Regulation of the synthesis of lutropin-induced protein in rat testis Leydig cells

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Abstract
The mechanism of action of lutropin [luteinizing hormone] on the stimulation of the synthesis of a specific lutropin-induced protein in rat testis Leydig cells was investigated. Lutropin-induced protein has a MW of approx. [approximately] 21,000 and is detected by labeling the Leydig-cell proteins with [35S]methionine, followed by separation by polyacrylamide-gel electrophoresis and radioautography of the dried gel. The incorporation of 35S into lutropin-induced protein was used as an estimate for the synthesis of the protein. Incubation of Leydig cells with dibutyryl cyclic[c]AMP or cholera toxin also resulted in the stimulation of synthesis of the protein. Synthesis of lutropin-induced protein, when maximally stimulated with 100 ng of lutropin/ml, could not be stimulated further by addition of dibutyryl cAMP. Addition of 3-isobutyl-1-methylxanthine, a phosphodiesterase inhibitor, further increased synthesis of the protein in the presence of a submaximal dose of lutropin (10 ng/ml) but not in the presence of lutropin or with maximal amounts of lutropin (100 and 1000 ng/ml). Actinomycin D prevented the effect of lutropin on the stimulation of lutropin-induced protein synthesis when added immediately or 1 h after the start of the incubation, but not when added after 5-6 h. This is interpreted as reflecting that, after induction of mRNA coding for lutropin-induced protein, lutropin had no influence on the synthesis of the protein in the presence of actinomycin D. Synthesis of the protein was also stimulated in vivo by injection of choriogonadotropin into rats 1 day after hypophysectomy, and the time course of this stimulation of lutropin-induced protein synthesis in vivo was similar to that obtained by incubating Leydig cells in vitro with lutropin. From these results it is concluded that stimulation of lutropin-induced protein synthesis by lutropin is most probably mediated by cAMP and involves synthesis of mRNA.