Instituto de Medicina Molecular and Instituto de Histologia e Biologia do Desenvolvimento, Faculdade de Medicina da Universidade de Lisboa, Avenida Prof. Egas Moniz, Lisboa 1649-028, Portugal Champalimaud Neuroscience Programme, Champalimaud Centre for the Unknown, Avenida Brasilia - Doca de Pedrouços, Lisboa 1400-038, Portugal
Instituto de Medicina Molecular and Instituto de Histologia e Biologia do Desenvolvimento, Faculdade de Medicina da Universidade de Lisboa, Avenida Prof. Egas Moniz, Lisboa 1649-028, Portugal Champalimaud Neuroscience Programme, Champalimaud Centre for the Unknown, Avenida Brasilia - Doca de Pedrouços, Lisboa 1400-038, Portugal.
Development. 2014 Jul;141(14):2770-9. doi: 10.1242/dev.108910.
Heterogeneous expression of the transcription factor NANOG has been linked to the existence of various functional states in pluripotent stem cells. This heterogeneity seems to arise from fluctuations of Nanog expression in individual cells, but a thorough characterization of these fluctuations and their impact on the pluripotent state is still lacking. Here, we have used a novel fluorescent reporter to investigate the temporal dynamics of NANOG expression in mouse embryonic stem cells (mESCs), and to dissect the lineage potential of mESCs at different NANOG states. Our results show that stochastic NANOG fluctuations are widespread in mESCs, with essentially all expressing cells showing fluctuations in NANOG levels, even when cultured in ground-state conditions (2i media). We further show that fluctuations have similar kinetics when mESCs are cultured in standard conditions (serum plus leukemia inhibitory factor) or ground-state conditions, implying that NANOG fluctuations are inherent to the pluripotent state. We have then compared the developmental potential of low-NANOG and high-NANOG mESCs, grown in different conditions, and confirm that mESCs are more susceptible to enter differentiation at the low-NANOG state. Further analysis by gene expression profiling reveals that low-NANOG cells have marked expression of lineage-affiliated genes, with variable profiles according to the signalling environment. By contrast, high-NANOG cells show a more stable expression profile in different environments, with minimal expression of lineage markers. Altogether, our data support a model in which stochastic NANOG fluctuations provide opportunities for mESCs to explore multiple lineage options, modulating their probability to change functional state.
转录因子 NANOG 的异质性表达与多能干细胞中各种功能状态的存在有关。这种异质性似乎源于单个细胞中 Nanog 表达的波动，但对这些波动及其对多能状态的影响的全面描述仍然缺乏。在这里，我们使用一种新的荧光报告基因来研究小鼠胚胎干细胞（mESCs）中 NANOG 表达的时间动态，并剖析不同 NANOG 状态下 mESCs 的谱系潜能。我们的结果表明，随机的 NANOG 波动在 mESCs 中广泛存在，即使在基础状态（2i 培养基）下培养时，基本上所有表达细胞的 NANOG 水平都显示出波动。我们进一步表明，当 mESCs 在标准条件（血清加白血病抑制因子）或基础状态下培养时，波动具有相似的动力学，这意味着 NANOG 波动是多能状态所固有的。然后，我们比较了在不同条件下培养的低 NANOG 和高 NANOG mESCs 的发育潜能，并证实 mESCs 在低 NANOG 状态下更容易进入分化。通过基因表达谱分析的进一步分析表明，低 NANOG 细胞具有明显的谱系相关基因表达，根据信号环境的不同，其表达谱也不同。相比之下，高 NANOG 细胞在不同环境中表现出更稳定的表达谱，谱系标记物的表达最小。总之，我们的数据支持这样一种模型，即随机的 NANOG 波动为 mESCs 提供了探索多种谱系选择的机会，从而调节它们改变功能状态的概率。
