High-affinity binding of [3H]estradiol-17 beta by an estrogen receptor in the liver of the turtle.
Specific [3H]estradiol-17 beta ([3H]E2) binding activity (EBA) with characteristics of an estrogen receptor (ER) was demonstrated in cytosols and nuclear extracts of the female turtle, Chrysemys picta. Three different receptor assays (dextran-coated charcoal assay, hydroxylapatite batch procedure, and DNA-cellulose chromatography) were evaluated in terms of their applicability in analyzing large numbers of samples. For the measurement of cytosolic EBA, the hydroxylapatite batch procedure was found to be the most reliable assay. On the other hand, the dextran-coated charcoal assay was found to be the most appropriate method for the measurement of nuclear EBA. Turtle hepatic EBA binds [3H]E2 with high affinity (cytosolic, 17.4 +/- 2.8 X 10(9) M-1; nuclear, 17.7 +/- 1.9 X 10(9) M-1), limited capacity (cytosolic, 133.7 +/- 4.6 fmol/g tissue; nuclear, 81.1 +/- 9.0 fmol/g tissue), and strict steroid specificity. The EBA bound natural estrogens (E2, estrone, estriol) as well as the nonsteroidal estrogen, diethylstilbestrol, but exhibited little affinity for androgens, progesterone, or corticosterone. The turtle hepatic EBA resembled mammalian and avian ERs in terms of binding characteristics; however, unlike mammalian and avian ERs it was shown to be heat-labile. Incubation at 30 degrees caused rapid loss of [3H]E2 binding activity in both cytosolic and nuclear fractions. The exchange between [3H]E2 and the endogenously bound estrogen was slow at 4 and 15 degrees, but the exchange process was facilitated in the presence of the chaotropic salt, NaSCN. Establishment of quantitation methods for both cytosolic and nuclear forms of EBA will enable future investigation of the mechanism and regulation of estrogen action in the liver of this turtle species.
Department of Biology, Tufts University, Medford, Massachusetts 02155.
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D000818:Animals; D002467:Cell Nucleus / Q000032:analysis; D003600:Cytosol / Q000032:analysis; D004958:Estradiol / Q000378:metabolism*; D005260:Female; D008024:Ligands; D008099:Liver / Q000166:cytology / Q000378:metabolism* / Q000648:ultrastructure; D008722:Methods; D011960:Receptors, Estrogen / Q000032:analysis / Q000378:metabolism*; D013861:Thiocyanates / Q000494:pharmacology; D014316:Tritium; D014426:Turtles / Q000378:metabolism*
D008024:Ligands; D011960:Receptors, Estrogen; D013861:Thiocyanates; D014316:Tritium; D004958:Estradiol