State Key Laboratory of Animal Nutrition, Institute of Animal Science, Chinese Academy of Agricultural Sciences, Beijing, China.
Guangdong Provincial Key Laboratory of Malignant Tumor Epigenetics and Gene Regulation, Guangdong-Hong Kong Joint Laboratory for RNA Medicine, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, Guangzhou, China.
Cell Prolif. 2021 Dec;54(12):e13131. doi: 10.1111/cpr.13131. Epub 2021 Oct 14.
RNF20 is recognized as a main E3 ligase for monoubiquitination of histone H2B at lysine 120 (H2Bub). The critical role of RNF20 and H2Bub in various molecular events, such as DNA replication, RNA transcription, and DNA damage response, has been widely investigated and documented. However, its role in porcine adipogenesis remains unknown. In this study, we aimed to clarify the effect of RNF20 on porcine preadipocyte differentiation.
Backfat tissues from fat-type pigs (Bama and Meishan) and lean-type pigs (Yorkshire and Landrace) were collected to detect the expression level of RNF20. Preadipocytes were isolated from Bama piglets and induced to differentiation. Small interfering RNAs were applied to deplete RNF20. Oil Red O staining, quantitative real-time PCR, RNA-seq, Western blot analysis, and EdU assays were performed to study the regulatory mechanism of RNF20 during adipogenesis.
We found that the expression levels of RNF20 and H2Bub were significantly higher in backfat tissues from fat-type pigs than in those from lean-type pigs. Consistently, the significantly induced expression of RNF20 and H2Bub was also observed in porcine differentiated adipocytes. In addition, knockdown of RNF20 greatly inhibited porcine adipogenesis, as evidenced by dramatically decreased lipid droplet formation and lower expression levels of adipogenic transcription masters in RNF20 knockdown cells. Mechanistically, the depletion of RNF20 decreases the cell proliferation and the level of p-C/EBPβ via the Ras-Raf-MEK1/2-ERK1/2 cascade pathway at the mitotic clonal expansion phase and therefore suppresses cell differentiation.
Our results demonstrate that RNF20 is required for porcine preadipocyte differentiation.
RNF20 被认为是组蛋白 H2B 赖氨酸 120 单泛素化（H2Bub）的主要 E3 连接酶。RNF20 和 H2Bub 在多种分子事件中的关键作用，如 DNA 复制、RNA 转录和 DNA 损伤反应，已被广泛研究和记录。然而，其在猪脂肪生成中的作用尚不清楚。本研究旨在阐明 RNF20 对猪前体脂肪细胞分化的影响。
收集来自脂肪型猪（巴马和梅山）和瘦肉型猪（约克夏和长白）的背脂组织，检测 RNF20 的表达水平。从巴马仔猪中分离前体脂肪细胞并诱导分化。应用小干扰 RNA 耗尽 RNF20。进行油红 O 染色、定量实时 PCR、RNA-seq、Western blot 分析和 EdU 测定，以研究 RNF20 在脂肪生成过程中的调节机制。
我们发现 RNF20 和 H2Bub 的表达水平在脂肪型猪的背脂组织中明显高于瘦肉型猪。同样，在猪分化的脂肪细胞中也观察到 RNF20 和 H2Bub 的显著诱导表达。此外，RNF20 的敲低极大地抑制了猪脂肪生成，这表现为脂质滴形成显著减少，RNF20 敲低细胞中脂肪生成转录大师的表达水平降低。在机制上，RNF20 的耗竭通过 Ras-Raf-MEK1/2-ERK1/2 级联途径降低有丝分裂克隆扩展阶段的细胞增殖和 p-C/EBPβ 的水平，从而抑制细胞分化。
我们的结果表明，RNF20 是猪前体脂肪细胞分化所必需的。
