Cloning and Expression Analysis of figla During Ovarian Differentiation in Turbot (Scophthalmus maximus)

Abstract:

The transcription factor gene figla (factor in the germ line, alpha), a member of the basic helix-loop-helix (bHLH) family, has been extensively documented for its pivotal role in mammalian ovarian development and primordial follicle formation. However, studies of figla in teleosts are less detailed than those in mammals. Turbot (Scophthalmus maximus), an important aquaculture species in Europe and China, exhibits sexual dimorphism in growth and body size and possesses a female heterogametic sex determination system (ZW female and ZZ male). Therefore, it is crucial to understand the mechanisms underlying sex determination and differentiation. This study exclusively used female individuals as experimental subjects to elucidate the complete cDNA sequence of figla in turbot and its role during early ovarian development. The full-length cDNA sequence of figla was cloned using RT-PCR and rapid amplification of cDNA ends techniques. Tissue expression distribution and spatiotemporal expression patterns during early ovarian development were investigated using semi-quantitative RT-PCR, real-time fluorescence quantitative PCR, and in situ hybridization. The full length of the figla gene cDNA had 1,006 base pairs (bp), with an open reading frame ranging from 150 bp to 758 bp, encoding 202 amino acids with a conserved bHLH domain. In the turbot gonads, figla exhibited specific expression, showing higher levels in the ovaries than in the testes. The expression pattern of figla mRNA was detected throughout various stages of ovarian development, commencing at 25 dph and progressively increasing until reaching its peak at 90 dph. In situ hybridization experiments revealed the predominant localization of figla mRNA within the cytoplasmic region of oocytes. In conclusion, the distinct expression pattern of figla in male and female turbot, its localization within primary oocytes, and its expression patterns during early ovarian development suggest a crucial role for this gene in the normal development of oocytes and ovaries. Further research is required to investigate the regulatory mechanisms underlying oocyte development and gonadal differentiation. The study of figla function not only enhances our understanding of sex determination and differentiation mechanisms in turbot but also provides novel insights and methodologies for sex control and breeding applications in this species.