Biochemistry and Molecular Biology Department, Johns Hopkins University Bloomberg School of Public Health, Baltimore, MD 21205, USA.
Biochemistry and Molecular Biology Department, Johns Hopkins University Bloomberg School of Public Health, Baltimore, MD 21205, USA.
Stem Cell Res. 2020 Dec;49:102078. doi: 10.1016/j.scr.2020.102078. Epub 2020 Nov 5.
The auxin-inducible degron (AID) system is becoming a widely used method for rapid and reversible degradation of target proteins. This system has been successfully used to study gene and protein functions in eukaryotic cells and common model organisms, such as nematode and fruit fly. To date, applications of the AID system in mammalian stem cell research are limited. Furthermore, standard mouse models harboring the AID system have not been established. Here we have explored the utility of the H11 safe-harbor locus for integration of the TIR1 transgene, an essential component of auxin-based protein degradation system. We have shown that the H11 locus can support constitutive and conditional TIR1 expression in mouse and human embryonic stem cells, as well as in mice. We demonstrate that the AID system can be successfully employed for rapid degradation of stable proteins in embryonic stem cells, which is crucial for investigation of protein functions in quickly changing environments, such as stem cell proliferation and differentiation. As embryonic stem cells possess unlimited proliferative capacity, differentiation potential, and can mimic organ development, we believe that these research tools will be an applicable resource to a broad scientific audience.
生长素诱导的降解结构域（AID）系统正成为一种快速且可逆地降解靶蛋白的广泛应用方法。该系统已成功用于研究真核细胞和常见模式生物（如线虫和果蝇）中的基因和蛋白质功能。迄今为止，AID 系统在哺乳动物干细胞研究中的应用有限。此外，还没有建立携带 AID 系统的标准小鼠模型。在这里，我们探索了 H11 安全港基因座用于整合 TIR1 转基因的用途，TIR1 是基于生长素的蛋白质降解系统的一个重要组成部分。我们已经表明，H11 基因座可以支持 TIR1 在小鼠和人类胚胎干细胞以及小鼠中的组成型和条件表达。我们证明 AID 系统可成功用于快速降解胚胎干细胞中的稳定蛋白，这对于研究快速变化环境中的蛋白质功能（如干细胞增殖和分化）至关重要。由于胚胎干细胞具有无限的增殖能力、分化潜力并且可以模拟器官发育，我们相信这些研究工具将成为广大科学界的一种适用资源。
