Abundance of Early Embryonic Primordial Germ Cells Promotes Zebrafish Female Differentiation as Revealed by Lifetime Labeling of Germline.

Teleost sex differentiation largely depends on the number of undifferentiated germ cells. Here, we describe the generation and characterization of a novel transgenic zebrafish line, Tg(piwil1:egfp-UTRnanos3), which specifically labels the whole lifetime of germ cells, i.e., from primordial germ cells (PGCs) at shield stage to the oogonia and early stage of oocytes in the ovary and to the early stage of spermatogonia, spermatocyte, and spermatid in the testis. By using this transgenic line, we carefully observed the numbers of PGCs from early embryonic stage to juvenile stage and the differentiation process of ovary and testis. The numbers of PGCs became variable at as early as 1 day post-fertilization (dpf). Interestingly, the embryos with a high amount of PGCs mainly developed into females and the ones with a low amount of PGCs mainly developed into males. By using transient overexpression and transgenic induction of PGC-specific bucky ball (buc), we further proved that induction of abundant PGCs at embryonic stage promoted later ovary differentiation and female development. Taken together, we generate an ideal transgenic line Tg(piwil1:egfp-UTRnanos3) which can visualize zebrafish germline for a lifetime, and we have utilized this line to study germ cell development and gonad differentiation of teleost and to demonstrate that the increase of PGC number at embryonic stage promotes female differentiation.