Yu_2025_Anim.Reprod.Sci_286_108092

This study aimed to investigate the effects of microRNA-150 (miR-150) on progesterone (P4) synthesis and apoptosis in goat luteal cells. Corpus luteum (CL) tissues were collected from non-pregnant goats. qPCR and in situ hybridization showed that miR-150 was highly expressed in the early CL but decreased in mid and late CL, and miR-150 was positively localized in luteal cells. Luteal cells from mid-cycle CL were then transfected with Agomir-150 (mimics) or Antagomir-150 (inhibitor). Immunoassays and qPCR results showed that Agomir-150 suppressed P4 levels and down-regulated the mRNA and protein expressions of hormonesensitive lipase (HSL), phosphorylated HSL (p-HSL), steroidogenic acute regulatory protein (StAR), and 3betahydroxysteroid dehydrogenase (HSD3B), whereas Antagomir-150 had the opposite effects. In addition, flow cytometry, qPCR, and Western blot results showed that Agomir-150 had no significant impact on cell apoptosis, while Antagomir-150 promoted apoptosis, up-regulated the mRNA and protein expressions of BCL2-associated X protein (BAX), Fas cell surface death receptor (FAS), tumor necrosis factor receptor 1 (TNFR1), Caspase-3, cleaved Caspase-3, and down-regulated B-cell lymphoma 2 (BCL-2) expression. Bioinformatic analysis identified nuclear receptor subfamily 4, group A, member 1 (NR4A1) as a potential miR-150 target. Dual-luciferase assays confirmed that miR-150 directly binds to the NR4A1-3'UTR and represses its expression, which was further validated by qPCR and Western blot. A rescue experiment revealed that NR4A1 knockdown partially reversed the Antagomir-150-induced up-regulation of P4 synthesis and apoptosis. Overall, this study demonstrates that miR-150 contributes to P4 synthesis and apoptosis in goat luteal cells by targeting NR4A1.