RAPID UPTAKE AND BINDING OF ESTRADIOL-17-BETA-6-(O-CARBOXYMETHYL)OXIME - I-125 LABELED BSA BY FEMALE RAT-LIVER

To investigate potential membrane-mediated responses to estrogen, a me mbrane-impermeant, radioiodinated, steroid-BSA conjugate-estradiol-17 beta-6-(O-carboxymethyl)oxime: I-125-labeled BSA (17 beta-E-6-I-125-BS A)-or related steroid conjugates, or I-125-BSA was injected into femal e Sprague-Dawley rats, and tissues were collected at varying times pos tinjection. The liver, adrenal, and spleen displayed the most prominen t uptake of 17 beta-E-6-I-125-BSA, reaching a maximum of 43 times bloo d levels in sonicated liver samples at 5 min postinjection, but no upt ake of I-125-BSA. Isolation of liver membranes by differential centrif ugation showed that over 50% of recovered radioactivity was in associa tion with microsomes and plasmalemma (P3 fraction) at 30 sec postinjec tion. By 60 min postinjection, over 75% of recovered radioactivity was in association with mitochondrial and lysosomal membranes (P2 fractio n), and less than 10% remained in the P3 fraction, In vitro competitio n assays demonstrated two binding sites in liver P3 fractions, The spl een and liver also showed saturable binding in vivo. These data sugges t the presence of at least one membrane-binding protein for estrogen i n liver, adrenal, and spleen. Initial studies of affinity-purified liv er P3 fractions using ligand blots indicated the presence of two bindi ng proteins, These potential membrane estrogen-binding proteins may be involved in a very rapid shuttling of estrogen from the plasmalemma t o mitochondria and lysosomes.