Isolation and characterization of the CYP17A1 gene and its processed pseudogene in Rana rugosa

CYP17A1 expression is up-regulated in the gonad in Rana (Glandirana) rugosa tadpoles treated with androgens to induce female-to-male sex-reversal. In this study, we isolated the CYP17A1 gene and its processed pseudogene from R. rugosa. The former was found to consist of 8 exons, and the latter a single-exon gene, designated CYP17A1P. The sequence of the promoter region of CYP17A1 differed from that of CYP17A1P. We found several consensus binding-sites for candidate transcription factors including androgen receptor (AR), Sox and FoxL2 in the CYP17A1 promoter region, but an AR-binding site was absent from CYP17A1P. When AR was over-expressed in Xenopus A6 cells, it did not increase CYP17A1 transcription in luciferase assays. CYP17A1 was strongly expressed in indifferent male gonads during sex determination and exclusively in testis, among eight adult tissues of R. rugosa. By contrast, CYP17A1P was expressed at very low, and similar levels in the adult tissues of both sexes. Fluorescent In-Situ Hybridization (FISH) analysis showed that CYP17A1P is localized to chromosome 4, while CYP17A1 is on chromosome 9. These results collectively suggest that CYP17A1, but not CYP17A1P is involved in male sex-determination in R. rugosa, and that androgens may not have a direct effect on the CYP17A1 transcription.