Dynamic miRNA landscapes unlock sexual plasticity in protogynous orange-spotted grouper under MT-implantation

Abstract Sex change, a manifestation of sexual plasticity, is widespread among fish and can occur spontaneously or in response to environmental stimuli. Despite its prevalence, the molecular regulatory mechanisms underlying sex change, particularly those involving miRNAs, remain largely unresolved. Orange-spotted grouper (Epinephelus coioides), a hermaphroditic species exhibiting natural female-to-male sex reversal, serves as an excellent model for investigating these processes. In this study, we established a reversible sex change model in E. coioides by inducing precocious female-to-male transition through 17α-methyltestosterone implantation, followed by spontaneous male-to-female reversion. Comprehensive transcriptome analyses were conducted during both transitions to identify differentially expressed miRNAs and their putative target mRNAs. Regulatory networks were constructed, revealing dynamic miRNA-mRNA interactions associated with sex differentiation. Key miRNAs were found to be inversely correlated with sex-related gene expression, suggesting their involvement in the regulation of sex reversal. Functional validation further supported the existence of targeted negative regulatory relationships between specific miRNAs and sex-related genes. Collectively, these findings demonstrate that miRNAs play essential roles in orchestrating bidirectional sex change in orange-spotted grouper, providing new insights into the molecular basis of sex determination and differentiation in teleosts.