Sexually Divergent Role of Germ Cells in Mammalian Gonadogenesis

The formation of supporting “nurse” cells, Sertoli cells in the testis, and granulosa cells in the ovary is a defining event in gonadal sex differentiation. However, the contribution of germ cells to this process remains unclear. We generated NANOS3-null porcine fetuses to examine how germ cell loss influences gonadal development. Loss of NANOS3 did not impair primordial germ cell specification or migration but induced apoptosis and loss of pluripotency markers, resulting in complete germ cell depletion by embryonic day 35. Despite comparable gross morphology, NANOS3-null ovaries displayed disorganized cords, mesenchymal cell death, and arrested maturation of supporting cells, whereas testes maintained organized, non-lumenized cords with preserved architecture. Single-nucleus RNA sequencing and cell–cell communication analyses revealed sex-specific divergence in developmental trajectories. In the ovary, germ cells act as a critical source of BMP and WNT/RSPO signals that direct coelomic and supporting-cell differentiation; their absence led to abnormal expansion of immature supporting cells and failure to form pre-granulosa and thecal lineages. In contrast, testicular somatic populations expanded but retained identity through DHH, AMH, and PDGF/FGF signaling. Together, these findings uncover a germline-dependent program for ovarian differentiation and a germline-autonomous program for testicular development, highlighting intrinsic sexual divergence in mammalian gonadogenesis.