The gonads and adrenal cortex are the tissues producing steroid hormones such as sex steroids, mineralocorticoid and glucocorticoid. These steroids are synthesized by consecutive enzymatic reactions by P450s and hydroxysteroid dehydrogenases. Studies of the tissue specific expression of these gene involved in steroidogenesis identified an orphan nuclear receptor Adrenal-4-Binding Protein (Ad4BP, NR5A1), also known as Steroidogenic Factor-1 (SF-1). In addition to these steroidogenic tissuses, Ad4BP/SF-1 was shown to be expressed in the pituitary gonadtrophe and ventromedial hypothalamic nucleus (VMH), both of which are closely related to reproductive activity. The gene disruption study revealed that Ad4BP/SF-1 is required for the development and functions of these tissues. Considering that Ad4BP/SF-1 gene expression at the tissues is crucial for the tissue development, the mechanisms underlying the tissue-specific expression should be essential for understanding the whole process of the tissue development. Thus, in vivo transgenic mouse assay was undertaken to ask which part of the Ad4BP/SF-1 gene locus is implicated in the tissue-specific expression. Our trial so far has successfully identified the enhancer for the fetal adrenal cortex, pituitary gonadtrophe, VMH, and recently fetal male gonad. Structural and functional analyses of the enhancers gradually revealed the upstream factors/signals to induce the expression of Ad4BP/SF-1.