Post-Translational Regulation of Oct4 Transcriptional Activity

Oct4 is a key component of the molecular circuitry which regulates embryonic stem cell proliferation and differentiation. It is essential for maintenance of undifferentiated, pluripotent cell populations, and accomplishes these tasks by binding DNA in multiple heterodimer and homodimer configurations. Very little is known about how formation of these complexes is regulated, or the mechanisms through which Oct4 proteins respond to complex extracellular stimuli which regulate pluripotency. Here, we provide evidence for a phosphorylation-based mechanism which regulates specific Oct4 homodimer conformations. Point mutations of a putative phosphorylation site can specifically abrogate transcriptional activity of a specific homodimer assembly, with little effect on other configurations. Moreover, we performed bioinformatic predictions to identify a subset of Oct4 target genes which may be regulated by this specific assembly, and show that altering Oct4 protein levels affects transcription of Oct4 target genes which are regulated by this assembly but not others. Finally, we identified several signaling pathways which may mediate this phosphorylation and act in combination to regulate Oct4 transcriptional activity and protein stability. These results provide a mechanism for rapid and reversible alteration of Oct4 transactivation potential in response to extracellular signals.

Oct4是调控胚胎干细胞增殖与分化的分子调控环路的关键组分。其对于维持未分化的多能性细胞群体至关重要，并通过以多种异二聚体（heterodimer）与同二聚体（homodimer）构象结合DNA来完成上述功能。目前，学界对于此类复合物的形成调控机制，以及Oct4蛋白如何响应调控多能性的复杂细胞外刺激的分子机制，仍知之甚少。本研究为基于磷酸化的Oct4特定同二聚体构象调控机制提供了实验证据。对推定磷酸化位点的点突变可特异性阻断特定同二聚体组装体的转录活性，而对其他构象几乎无影响。此外，我们通过生物信息学预测，鉴定出可能受该特定组装体调控的Oct4靶基因子集，并证实改变Oct4蛋白水平会影响该组装体调控的Oct4靶基因的转录，而非其他靶基因。最后，我们鉴定出数种可介导该磷酸化过程，并协同调控Oct4转录活性与蛋白稳定性的信号通路。本研究结果为细胞外信号刺激下Oct4转录激活潜能的快速可逆改变提供了潜在的分子机制。