Assessment of mechanisms of thyroid hormone action in mouse Leydig cells: regulation of the steroidogenic acute regulatory protein, steroidogenesis, and luteinizing hormone receptor function.

Recently, we demonstrated that triiodothyronine (T(3)) stimulated steroid hormone biosynthesis and steroidogenic acute regulatory (StAR) protein expression in mLTC-1 mouse Leydig tumor cells through the mediation of steroidogenic factor 1 (SF-1). We now report a dual response mechanism of T(3) on steroidogenesis and StAR expression, and on LH receptor (LHR) expression and binding in mLTC-1 cells. T(3) acutely (8 h), induced a 260% increase in StAR messenger RNA (mRNA) expression over the basal level which was coincident with an increase in progesterone (P) production. In contrast, chronic stimulation with T(3) (beyond 8 h), resulted in an attenuation of StAR expression and P production. This attenuation was most likely caused by a decrease in cholesterol delivery to the inner mitochondrial membrane as demonstrated by incubations with the hydrophilic steroid precursors, 22R hydroxycholesterol and pregnenolone, which restored P synthesis. In similar studies, chronic treatment with T(3) increased the levels of cytochrome P450scc mRNA by 83%, whereas those of cytochrome P450 17alpha-hydroxylase and 3ss-hydroxysteroid dehydrogenase decreased. The diminished response in steroidogenesis following chronic T(3) exposure was not a result of alterations in StAR mRNA stability, but rather was due to inhibition of transcription of the StAR gene. Similar acute stimulatory and chronic inhibitory responses to T(3) were found when LHR mRNA expression and LHR ligand binding were examined. Transfections with an LHR or StAR promoter/luciferase reporter construct demonstrated that a 173-bp fragment of the LHR promoter containing an SF-1 binding motif was involved in T(3) response, as was the SF-1 recognition site at -135 bp in the StAR promoter. Furthermore, the importance of SF-1 in T(3) function was also verified employing mutation in the bases of SF-1 sequences using electrophoretic mobility shift assays. The potential physiological relevance of these findings was demonstrated when similar responses were obtained in mice rendered hypo and hyperthyroid. Collectively, these observations further characterize the thyroid-gonadal connection and provide insights into the mechanisms for a dual regulatory role of thyroid hormone in Leydig cell functions.