Abstract
Mammalian folliculogenesis is a complex process in which primordial follicles develop into pre-ovulatory follicles, followed by ovulation to release mature oocytes. In this study, we explored the role of miR-144 in ovulation. miR-144 was one of the differentially expressed microRNAs, which showed 5.59-fold changes, in pre-ovulatory ovarian follicles between Large White and Chinese Taihu sows detected by Solexa deep sequencing. We demonstrated that overexpression of miR-144 significantly decreased the luciferase reporter activity under the control of the cyclooxygenase-2 (COX-2) or mothers against decapentaplegic homologue 4 (Smad4) 3'-untranslated region (3'-UTR) and suppressed COX-2 and Smad4 expression. In contrast, a miR-144 inhibitor increased COX-2 and Smad4 expression in mouse granulosa cells (mGCs). Meanwhile, Smad4 upregulated COX-2 expression, but this effect was abolished when the mGCs were treated with the transforming growth factor beta signalling pathway inhibitor SB431542. Moreover, luciferase reporter, chromatin immunoprecipitation and electrophoretic mobility shift assay results showed that the transcription factor CP2 upregulated miR-144 expression, which partially contributed to the suppression of COX-2 in mGCs. Both CP2 and miR-144 alter prostaglandin E2 (PGE2) production by regulating COX-2 expression. In addition, miR-144 regulated mGC apoptosis and affected follicular atresia, but these activities did not appear to be through COX-2 and Smad4. Taken together, we revealed an important CP2/miR-144/COX-2/PGE2/ovulation pathway in mGCs.
Highlights
Chinese Taihu (CT) pigs farrow three to five more piglets per litter than American or European pig breeds, which has been intensely studied.[1]
Two independent small RNA libraries from pre-ovulatory ovarian follicles of CT and Large White (LW) sows were sequenced with the high-throughput Illumina Solexa system (Beijing Genomics Institute, Shenzhen, China)
Three hundred ninety miRNAs were differentially expressed between the two breeds, with 124 miRNAs upregulated and 266 miRNAs downregulated in CT sows (Po0.05, |Fold change|41) (Supplementary Table S1)
Summary
Chinese Taihu (CT) pigs farrow three to five more piglets per litter than American or European pig breeds, which has been intensely studied.[1]. MicroRNAs (miRNAs) are short, non-coding RNAs, 19–22 nucleotides in length, which act by targeting partially complementary sequences within the mRNA 3ˈ-untranslated region (3ˈ-UTR), leading to functional repression of target transcripts. These molecules are involved in multiple biological processes.[16] Many studies have shown that miRNAs have important roles in follicular development, follicular atresia, hormone modulation, cell apoptosis and cell proliferation.[17,18,19,20]. Received 02.8.16; revised 28.11.16; accepted 21.12.16; Edited by G Calin miR-144 regulates ovulation via targeting COX-2 J Zhou et al
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