Further evidence that the mitochondrial proteins induced by hormone stimulation in MA-10 mouse Leydig tumor cells are involved in the acute regulation of steroidogenesis.

In previous studies we and others have described several mitochondrial proteins which are synthesized in response to acute hormone stimulation in several steroidogenic tissues. In both MA-10 mouse Leydig tumor cells and primary cultures of rat adrenal cortex cells, these proteins consist of a family of 37 kilodalton (kDa) and 32 kDa precursor forms and fully processed forms which are 30 kDa in molecular weight. The nature of the appearance of these proteins and their subcellular localization to the mitochondria, the site of the rate limiting step in steroidogenesis, has led to the speculation that they may be involved in the acute regulation of steroidogenesis. In the present study we have taken advantage of another steroidogenic cell, the R2C rat Leydig tumor cell, to perform studies which further indicate that these mitochondrial proteins are involved in the regulation of steroidogenesis. Unlike the MA-10 cell which requires hormone stimulation for steroid production, the R2C cell is a constitutive progesterone producer whose steroid production cannot be further increased with hormone stimulation. We have shown that the R2C cell line is less sensitive to the inhibition of steroid production by the metal chelator orthophenanthroline (OP) than is the MA-10 cell. We have demonstrated that progesterone production and the 30 kDa mitochondrial proteins remain present in the R2C cells at a concentration of OP which completely inhibits progesterone production and totally eliminates the 30 kDa proteins in MA-10 cells. As further evidence for the role of these proteins in steroidogenic regulation, we have isolated several revertants of the R2C parent (P) cell line which have lost the ability to synthesize progesterone constitutively, but which can be stimulated to synthesize this steroid by trophic hormone and cAMP analog. In these revertants, designated (R), the normally constitutively present 30 kDa proteins are greatly decreased compared to controls, but reappear in large amounts following hormone stimulation. Taken together, these data provide further evidence that the 30 kDa mitochondrial proteins are involved in the acute regulation of steroidogenesis in Leydig cells.