Single-cell atlas reveals NLRP2 associated mitochondrial dysfunction in estrogen induced Macrobrachium nipponense sex reversal

Sex reversal is used in aquaculture breeding, but its cellular basis in crustaceans remains unclear. Here we combine estrogen induced feminization, single-cell RNA sequencing, histology and RNA interference to study gonadal development in the oriental river prawn (Macrobrachium nipponense). Feeding post-larval males with 17β-estradiol produced neo-females with ovary like gonads. Single-cell profiling of testes, ovaries and neo-female gonads resolved major oogenic and spermatogenic cell types and showed that neo-female gonads were enriched in early germ cells, whereas mature oocytes and spermatozoa were depleted. Cell type matched analyses identified oxidative phosphorylation as a consistently altered pathway. The gene MnNLRP2 showed primary spermatocyte specific and testis enriched expression. Knockdown of MnNLRP2 shifted sex ratios toward females in juveniles, altered sex hormone levels and caused spermatogenic arrest in adults. These results link MnNLRP2 to mitochondrial dysfunction during incomplete crustacean sex reversal. Single-cell analysis reveals incomplete estrogen induced sex reversal in Macrobrachium nipponense and identifies MnNLRP2 associated mitochondrial dysfunction in germ-cell development.