Science for Life Laboratory, Department of Medical Biochemistry and Microbiology, Uppsala University, Uppsala, Sweden.
Science for Life Laboratory, Department of Medical Cell Biology, Uppsala University, Uppsala, Sweden.
FASEB J. 2020 Aug;34(8):10250-10266. doi: 10.1096/fj.201901321R. Epub 2020 Jun 19.
The transcription factor ZBED6 acts as a repressor of Igf2 and affects directly or indirectly the transcriptional regulation of thousands of genes. Here, we use gene editing in mouse C2C12 myoblasts and show that ZBED6 regulates Igf2 exclusively through its binding site 5'-GGCTCG-3' in intron 1 of Igf2. Deletion of this motif (Igf2 ) or complete ablation of Zbed6 leads to ~20-fold upregulation of the IGF2 protein. Quantitative proteomics revealed an activation of Ras signaling pathway in both Zbed6 and Igf2 myoblasts, and a significant enrichment of mitochondrial membrane proteins among proteins showing altered expression in Zbed6 myoblasts. Both Zbed6 and Igf2 myoblasts showed a faster growth rate and developed myotube hypertrophy. These cells exhibited an increased O consumption rate, due to IGF2 upregulation. Transcriptome analysis revealed ~30% overlap between differentially expressed genes in Zbed6 and Igf2 myotubes, with an enrichment of upregulated genes involved in muscle development. In contrast, ZBED6-overexpression in myoblasts led to cell apoptosis, cell cycle arrest, reduced mitochondrial activities, and ceased myoblast differentiation. The similarities in growth and differentiation phenotypes observed in Zbed6 and Igf2 myoblasts demonstrates that ZBED6 affects mitochondrial activity and myogenesis largely through its regulation of IGF2 expression. This study adds new insights how the ZBED6-Igf2 axis affects muscle metabolism.
转录因子 ZBED6 作为 Igf2 的抑制剂，直接或间接影响数千个基因的转录调控。在这里，我们使用基因编辑技术在小鼠 C2C12 成肌细胞中进行研究，结果表明 ZBED6 通过其在 Igf2 内含子 1 中的结合位点 5'-GGCTCG-3' 来特异性调节 Igf2。该基序缺失（Igf2 ）或完全敲除 Zbed6 会导致 IGF2 蛋白的表达上调约 20 倍。定量蛋白质组学显示，在 Zbed6 和 Igf2 成肌细胞中均激活了 Ras 信号通路，并且在 Zbed6 成肌细胞中表达改变的蛋白质中，线粒体膜蛋白显著富集。Zbed6 和 Igf2 成肌细胞的生长速度均加快，并发生肌管肥大。由于 IGF2 的上调，这些细胞的 O 消耗率增加。转录组分析显示，Zbed6 和 Igf2 肌管中差异表达基因之间有 ~30%的重叠，上调基因富集参与肌肉发育。相比之下，ZBED6 在成肌细胞中的过表达会导致细胞凋亡、细胞周期停滞、线粒体活性降低和成肌细胞分化停止。Zbed6 和 Igf2 成肌细胞中观察到的生长和分化表型的相似性表明，ZBED6 通过调节 IGF2 的表达来影响线粒体活性和肌肉生成。本研究为 ZBED6-Igf2 轴如何影响肌肉代谢提供了新的见解。
