Loss of growth differentiation factor 9 causes an arrest of early folliculogenesis in zebrafish–A novel insight into its action mechanism.

Growth differentiation factor 9 (GDF9) was the first oocyte-specific growth factor identified; however, most information about GDF9 functions comes from studies in the mouse model. In this study, we created a mutant for Gdf9 gene (gdf9-/-) in zebrafish using TALEN approach. The loss of Gdf9 caused a complete arrest of follicle development at primary growth (PG) stage. These follicles eventually degenerated, and all mutant females gradually changed to males through sex reversal, which could be prevented by mutation of the male-promoting gene dmrt1. Interestingly, the phenotypes of gdf9-/- could be rescued by simultaneous mutation of inhibin α (inha-/-) but not estradiol treatment, suggesting a potential role for the activin-inhibin system or its signaling pathway in Gdf9 actions. In gdf9 null follicles, the expression of activin βAa (inhbaa), but not βAb (inhbab) and βB (inhbb), decreased dramatically; however, its expression rebounded in the double mutant (gdf9-/-;inha-/-). These results indicate clearly that the activation of PG follicles to enter the secondary growth (SG) requires intrinsic factors from the oocyte, such as Gdf9, which in turn works on the neighboring follicle cells to trigger follicle activation, probably involving activins. In addition, our data also support the view that estrogens are not involved in follicle activation as recently reported. Author summary: Follicles are the basic structural and functional units of the ovary. Each follicle consists of an oocyte and surrounding somatic follicle cells. The growth and maturation of follicles or folliculogenesis is controlled by a variety of hormones and local factors. It has been known for decades that the oocyte in the follicle plays an active role in controlling folliculogenesis by releasing various regulatory factors, among which the growth differentiation factor 9 (GDF9) is the best characterized one. In this study, we used gene editing method to delete the gene for GDF9 (gdf9) in the zebrafish. We discovered that without gdf9 gene (mutant), the follicles could not develop to advanced stages and the females were therefore infertile. Interestingly, when inhibin (a hormone from the ovary) was lost simultaneously, the follicles in the gdf9 mutant resumed growth and development. We hypothesized that the resumption of follicle growth in the double mutant of gdf9 and inha was likely due to increased activity of activin. [ABSTRACT FROM AUTHOR]