The piRNA protein Asz1 is essential for germ cell and gonad development in zebrafish and exhibits differential necessities in distinct types of RNP granules
Authors: Ahmad Adam , Bogoch Yoel , Shvaizer Gal , Elkouby Yaniv M. 
Published: July 2023

AbstractGerm cells are essential for fertility, embryogenesis, and reproduction. Germline development requires distinct types of RNA-protein (RNP) granules, including germ plasm in embryos, piRNA granules in gonadal germ cells, and the Balbiani body (Bb) in oocytes. However, the regulation of RNP assemblies in zebrafish germline development are still poorly understood. Asz1 is a piRNA protein in Drosophila and mice. Zebrafish Asz1 localizes to both piRNA and Bb granules, with yet unknown functions. Here, we hypothesized that Asz1 functions in RNP granule assemblies and germline development in zebrafish. We generatedasz1mutant fish to determine the roles of Asz1 in germ cell development. We show that Asz1 is dispensable for somatic development, but essential for germ cell and gonad development.asz1-/-fish developed exclusively as sterile males with severely underdeveloped testes that lacked germ cells, demonstrating that Asz1 is essential for spermatogenesis. Mechanistically, we provide evidence to conclude that zygotic Asz1 is not required for primordial germ cell specification or migration to the gonad, but is essential for germ cell survival during post-embryonic gonad development, likely by suppressing the expression of germline transposons. Increased transposon expression and morphologically mis-organized piRNA granules inasz1mutants, argues that zebrafish Asz1 functions in the piRNA pathway. We generatedasz1;tp53fish to partially rescue ovarian development, revealing underdeveloped mutant ovaries with defective oocytes, and that Asz1 is also essential for oogenesis. We further showed that in contrast with piRNA granules, Asz1 is dispensable for Bb granule formation, as shown by normal Bb localization of Buc anddazl. By uncovering Asz1 as an essential regulator of germ cell survival and gonadogenesis in zebrafish, and determining its differential necessity in distinct RNP granule types, our work advances our understanding of the developmental genetics of reproduction and fertility, as well as of RNP granule biology.Author SummaryGerm cells undergo a highly dynamic developmental program that begins in the early embryo and continues through juvenile and adult life. Identifying functional regulators and deciphering the developmental mechanisms of germ cells are critical for advancing our understanding of fertility and reproduction, as well as their associated diseases. Here, we identified Asz1 as an essential regulator of germ cell and gonad development in zebrafish. We demonstrate that zygotic Asz1 is dispensable for the specification and migration of primordial germ cells in the embryo, but is necessary for germ cell survival in the developing gonad, likely by protecting them from transposable elements. Upon loss of asz1, expression of germline transposons was induced, and piRNA granules were mis-organized, suggesting a conserved role for zebrafish Asz1 in the piRNA pathway. We show that Asz1 is required for both spermatogenesis and oogenesis. However, unlike RNA-protein (RNP) granules of the piRNA pathway, Asz1 was not required for RNP granules of the Balbiani body in differentiating oocytes, revealing its differential necessity in distinct types of germline RNP granules. In mice, Asz1 was shown to be essential for spermatogenesis but not oogenesis, but its functions in human gonads are unclear. Our work reports the functional requirements of Asz1 in both sexes in zebrafish, contributes to our knowledge of developmental reproduction biology, and sheds new light on the complexity of RNP granule assemblies.