Abstract
Ovarian development in crustaceans is characterized by rapid production of egg yolk protein in a process called vitellogenesis. In the present study, we investigated the involvement of a DEAD (Asp-Glu-Ala-Asp) box RNA helicase 20 (DDX20), forkhead transcription factor (FOXL)2 and fushi tarazu factor (FTZ-F)1 in the regulation of vitellogenesis. Based on ESTs from the testis and accessory gland of Eriocheir sinensis, we cloned the full-length cDNAs of foxl2 and fushitarazu factor 1 (ftz-f1), which include the conserved structural features of the forkhead family and nuclear receptor 5A (NR5A) family respectively. The expression of foxl2 mRNA surged at the mature stage of the ovary, when vtg mRNA swooped, suggesting that foxl2 negatively affects the vitellogenin (VTG) synthesis at this developmental stage. Etoposide (inducing germ cell apoptosis) treatment up-regulated FOXL2 and DDX20 at both the mRNA and the protein levels, primarily in the follicular cells as shown by immunofluorescence analysis. Furthermore, foxl2, ddx20 and ftz-f1 mRNA levels increased significantly with right-eyestalk ablation. Interactions between FOXL2 and DDX20 or FTZ-F1 were confirmed by co-immunoprecipitation and the forkhead domain of FOXL2 was identified as the specific structure interacting with FTZ-F1. In conclusion, FOXL2 down-regulates VTG expression by binding with DDX20 in regulation of follicular cell apoptosis and with FTZ-F1 to repress the synthesis of VTG at the mature stage. This report is the first to describe the molecular mechanism of VTG synthesis in E. sinensis and may shed new light on the regulation of cytochrome P450 enzyme by FOXL2 and FTZ-F1 in vitellogenesis.