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In the present study, steroidogenesis in two different Leydig tumor cell lines was compared. One, the MA-10 mouse tumor cell line, produces steroids and the steroidogenic acute regulatory (StAR) protein only when stimulated by trophic hormones and cAMP analogs. The other, the R2C rat tumor cell line, produces steroids and the StAR protein constitutively without stimulation. We observed that high levels of DAX-1 (dosage-sensitive sex reversal, adrenal hypoplasia congenita, critical region on the X chromosome, gene-1), a repressor of steroidogenesis and StAR gene expression, were present in MA-10 cells but not in R2C cells. Based upon this observation, we hypothesized that the absence of DAX-1 might result in constitutive steroidogenesis in R2C cells. To test this hypothesis, DAX-1 was overexpressed in the R2C cells using the Tet-on inducible gene expression system and resulted in a 45% decrease in steroid production, a 35% decrease in StAR protein, and a 39% decrease in cytochrome P450 side chain cleavage expression. Further, using retroviral infection with DAX-1, StAR expression and steroidogenesis were decreased 50-60% and 60% in R2C cells, respectively. These results corroborate previous findings that DAX-1 negatively regulates steroid synthesis through the inhibition of StAR expression and indicate that the absence of DAX-1 in R2C cells is, at least in part, responsible for the constitutive steroidogenesis and StAR expression observed.

endocrinology

Regulation of Steroidogenesis and Steroidogenic Acute Regulatory Protein in R2C Cells by DAX-1 (Dosage-Sensitive Sex Reversal, Adrenal Hypoplasia Congenita, Critical Region on the X Chromosome, Gene-1)