TRANSCRIPTIONAL REGULATION OF P450SCC GENE-EXPRESSION IN NEURAL AND STEROIDOGENIC CELLS - IMPLICATIONS FOR REGULATION OF NEUROSTEROIDOGENESIS

Regulation of steroidogenesis in classic endocrine tissues is mediated by transcriptional regulation of the P450scc gene, which encodes the first and rate-limiting cholesterol side-chain cleavage enzyme. We pre viously showed that P450scc messenger RNA is regionally expressed in t he adult rat brain, primary glial cultures, and C6 glioma cells. Expre ssion of P450scc in the brain results in the de novo synthesis of neur osteroids, a class of steroid hormones that are active at gamma-aminob utyric acid, and N-methyl-D-aspartate receptors. We determined whether P450scc expression is transcriptionally regulated in neural cells, us ing the same DNA sequences and nuclear proteins as classic steroidogen ic adrenal and Leydig cells. The transcriptional activity of deletiona l mutants of 2.5 kilobases of the 5'-flanking regulatory region of the rat P450scc gene cloned into a luciferase reporter gene was assessed in mouse adrenocortical Y-1, mouse Leydig MA-10, rat C6 glioma, rat GC somatotrope, and mouse GT1-7 neurosecretory cell lines. P450scc was t ranscriptionally regulated in Y-1, MA-10, and C6 glioma cells, but not in GC or GT1-7 cells. In one region (-94/-35), putative steroidogenic factor-1-binding sites appeared to be critical for the basal transcri ptional activity and cAMP responsiveness in steroidogenic Y-1 and MA-1 0 cells, but had no function in rat C6 cells. DNA sequences between -9 4/-130 mediated both basal and cAMP-inducible transcriptional activity in C6 cells. Gel mobility shift assays showed that one nuclear protei n binding to DNA sequences between -54 and -35 was abundant in MA-10 a nd Y-1 cells, but was absent from C6 cells, whereas another nuclear pr otein, binding to DNA sequences between -94 and -130 was abundant in C 6 cells, but was rare in MA-10 cells and absent from Y-1 and other adr enocortical cells. Although the DNA sequence between -94 and -130 cont ains an Sp1 site, Sp1 did not bind to this site. Nevertheless, this GC -rich region was critical for nuclear protein binding and for basal an d cAMP-induced transcriptional regulation in both C6 and MA-in cells. These observations demonstrate that the rat P450scc gene is transcript ionally regulated in glioma cells, but its regulation in glial cells i nvolves a DNA element different from those used in classic steroidogen ic tissues. The results further suggest that steroidogenic factor-1 is not involved in regulating neurosteroidogenesis.