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. 2025 Jul 31;26(1):707.
doi: 10.1186/s12864-025-11408-0.

Integrated analysis of microRNA and mRNA interactions regulating fecundity in the ovaries of two distinct sheep breeds

Salsabeel Yousuf  1 , Waqar Afzal Malik  2 , Hui Feng  1 , Tianyi Liu  1 , Lingli Xie  1 , Xiangyang Miao  3
Affiliations

Integrated analysis of microRNA and mRNA interactions regulating fecundity in the ovaries of two distinct sheep breeds

Salsabeel Yousuf et al. BMC Genomics. .

Abstract

Sheep production is vital to the global agricultural economy, particularly for meat and wool. However, fertility varies significantly among breeds, influenced by genetics, nutrition, and environment. MicroRNAs (miRNAs) are crucial post-transcriptional regulators of gene expression, binding to target messenger RNAs (mRNAs) to inhibit translation or trigger degradation. Despite their importance, little is known about ovary-specific miRNAs and their target genes in high and low fecundity sheep breeds. Here, we investigated miRNA expression profiles in ovarian tissues from Small Tail Han (high fecundity) and Dolang sheep (low fecundity) using miRNA sequencing. The analysis identified 51 significantly differentially expressed miRNAs (DEmiRNAs), with only nine mapping to the reference genome (including oar-miR-103, oar-miR-16b). We predicted 88,699, 176,629, and 240,753 target mRNAs for these DEmiRNAs from sheep, goat, and cow, respectively. Functional enrichment analysis revealed significant biological processes and signaling pathways potentially associated with fecundity. Subsequently, we constructed a DEmiRNA-mRNA regulatory network, highlighting candidate miRNA-mRNA pairs like chi-miR-130a-3p-WNT2B, oar-miR-103-FSHR, and oar-miR-16b-AMH. These findings provide insights into the potential regulators of sheep fecundity. We further validated these candidate miRNA-mRNA interactions using qRT-PCR to explore their roles in regulating sheep fecundity. Overall, this study utilized miRNA sequencing to analyze ovarian miRNA expression in sheep and identified potential miRNA-mRNA targets associated with fecundity. This lays the foundation for future research on the molecular mechanisms underlying sheep ovarian physiology and improving reproductive traits.

Keywords: Fecundity; Ovary; Ovine tissue; Sheep; Transcriptome; miRNA.

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Conflict of interest statement

Declarations. Ethics approval and consent to participate: All the procedures involving animals were approved by the animal care and use committee at the Institute of Animal Sciences, Chinese Academy of Agricultural Sciences (NO. IAS2019-82), where the study was conducted. All the experiments were performed in accordance with ARRIVE guidelines and regulations set by the Ministry of Agriculture of the People’s Republic of China. Consent for publication: Not applicable. Competing interests: The authors declare no competing interests.

Figures

Fig. 1
Fig. 1
Structural length characteristics of small RNAs. A-B-C-D-E–F Nucleotide composition profiles of mature small RNAs in X_LC and D_LC groups. G Line plot reveal the miRNA length distribution in each sample
Fig. 2
Fig. 2
Sample wise annotation summary of small RNAs in two sheep breeds. A-B Total small RNA annotation summary in D_LC and X_LC. C-D Annotation summary for unique small RNAs in D_LC and X_LC
Fig. 3
Fig. 3
miRNA expression analysis. A Boxplot log (TPM) sample distribution. B miRNA expression density distribution of each sample in two group X_LC and D_LC. C Sample-to-sample correlation coefficient heatmap. D Correlation between the samples of miRNA expression level. E Sample-to-sample cluster plot of each sample
Fig. 4
Fig. 4
Differentially expressed miRNAs and their expression. A Volcano map of differentially expressed miRNAs. The differences resulting from the comparison are reflected in the volcano map, with non-differential miRNAs in gray, significant upregulated miRNAs in red, significantly differential miRNAs in green, and log in the X axis2FoldChange with a Y axis orientation of -log10pValue or -log10qValue. B Differentially expressed miRNAs heat expression
Fig. 5
Fig. 5
Functional enrichment analysis of DEmiRNA target genes. A GO analysis of DEmiRNAs target genes from ovarian tissue. B KEGG Pathway enrichment analysis for miRNA target genes
Fig. 6
Fig. 6
Co-expression network of DEGs and DEmiRNAs. A Differentially Expressed miRNAs and their target mRNAs from Ovarian Tissue in X_LC vs. D_LC. B Subnetwork of Key DEmiRNAs and mRNAs
Fig. 7
Fig. 7
miRNA-mRNA Regulatory Network in X_LC vs. D_LC. The triangle (red) node represents a differentially expressed miRNA (DEmiRNA), circle (yellow) node represents a messenger RNA (mRNA), and rectangle (green) node represents a biological pathway
Fig. 8
Fig. 8
Protein protein interaction (PPI) network of differentially expressed mRNAs in Ovine tissue of Small Tail Han and Dolang sheep
Fig. 9
Fig. 9
Validation of DE miRNAs, DE mRNAs, and their expression patterns. The qRT-PCR verification result of shorlisted (A-B) differentially expressed miRNAs and their corresponding target genes (mRNAs) and fold change expression of (C-D) shortlisted DEGs and DEmiRNAs in Ovine tissue of Small Tail Han and Dolang sheep (X_LC -vs-D_LC). Red indicates upregulation; green indicates downregulation
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