Comparison of the binding of [2,4,6,7-3H] estradiol-17β [E23H] to the immature rat uterus under in vivo, in vitro and “cell-free” conditions—II. Nuclear binding

Abstract A study of nuclear-binding of cytosol estradiol-receptor complex in immature rat uteri under in vivo , in vitro and “cell-free” experimental conditions showed that nuclear-binding occurred under different forms which were related to the state and molecular form of the receptor present in the cytosol fraction. Analysis of the KCl soluble nuclear extracts on 0.3 M KCl containing sucrose gradients showed that under in vivo conditions a distinct “5S” peak was present unlike under in vitro and “cell-free” conditions where both “4S” and “5S” peaks were simultaneously present, incubation at higher temperature favoring a higher proportion of “5S”. Under “cell-free” conditions, in the presence of EDTA, the KCl soluble nuclear extract sedimented as a “4S” peak only. On treatment with Dextran-coated charcoal no significant difference in the relative stabilities of the KCl soluble extracts obtained under the three experimental conditions was observed. Addition of excess unlabeled estradiol under in vitro and “cell-free” conditions resulted in a disappearance of the bound E 2 3 H both in the “4S” and “5S” forms, both in the cytosol and the nuclear extracts, demonstrating saturability of these bindings. These data would suggest that nuclear binding of estradiol under in vitro and “cell-free” conditions involve components other than that observed in vivo . Although in vitro and “cell-free” nuclear bindings differ from that observed in vivo , yet activation of the cytosol by temperature promotes nuclear binding in a form approaching that observed under in vivo conditions.