binding in the nuclear pellets during the post-assay washing procedure (Walters & Clark, 1978). If this loss were due to the dissociation of receptor-progesterone complexes from nuclear sites, it should be possible to detect and recover them by adding hydroxylapatite to the nuclear pellets at the end of the incubation. This modification of the nuclear exchange assay resulted in 83% greater recovery of specific [3H]-progesterone binding than in the nuclear pellet alone and accounted for the remainder of the translocated nuclear receptors. Thus, quantitation of nuclear progesterone receptors can be achieved in the rat uterus with this nuclear exchange-HAP assay. This method of trapping solubilized hormone complexes also increased recovery of progesterone receptors from the chick oviduct. In contrast, this procedure did not significantly increase the amount of rat uterine estrogen receptor detected, suggesting that little solubilization of nuclear bound estrogen receptors takes place during incubation and nuclear washing procedures.This simple modification of the exchange assay permits measurement of the total quantity of nuclear bound progesterone receptors in the rat uterus. These results emphasize that receptors may dissociate from nuclear pellets under in vitro conditions which results in an underestimate of the nuclear receptor level that actually existed in vivo.