转录共抑制因子Tle1上游调控因子的筛选
Screening of upstream regulatory factors for the transcriptional repressor Tle1
徐纪 1潘德京1
作者信息
- 1. 苏州大学苏州医学院剑桥-苏大基因组资源中心，江苏省苏州市 215123
- 折叠
摘要
目的探讨成脂分化早期Tle1的上游转录调控网络，明确关键候选转录因子对Tle1表达的直接调控作用。方法基于实验室前期RNA-seq与Genecard网站预测联合筛选3T3-L1细胞成脂早期Tle1的潜在上游调控因子，采用实时荧光定量PCR技术检测候选基因与Tle1表达波动的时序相关性，通过双荧光素酶报告基因实验验证转录因子对Tle1启动子活性的影响，并利用PiggyBac转座系统构建多西环素（Dox）诱导的稳定过表达细胞系。结果 Cebpb与Nfil3的表达波动在成脂分化早期与Tle1呈现高度的时序一致性；双荧光素酶实验证实过表达Cebpb、Nfil3及Rara可显著激活Tle1的启动子活性，而过表达Egr2则显著抑制其启动子活性；成功建立并鉴定了受Dox严密调控的Cebpb与Nfil3稳定过表达3T3-L1细胞系。结论 Cebpb与Nfil3是成脂分化早期调控Tle1表达的核心上游正向转录因子，能直接结合并激活Tle1转录，该调控轴的发现为进一步阐明脂肪细胞发育的早期分子机制奠定了理论基础与模型工具。
Abstract
In this paper, we aimed to explore the upstream transcriptional regulatory network of Tle1 during the early stage of adipogenesis and to clarify the direct regulatory effects of key candidate transcription factors on Tle1 expression. Based on a combined analysis of RNA-seq and predictions from the Genecard website, we screened potential upstream regulatory factors of Tle1 during the early adipogenic differentiation of 3T3-L1 cells. Real-time quantitative PCR (RT-qPCR) was employed to detect the temporal correlation between the expression fluctuations of candidate genes and Tle1. Furthermore, dual-luciferase reporter assays were conducted to verify the effects of these transcription factors on Tle1 promoter activity, and the PiggyBac transposon system was utilized to construct doxycycline (Dox)-inducible stable overexpression cell lines. The results showed that the expression fluctuations of Cebpb and Nfil3 exhibited a high degree of temporal consistency with Tle1 during early adipogenesis. Dual-luciferase assays confirmed that the overexpression of Cebpb, Nfil3, and Rara significantly activated the promoter activity of Tle1, whereas the overexpression of Egr2 significantly inhibited it. Additionally, 3T3-L1 cell lines with stable overexpression of Cebpb and Nfil3, strictly regulated by Dox, were successfully established and identified. In conclusion, Cebpb and Nfil3 are core upstream positive transcription factors that regulate Tle1 expression during early adipogenesis by directly binding to and activating its transcription. The discovery of this regulatory axis lays a theoretical foundation and provides a model tool for further elucidating the early molecular mechanisms of adipocyte development.关键词
Tle1/转录调控/稳转细胞系/双荧光素酶报告基因Key words
Tle1/Transcriptional regulation/Stable cell line/Dual-luciferase reporter gene引用本文复制引用
徐纪,潘德京.转录共抑制因子Tle1上游调控因子的筛选[EB/OL].(2026-04-29)[2026-04-30].http://www.paper.edu.cn/releasepaper/content/202604-212.学科分类
生物科学研究方法、生物科学研究技术/分子生物学
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