miR-33a靶向Lipin1和IRS2调节绵羊前体脂肪细胞分化的研究

doi:10.11843/j.issn.0366-6964.2020.04.006

Abstract

Abstract: The aim of this study was to reveal the biological functions of miR-33a during the differentiation of ovine preadipocytes. In this study, preadipocytes from the back fat of 15-day-old male lambs were used as research material. All the experiments in this study had 3 replicates. Bioinformatics software was used to predict the target genes of miR-33a. The potential target genes were verified by double luciferase reporter system. In order to reveal the expression pattern of miR-33a and its targeting genes during the differentiation of ovine preadipocytes, qPCR and Western blotting were used to detect the expressions of miR-33a, Lipin1, and IRS2 and proteins. In order to elucidate the regulatory mechanism of miR-33a on its target genes, miR-33a was overexpressed or interfered using lentivirus-mediated method. The expressions of Lipin1, IRS2 and adipogenic marker genes in ovine preadipocytes were detected. The lipid droplet deposition ability was measured by Oil Red O staining. The binding sites of miR-33a with 3'-UTR of Lipin1 and IRS2 were found using bioinformatics prediction. miR-33a significantly down-regulated the relative fluorescence activity of Lipin1 and IRS2 wild-type double fluorescent plasmids (P<0.05). The expression levels of miR-33a and Lipin1, IRS2 exhibited opposite trends during ovine preadipocyte differentiation. The overexpression of miR-33a significantly down-regulated the expressions of Lipin1 (P<0.01), IRS2 (P<0.05) and their encoded proteins, and adipogenic marker genes. After the miR-33a was interfered, the expressions of these genes and proteins were significantly up-regulated. Overexpression of miR-33a reduced lipid droplet deposition, and interference of miR-33a promoted lipid droplet deposition. In conclusion, the expressions of miR-33a and Lipin1, IRS2 were negative correlated during ovine preadipocyte differentiation. miR-33a negatively regulates the differentiation of ovine preadipocytes and the lipid droplets deposition by targeting the 3'-UTR of Lipin1 and IRS2.

Key words: miR-33a, Lipin1, IRS2, sheep, adipocyte differentiation

CLC Number:

S826.2

WANG Qiang, PAN Yangyang, QIAO Liying, LIU Jianhua, ZHAO Bishi, LIU Xuying, WANG Feng, LIANG Yu, LIU Wenzhong. miR-33a Regulates Ovine Preadipocyte Differentiation by Targeting Lipin1 and IRS2[J]. Acta Veterinaria et Zootechnica Sinica, 2020, 51(4): 701-712.

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miR-33a Regulates Ovine Preadipocyte Differentiation by Targeting Lipin1 and IRS2

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