Fuchou Tang

BACKGROUND: MicroRNAs (miRNAs) are critical regulators of transcriptional and post-transcriptional gene silencing, which are involved in multiple developmental processes in many organisms. Apart from miRNAs, mouse germ cells express another type of small RNA, piwi-interacting RNAs (piRNAs). Although it has been clear that piRNAs play a role in repression of retrotransposons during spermatogenesis, the function of miRNA in mouse germ cells has been unclear. METHODOLOGY/PRINCIPAL FINDINGS: In this study, we first revealed the expression pattern of miRNAs by using a real-time PCR-based 220-plex miRNA expression profiling method. During development of germ cells, miR-17-92 cluster, which is thought to promote cell cycling, and the ES cell-specific cluster encoding miR-290 to -295 (miR-290-295 cluster) were highly expressed in primordial germ cells (PGCs) and spermatogonia. A set of miRNAs was developmentally regulated. We next analysed function of miRNA biogenesis in germ cell development by using conditional Dicer-knockout mice in which Dicer gene was deleted specifically in the germ cells. Dicer-deleted PGCs and spermatogonia exhibited poor proliferation. Retrotransposon activity was unexpectedly suppressed in Dicer-deleted PGCs, but not affected in the spermatogonia. In Dicer-deleted testis, spermatogenesis was retarded at an early stage when proliferation and/or early differentiation. Additionally, we analysed spermatogenesis in conditional Argonaute2-deficient mice. In contrast to Dicer-deficient testis, spermatogenesis in Argonaute2-deficient testis was indistinguishable from that in wild type. CONCLUSION/SIGNIFICANCE: These results illustrate that miRNAs are important for the proliferation of PGCs and spermatogonia, but dispensable for the repression of retrotransposons in developing germ cells. Consistently, miRNAs promoting cell cycling are highly expressed in PGCs and spermatogonia. Furthermore, based on normal spermatogenesis in Argonaute2-deficient testis, the critical function of Dicer in spermatogenesis is independent of Argonaute2.

BACKGROUND: Argonaute2 protein (Ago2) is a key component of RNA-induced gene silencing complex, which is crucial for microRNA-mediated repression of target genes. The function of Ago2 in the mouse oocyte and early embryonic development is less well characterized but it is likely to have an important role in regulating maternally inherited mRNA. We have examined the role of Ago2 by conditional deletion of the gene in developing oocytes. RESULTS: Ago2 was deleted specifically in the growing oocytes. Although the Ago2-deficient oocytes are able to develop to mature oocytes, they have abnormal spindles and chromosomes that are unable to cluster together properly. This phenotype is very similar to the phenotype of Dicer-deficient oocytes. We examined the microRNA expression profile in the Ago2-deficient oocyte and found that the expression of most microRNAs was reduced by more than 80%. To determine the downstream genes that are regulated by Ago2, we used microarray analysis on Ago2-deficient oocytes and found that 512 genes were upregulated and 1,073 genes were downregulated (FC > 2, P < 0.05). CONCLUSION: Our study shows that Ago2 has a key function in the mouse oocyte through global regulation of microRNA stability, and through this mechanism it affects gene expression in developing oocytes.