Jang Hye-Jeong, Park Hwan Hee, Linh Tran Thi Thuy, Lee Hak-Kyo, Song Ki-Duk, Lee Woon Kyu
Department of Animal Biotechnology, Chonbuk National University, Jeonju 561-756, Korea .
Asian-Australas J Anim Sci. 2015 Dec;28(12):1721-8. doi: 10.5713/ajas.15.0497.
Embryonic stem cells (ESCs) have been used as a powerful tool for research including gene manipulated animal models and the study of developmental gene regulation. Among the critical regulatory factors that maintain the pluripotency and self-renewal of undifferentiated ESCs, NANOG plays a very important role. Nevertheless, because pluripotency maintaining factors and specific markers for livestock ESCs have not yet been probed, few studies of the NANOG gene from domestic animals including bovine have been reported. Therefore, we chose mouse ESCs in order to understand and compare NANOG expression between bovine, human, and mouse during ESCs differentiation. We cloned a 600 bp (-420/+181) bovine NANOG 5'-flanking region, and tagged it with humanized recombinant green fluorescent protein (hrGFP) as a tracing reporter. Very high GFP expression for bovine NANOG promoter was observed in the mouse ESC line. GFP expression was monitored upon ESC differentiation and was gradually reduced along with differentiation toward neurons and adipocyte cells. Activity of bovine NANOG (-420/+181) promoter was compared with already known mouse and human NANOG promoters in mouse ESC and they were likely to show a similar pattern of regulation. In conclusion, bovine NANOG 5-flanking region functions in mouse ES cells and has characteristics similar to those of mouse and human. These results suggest that bovine gene function studied in mouse ES cells should be evaluated and extrapolated for application to characterization of bovine ES cells.
胚胎干细胞（ESCs）已成为一种强大的研究工具，用于包括基因操作动物模型和发育基因调控研究在内的多个领域。在维持未分化胚胎干细胞的多能性和自我更新的关键调控因子中，NANOG起着非常重要的作用。然而，由于尚未探究家畜胚胎干细胞的多能性维持因子和特异性标记物，关于包括牛在内的家畜NANOG基因的研究报道较少。因此，为了了解和比较牛、人和小鼠胚胎干细胞分化过程中NANOG的表达情况，我们选择了小鼠胚胎干细胞。我们克隆了一段600 bp（-420/+181）的牛NANOG 5'侧翼区域，并用人源化重组绿色荧光蛋白（hrGFP）作为追踪报告基因对其进行标记。在小鼠胚胎干细胞系中观察到牛NANOG启动子的绿色荧光蛋白表达非常高。在胚胎干细胞分化过程中监测绿色荧光蛋白的表达，随着向神经元和脂肪细胞的分化，其表达逐渐降低。在小鼠胚胎干细胞中，将牛NANOG（-420/+181）启动子的活性与已知的小鼠和人NANOG启动子进行比较，它们可能表现出相似的调控模式。总之，牛NANOG 5'侧翼区域在小鼠胚胎干细胞中发挥作用，并且具有与小鼠和人相似的特征。这些结果表明，在小鼠胚胎干细胞中研究的牛基因功能应进行评估并外推，以应用于牛胚胎干细胞的特性表征。
