Fig. 13

Combined effect of triploidy and EE2 administration in hormonal and paracrine regulation in zebrafish gonads. (A). Differentially expressed genes in EE-treated triploid (3n + EE2) compared to diploid control (2n) zebrafish gonads: follicle-stimulating hormone receptor (fshr), antimüllerian hormone (amh), naked cuticle homolog 3-like (nkd3l), inhibin a (inha), Dishevelled-binding antagonist of beta-catenin 1 (dac1), dehydrogenase/reductase member 3a (dhrs3a), microRNA 202 (mir202), Kelch-like protein 10 (klhl10), deiodinase 1 (dio1) and deiodinase 3 (dio3). Asterisks denote significant differences between treatments. Normalized counts and statistical significance were obtained using DESeq2 median of ratios normalization and negative binomial differential expression modeling. (B). Gene regulatory network in the gonads of 3n + EE2 compared to 2n gonads at 120 days post-fertilization. EE2-treated triploids showed downregulation of fshr, amh, and inha, which favours spermatogonial proliferation. Suppression of nkd3l and dact1 indicated sustained β-catenin activation, reinforcing the male trajectory. dio1 and dio3 were downregulated, while notch1 and the female-biased mir202 were also reduced, reflecting lack of ovarian inputs. In the Retinoic Acid (RA) pathway, dhrs3a was downregulated and cyp26b1 unchanged, suggesting constrained RA signaling. Finally, reduced klhl10 expression aligned with impaired spermatogenesis observed in triploid testes. Upward/downward arrows indicate up- or downregulation of the adjacent gene; colored solid lines represent upregulation, colored dashed lines downregulation, and grey lines genes not differentially expressed but active in baseline gonadal physiology. Line endings denote stimulatory or inhibitory interactions. Abbreviations: spermatogonia An undifferentiated (SpgAund), spermatogonia A differentiated (SpgAdiff), spermatogonia B (SpgB), spermatocyte (Spc), spermatid (Spt), spermatozoa (Stz). Created in BioRender. MARTINEZ, A. (2025) https://BioRender.com/q3sjavl.