Recently we reported the isolation of two types of cytochrome P450cl7s (steroid 17 alpha-hydroxylase/C17, 20 lyase) encoded by two different genes, from genomes of teleost fish. In this study, we characterized the expression profile, enzymatic activity and transcriptional regulation of P450c174 and -II in the medaka ovary. Similar to tilapia, medaka P450c17-I possessed both hydroxylase and lyase activities, while P450c17-II possessed only the hydroxylase activity. In situ hybridization and gene expression profiles during 48 h prior to spawning indicated that P450c17-is responsible for the production of estradiol-17 beta during oocyte growth, while P450c17-II for the production of 17 alpha, 20 beta-dihydroxy-4-pregnen-3-one during oocyte maturation and cortisol production in the head kidney. Luciferase assays and expression profiles of transcription factors as revealed by real time PCR suggested that P450c17-I and -II expression are tightly controlled by Ad4BP/SF-1, Lrb-1, Fox12, and Dax1 during the 48 h prior to spawning. (c) 2007 Elsevier Inc. All rights reserved.