THE ROLE OF ERK2 1 IN STEROID PRODUCTION AND StAR PROTEIN EXPRESSION BY Y1 CELLS: STUDIES USING StAR OVER-EXPRESSING TRANSFECTS

In studies using inhibitors of MEK-1, the upstream activator of ERK2 1, we have recently demonstrated that the 42 and 44 kDa extracellular signal-regulated kinases (ERK2 1, respectively) play a major role in regulating cyclic AMP-stimulated StAR gene expression in Y1 mouse adrenocortical cells. We have now extended these studies with the use of a stably-transfected Y1 cell-line over-expressing StAR mRNA under control of CMV promoter control. In these cells, increased StAR expression was correlated with enhanced basal steroidogenesis that was not inhibited by MEK-1 inhibitors (untransfected: 0.7 ± 0.3 pmol pregnenolone 106 cells, transfected: 4.8 ± 1.7 pmol pregnenolone 106 cells), in accordance with ERK2 1 acting to regulate steroid production before transcription of the StAR gene. However, StAR mRNA and protein expression and steroid production by transfected cells were further enhanced in the presence forskolin (FSK, 1 µM) and to a much greater extent than in control cells (untransfected: 4.4 ± 0.2 pmol pregnenolone 106 cells, transfected: 10.4 ± 0.3 pmol pregnenolone 106 cells). Furthermore, MEK-1 inhibitors did not reduce the FSK-induced increase in StAR mRNA and protein expression in transfected cells. These results suggest that activation of adenylate cyclase by FSK can regulate the expression of StAR by both increasing transcription of the endogenous gene and by enhancing stability of existing mRNA. They further suggest that ERK2 1 activation is required for FSK-induced StAR mRNA transcription but is not involved in the regulation of mRNA stability by FSK.