Isolation of mRNA from Bovine Pituitary. The Cell-Free Synthesis of the alpha and beta Subunits of Luteinizing Hormone

Abstract

RNA derived from bovine steer pituitary was translated in wheat germ cell-free extracts containing [35S]methionine. Antisera generated against purified denatured .alpha. and .beta. subunits of lutropin were used to demonstrate the synthesis of both proteins in vitro. The immunoprecipitated products of the cell-free system were resolved on sodium dodecyl sulfate/polyacrylamide gels; the MW of the immunoprecipitated .alpha. subunit protein was .apprx. 14,000, while that of the .beta. protein was estimated to be 16,000. Since the MW of authentic .alpha. and .beta. subunits are 10,600 and 14,000, respectively, the cell-free products presumably represented their pre-protein forms. The ratio of the immunoprecipitated subunit pre-proteins was dependent on the Mg concentration in the translation mixtures; at 2.1 mM, translation of lutropin .alpha. and .beta. mRNA was comparable. RNA isolated from cow pituitary tissue directed the synthesis of 5-fold less of the .alpha. and .beta. immunoprecipitated proteins than did steer RNA. Since the blood levels of gonadal steroids are higher in the cow, lutropin .alpha. and .beta. mRNA biosynthesis apparently is repressed by these steroids. Synthesis of lutropin .alpha. and .beta. subunits may be coordinately expressed in certain physiological situations.