ABSTRACT: Female endocrinological symptoms, such as premature ovarian inefficiency (POI) are caused by diminished ovarian reserve and chemotherapy. The etiology of POI remains unknown, but this can lead to infertility. This has accelerated the search for master regulator genes or other molecules that contribute as enhancers or silencers. The impact of regulatory microRNAs (miRNAs) on POI has gained attention; however, their regulatory function in this condition is not well known. RNA sequencing was performed at four stages, 2-(2 W), 6-(6 W), 15-(15 W), and 20-(20 W) weeks, on ovarian tissue samples and 5058 differentially expressed genes (DEGs) were identified. Gene expression and enrichment were analyzed based on the gene ontology and KEGG databases, and their association with other proteins was assessed using the STRING database. Gene set enrichment analysis was performed to identify the key target genes. The DEGs were most highly enriched in 6 W and 15 W groups. Figla, GDF9, Nobox, and Pou51 were significantly in-creased at 2 W compared with levels at 6 W and 20 W, whereas the expression of Foxo1, Inha, and Taf4b was significantly de-creased at 20 W. Ccnd2 and Igf1 expression was maintained at similar levels in each stage. In total, 27 genes were upregulated and 26 genes interacted with miRNAs; moreover, stage-specific upregulated and downregulated interactions were demonstrated. Increased and decreased miRNAs were identified at each stage in the ovaries. The constitutively expressed genes, Ccnd2 and Igf1, were identified as the major targets of many miRNAs (p < 0.05), and Fshr and Foxo3 interacted with miRNAs, namely mmu-miR-670-3p and mmu-miR-153-3p. miR-26a-5p interacted with Piwil2, and its target genes were downregulated in the 20 W mouse ovary. In this study, we aimed to identify key miRNAs and their target genes encompassing the reproductive span of mouse ovaries using mRNA and miRNA sequencing. These results indicated that gene sets are regulated in the reproductive stage-specific manner via interaction with miRNAs. Furthermore, consistent expression of Ccnd2 and Igf1 is considered crucial for the ovarian reserve and is regulated by many interactive miRNAs.
