miR-27b-3p inhibits estrogen secretion of goose granulosa cells by targeting CYP1B1 through the AMPK signaling pathway

Although miR-27b-3p has been evi-denced to regulate the proliferation, apoptosis, and dif-ferentiation of a variety of mammalian cell types, its actions and mechanisms on ovarian cell steroidogenesis remains largely unknown in both mammalian and avian species. In this study, we aimed to determine the expres-sion profiles of miR-27b-3p in granulosa cell layers dur-ing goose ovarian follicle development and to reveal its actions on estrogen (E2) secretion of goose granulosa cells as well as the underlying regulatory mechanisms. It was observed that miR-27b-3p was ubiquitously expressed throughout follicle development but exhibited much higher levels in hierarchical-than in prehierarchi-cal follicles. In cultured granulosa cells from the fourth through second largest preovulatory (F4-F2) follicles of goose, up-and downregulation of miR-27b-3p by using its mimic and inhibitor significantly decreased and increased E2 secretion, respectively. Meanwhile, themRNA levels of STAR and CYP19A1 were significantly reduced while those of CYP11A1 and 3bHSD were ele-vated in the mimic-transfected granulosa cells. By com-parison, downregulation of miR-27b-3p enhanced the mRNA levels of STAR but had no significant effects on those of CYP19A1, CYP11A1, and 3bHSD. Results from bioinformatic prediction and luciferase reporter assay demonstrated that CYP1B1 was a downstream target of miR-27b-3p. Although the siRNA-mediated downregulation of CYP1B1 did not significantly change E2 secretion by goose granulosa cells, it reduced the mRNA levels of STAR and CYP19A1 as well as those of LKB1 and AMPKa, which are involved in the AMPK signaling pathway. Taken together, these data suggest that miR-27b-3p plays an inhibitory role in E2 secretion by goose F4-F2 granulosa cells, at least in part, by targeting CYP1B1 through the AMPK signal-ing pathway.