Induction and repression of specific estradiol sensitive proteins in the rat corpus luteum

In the pregnant rat estradiol causes luteal cell hypertrophy and a dramatic increase in overall protein synthesis. The aim of this investigation was to determine whether estradiol induces the synthesis of specific proteins in luteal cell subcellular fractions and to examine estradiol-regulated translational products in order to identify a specific protein marker of estradiol action in the corpus luteum. Corpora lutea obtained from day 12 hypophysectomized-hysterectomized pregnant rats treated with or without estradiol (2 cm implant) for 3 days were incubated with Trans35S-label for 8 hours at 37 degrees C in an atmosphere of 95% O2 - 5% CO2. Nuclear, mitochondrial, microsomal and cytosolic subcellular compartments were obtained by differential centrifugation. RNA was extracted and translated using a rabbit reticulocyte lysate reaction mixture. Radiolabeled proteins were separated by SDS-PAGE, visualized by fluorography and quantified by densitometry. Estradiol-treatment significantly increased 35S-amino acid incorporation into luteal proteins. Specific estradiol-induced proteins within cellular fractions included mitochondrial 80, 50, and 32 x 10(-3) Mr proteins; microsomal 100, 80, 32 x 10(-3) Mr proteins; and cytosolic 80, 60, 50 and 14 x 10(-3) Mr proteins. Estradiol-treatment increased the secretion of two proteins (60 and 45 x 10(-3) Mr) while no major estradiol-induced proteins were noted in the nuclear fraction. In vitro translation of mRNA indicated that estradiol markedly enhances the message for a protein with apparent Mr 32 x 10(-3) Mr. The 32 x 10(-3) Mr translated protein appeared as the major estradiol-induced protein in mitochondrial and microsomal fractions. Estradiol also reduced the expression of a 38 x 10(-3) Mr protein (pI 7.0) in the cytosolic compartment. The major estradiol sensitive protein observed in luteal particulate and translation profiles was a 32 x 10(-3) Mr protein with a pI greater than or equal to 8.5. This protein may serve as a potential protein marker of estradiol action in the rat corpus luteum.