Scientia Agricultura Sinica

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miR-221-3p regulates ovarian granulosa cells apoptosis by targeting BCL2L11

LIU YuFang1,2, CHEN YuLin1,2, ZHOU ZuYang1,2, CHU MingXing1*   

  1. 1Key Laboratory of Animal Genetics, Breeding and Reproduction of Ministry of Agriculture and Rural Affairs, Institute of Animal Science, Chinese Academy of Agricultural Sciences, Beijing 100193; 2 College of Life Science and Food Engineering, Hebei University of Engineering, Handan 056001
  • Published:2021-11-30

Abstract: 【ObjectiveBCL2L11 promotes apoptosis in mammals and is involved in the development of tissues and organs related to reproductive traits and in disease treatment. To explore the effect of miR-221-3p on the regulation of granulosa cell apoptosis in Small-Tail Han sheep by targeting BCL2L11, so as to provide evidence for further study of the regulation of BCL2L11 in granulosa cell apoptosis and atresia of follicles. MethodBased on the whole transcriptome sequencing analysis of the ovarian tissue of the previous study in our group, the differentially expressed gene BCL2L11 and its regulatory element miR-221-3p were obtained in this study. Analysis of the expression of BCL2L11 in different tissues of the Small-Han Tail sheep using semi-quantitative and tissue fluorescence quantification (RT-qPCR). The expression of BCL2L11 and miRNA-221-3p was identified in the ovarian tissues of the Small-Tail Han sheep in the follicular and luteal phase by RT-qPCR. The miR-221-3p mimic, BCL2L11 wild-type and BCL2L11 mutant-type were co-transfected in HEK293T cells with negative control, the dual luciferase reporter gene detection system was used to determine the targeting relationship between miR-221-3p and BCL2L11. The miR-221-3p mimic and negative control were transfected into ovarian granulosa cells to achieve the overexpression of miR-221-3p. RT-qPCR was used to detect the effect of miR-221-3p on the expression levels of BCL2L11 and the marker genes of the apoptosis of ovarian granulosa cell gene XIAP and Fas at the mRNA level. At the same time, the changes in proliferation of granulocytes in the miR-221-3p overexpression and negative control groups were also analysed using the EdU assay. ResultsThe results showed that the expression of BCL2L11 in ovarian tissue was the highest, followed by spleen and lung tissues. RT-qPCR results showed that the expression of miR-221-3p and BCL2L11 was significantly different in the ovarian tissues of Small Tail Han sheep between follicular and luteal phases, the expression of miR-221-3p was higher in follicular than that in luteal phase ovaries, whereas BCL2L11 was less expressed in follicular than that in luteal phase ovaries, which showed the phenomenon of a negative regulation. Dual luciferase reporter analysis showed that overexpression of miR-221-3p significantly inhibited the activity of BCL2L11 3’UTR vector (P<0.05). The overexpression of miR-221-3p significantly reduced the mRNA level expression of target gene BCL2L11, while follicular granulosa cell apoptosis expression of marker genes XIAP and Fas were also significantly reduced (P<0.05). Analysis of the EdU assay showed that the proliferation rate of granulosa cells overexpressing miR-221-3p was 18.9%, which was significantly higher than that of the negative control group at 10.43% (P<0.01). 【ConclusionBCL2L11 and miR-221-3p are important genes and regulatory elements that regulate ovarian development in sheep. BCL2L11 is one of the target genes of miR-221-3p, and overexpression of miR-221-3p could inhibit granulosa cell apoptosis by target BCL2L11.



Key words: miR-221-3p, Small-Tail Han Sheep, granulosa cells apoptosis, BCL2L11

[1] LIU YuFang,CHEN YuLin,ZHOU ZuYang,CHU MingXing. miR-221-3p Regulates Ovarian Granulosa Cells Apoptosis by Targeting BCL2L11 in Small-Tail Han Sheep [J]. Scientia Agricultura Sinica, 2022, 55(9): 1868-1876.
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