Expression dynamics of sex-biased development- and immune-related genes underpin early gonadal differentiation in grass carp

Immune-reproductive interactions play important regulatory roles in vertebrate gonadal development. However, the contribution of immune factors to early gonadal development in teleost fish remains poorly understood. To address this, we conducted transcriptome profiling of male and female gonads in grass carp, a representative teleost species, across six developmental time points, focusing on the sex-biased development and immune-related genes. Our results pinpointed sex determination in grass carp at around 30 days post-fertilization (dpf), followed by ovarian and testicular morphological differentiation at approximately 60 and 120 dpf, respectively. Time-series analysis of differentially expressed genes (DEGs) characterized the transcriptional dynamics during the transition from sex determination to differentiation, identifying key female- (e.g., cyp19a1a, foxl2a/b) and male-biased (e.g., dmrt1, amh) development-related genes. Interestingly, extensive sexual dimorphism was also observed in the expression of immune-related gene during early gonadal development. Specifically, females exhibited activation of early macrophage-related signaling (Stage 1) and late microenvironmental regulation (Stage 4), whereas males showed a delayed macrophage-related signaling (Stage 2) followed by a strong induction of type I interferon (IFN) signaling (Stage 3). Protein and protein interaction (PPI) network analysis further highlighted a divergence in regulatory architecture, demonstrating that ovarian hub genes were exclusively sex development-related (e.g., ccna1, fancd2) while testicular networks integrated both sex development and immune-related regulators (e.g., amh, isg15). Collectively, our findings characterize the dynamic and sex-specific coordination of developmental and immune programs, providing new insights into gonadal development in teleost.