miR-150靶向AOC3调控绵羊前体脂肪细胞分化的研究

doi:10.11843/j.issn.0366-6964.2023.08.013

Abstract

Abstract: The aim of this study was to reveal the action mechanism of miR-150 during the differentiation of preadipocytes in Guangling Large-Tailed sheep by investigating the relationship between miR-150 and target gene AOC3. In this study, three 5-day-old Guangling Large-Tailed male sheep with normal development and health were used as the research material. The expression level of miR-150 in subcutaneous, perirenal and tail adipose tissues of Guangling Large-Tailed sheep was detected by qPCR. Tail adipose tissue of Guangling large-tailed sheep was collected to isolate ovine preadipocytes. Cell purity was identified by cell immunofluorescence. Bioinformatics software was used to predict the target relationship between miR-150 and AOC3, and double luciferase reporter system was used to verify this relationship. miR-150 mimics and inhibitors were transfected into preadipocyte and the expression of AOC3 and adipogenic marker genes were detected. The overexpression and interference vector of ovine AOC3 were constructed and transfected into ovine preadipocytes. qPCR and Western blotting were used to detect the expression patterns of miR-150 and AOC3 during the differentiation of preadipocytes in Guangling Large-Tailed sheep. Lipid accumulation ability was detected by Oil Red O staining. Three biological replicates were set up for each of the above tests. The results showed that the expression of miR-150 was the highest in tail fat, and the difference was significant (P<0.001). After overexpression of miR-150, the expression of AOC3 and adipogenic marker genes were significantly down-regulated (P<0.05), lipid droplet accumulation decreased. These results implied that miR-150 inhibits the differentiation of ovine preadipocytes. The results were opposite after inhibition of miR-150. There was a binding site between miR-150 and the 3'-UTR of AOC3, and miR-150 was extremely significant decreased the relative fluorescence activity of wild type AOC3 (P<0.001). After overexpressing AOC3, the expression of adipogenic marker genes were significantly up-regulated (P<0.01) and more lipid droplets were produced, indicating that AOC3 promotes the differentiation of ovine preadipocytes. After interfering with AOC3, the result was reversed. In conclusion, there is a binding site between miR-150 and the 3'-UTR of AOC3. The expression level of miR-150 was negatively correlated with that of AOC3 during the differentiation of ovine preadipocyte. miR-150 inhibits the differentiation of ovine preadipocytes by targeting AOC3. The results provide a theoretical basis for the research of microRNAs (miRNAs) at the molecular level.

Key words: Guangling Large-Tailed sheep, miR-150, AOC3, adipocyte differentiation

CLC Number:

S826.2

ZHANG Hanyue, ZHAO Dan, LIANG Yu, ZHAO Bishi, FAN Mengdan, QIAO Liying, LIU Jianhua, YANG Kaijie, PAN Yangyang, LIU Wenzhong. miR-150 Regulates Ovine Preadipocyte Differentiation by Targeting AOC3[J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(8): 3262-3274.

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miR-150 Regulates Ovine Preadipocyte Differentiation by Targeting AOC3

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