Integrated chromatin accessibility and DNA methylation analysis to reveal the critical epigenetic modification and regulatory mechanism in gonadal differentiation of the sequentially hermaphroditic fish, Monopterus albus.

Background: Monopterus albus is a hermaphroditic and economically farmed fish that undergoes sex reversal from ovary to testis via ovotestis during gonadal development. The epigenetic changes that are associated with gonadal development in this species remain unclear. Methods: We produced DNA methylome, transcriptome, and chromatin accessibility maps of the key stages of gonad development: ovary, ovotestis, and testis. The expression of the key candidate genes was detected using qRT-PCR and in situ hybridization and the methylation levels were analysed using bisulphite sequencing PCR. Promoter activity and regulation were assessed using dual-luciferase reporter assays. Results: Gonadal development exhibits highly dynamic transcriptomic, DNA methylation, and chromatin accessibility changes. We found that DNA methylation status, especially of the transcription start site, was significantly negatively correlated with gene expression while chromatin accessibility exhibited no correlation with gene expression during gonadal development. The epigenetic signatures revealed many novel regulatory elements and genes involved in sex reversal, which were validated. DNA methylation detection and site mutation of plastin-2 promoter, as a candidate gene, revealed that DNA methylation could impact the binding of transcription factor dmrt1 and foxl2 through methylation and demethylation to regulate plastin-2 expression during gonadal development. Conclusions: These data provide novel insights into epigenetic modification and help elucidate the potential molecular mechanism by which dynamic modification of DNA methylation plays a crucial role in gonadal development. Highlights: DNA methylation modification plays a more crucial role than chromatin accessibility in gonad development. DNA methylation in the transcription start site plays a key role in gonad development. DNA methylation of the plastin-2 promoter inhibited gene expression. Plastin-2 is positively regulated by dmrt1 while negatively regulated by foxl2. [ABSTRACT FROM AUTHOR]