CYTOCHROME-P450 17-ALPHA-HYDROXYLASE GENE-EXPRESSION IN DIFFERENTIATING RAT TROPHOBLAST CELLS

Trophoblast giant cells of the rat placenta express cytochrome P450 17 alpha-hydroxylase (P450c17) and synthesize androgens. The purpose of this study was to investigate androgen production and expression of P4 50c17 in the Rcho-1 trophoblast cell line. These cells are capable of differentiating along the trophoblast giant cell lineage. Androstenedi one production increased approximately 70-fold as Rcho-1 trophoblast c ells progressed fi-om the proliferation to the differentiation state. P450c17 enzyme activity and mRNA also showed significant increases ass ociated with trophoblast giant cell differentiation. To study the tran scriptional regulation of the P450c17 gene, the activities of a series of P450c17 promoter-luciferase reporter constructs were evaluated fol lowing transient transfection into Rcho-1 trophoblast cells. A DNA reg ion located -98 bp upstream of the P450c17 gene transcriptional start site was the shortest promoter DNA construct consistently possessing a ctivity in Rcho-1 trophoblast cells. Activities of longer constructs ( -156 to -1560 bp) in this population of cells were significantly great er than the -98 bp promoter-reporter construct. The -476 bp P450c17 co nstruct showed maximal promoter activity in transiently transfected Rc ho-1 trophoblast cells and was developmentally activated in stably tra nsfected Rcho-1 trophoblast cells. Activation of the cyclic AMP/protei n kinase A pathway did not significantly affect P450c17 promoter activ ity in Rcho-1 trophoblast cells, in contrast to its effects in mouse M A-10 Leydig cells. In summary, Rcho-1 trophoblast cells are capable of endocrine differentiation and are a useful in vitro system for studyi ng the regulation of trophoblast androgen production and P450c17 gene expression.