Characterization of Forkhead Box Transcription Factor (foxl) in Sex Differentiation of Chinese Tongue Sole (Cynoglossus semilaevis)

Chinese tongue sole (Cynoglossus semilaevis) is an important mariculture product in northern China, exhibiting significant sexual dimorphism: females grow 2–4 times faster than males and ultimately attain much greater body weights. As a well-known transcription factor crucial for regulating sex differentiation, foxl2 has been characterized in various mammals. Herein, we identified and characterized three foxl genes, foxl1, foxl2a and foxl2l. Three foxl genes exhibited a gonad-biased expression pattern, where foxl2a showed higher expression in ovary than in testis, while foxl1 and foxl2l exhibited higher expression in testis. All foxl genes were detected in testes and ovaries by ISH; foxl1/foxl2l were expressed in oocytes and sperm, and foxl2a in granulosa cells and sperm. Overexpression of foxl in testicular cells led to KEGG enrichment in DNA repair, MAPK, FOXO and progesterone-mediated oocyte maturation pathways. In tongue sole testicular cell line, knockdown of foxl1 and foxl2l resulted in upregulation of multiple male-related genes. In contrast, knockdown of foxl2a led to decreased expression of aromatase genes and increased expression of ctnnb1, indicating that foxl2a is more closely associated with female differentiation and maintenance. Our study investigated the functions of the foxl gene family in teleosts and offers valuable insights into their role in sex differentiation and gonadal maintenance in teleost fish.