The 5'-flanking regions of genes for three mouse adrenal steroid hydro
xylases were analyzed for their ability to direct adrenal cortex-speci
fic beta-galactosidase (beta-gal) reporter expression both in cell cul
ture and transgenic mice. The 5'-flanking regions chosen were from the
genes for steroid 21-hydroxylase (21-OHase), expressed throughout the
adrenal cortex and mediating both glucocorticoid and mineralocorticoi
d synthesis, and aldosterone synthetase (AS) and steroid 11 beta-hydro
xylase (11 beta-OHase), which catalyze respectively the terminal steps
of mineralocorticoid synthesis in the zona glomerulosa and glucocorti
coid synthesis in the zona fasciculata/reticularis. While 5.0 kb of 11
beta-OHase gene 5'-flanking region and 5.4 kb of the AS gene 5'-flank
ing region mediated respectively moderate and low levels of beta-gal r
eporter expression in YI adrenocortical tumor cells, neither of these
5'-flanking regions was able to direct reporter expression to the appr
opriate adrenocortical zone of transgenic mice. This suggests that add
itional regulatory elements, lying outside these 5'-flanking regions,
are required for 11 beta-OHase and AS gene expression in the intact mo
use. In contrast, 6.4 kb of the mouse 21-OHase A gene 5' flanking regi
on was able to direct specific beta-galactosidase reporter expression,
in both Y1 cells and transgenic mice, indicating that elements direct
ing adrenal cortex-specific gene expression in vivo are located not mo
re than 6.4 kb 5' of the 21-OHase gene transcription start site.