Mutation of foxl2 or cyp19a1a Results in Female to Male Sex Reversal in XX Nile Tilapia

Abstract It is well accepted that Forkhead box protein L2 (Foxl2) and aromatase (Cyp19a1; the enzyme responsible for estrogen synthesis) are critical for ovarian development in vertebrates. Knockouts of Foxl2 and Cyp19a1 in goat, mouse, and zebrafish have revealed similar but not identical functions across species. Functional analyses of these two genes in other animals are needed to elucidate their conserved roles in vertebrate sexual development. In this study, we established foxl2 and cyp19a1a mutant lines in Nile tilapia. Both foxl2−/− and cyp19a1a−/− XX fish displayed female-to-male sex reversal. Sf1, Dmrt1, and Gsdf were upregulated in the foxl2−/− and the cyp19a1a−/− XX gonads. Downregulation of Cyp19a1a and serum estradiol-17β level, and upregulation of Cyp11b2 and serum 11-ketotestosterone level were observed in foxl2−/− XX fish. The mutant phenotype of foxl2−/− XX individuals could be rescued by 17β-estradiol treatment from 5 to 30 days after hatching (dah). Upregulation of Star1, the enzyme involved in androgen production in tilapia, was also observed in the foxl2−/− XX gonad at 30 and 90 dah. In vitro promoter analyses consistently demonstrated that Foxl2 could suppress the transcription of star1 in a dose-dependent manner. In addition, compared with the control XX gonad, fewer germ cells were detected in the foxl2−/− XX, cyp19a1a−/− XX, and control XY gonads 10 dah. These results demonstrate that Foxl2 promotes ovarian development by upregulating Cyp19a1a expression and repressing male pathway gene expression. These results extend the study of Foxl2 and Cyp19a1a loss of function to a commercially important fish species. Loss-of-function study showed that Foxl2 promoted ovarian development by upregulating Cyp19a1a expression, increasing germ cell number, and repressing male pathway gene expression in Nile tilapia.