Study the role(s) of p53 in embryonic stem cells

在2012财年，我们在绘制胚胎干细胞的全基因组p53信号图谱方面取得了重大进展。1)已知P53是ES细胞的有效分化诱导剂。然而，P53调控ES细胞分化的机制尚不完全清楚。包括我们在内的一些研究表明，p53通过抑制ES丰富的基因的转录而诱导分化，例如ES细胞的主要调节因子Nanog。然而，对于ES细胞的维持，Nanog并不是严格要求的，这表明存在其他机制。为了探索这些其他机制，我们在研究中引入了功能强大的芯片序列技术。利用这项技术，我们令人惊讶地发现，p53不仅下调ES丰富的基因，而且上调与分化相关的基因。2)此外，P53还通过不同的机制调控ES富集型基因和分化相关基因的表达。对于与分化相关的基因，p53往往通过作用于启动子区域来上调其表达。对于ES丰富的基因，P53通常与末端增强子结合并干扰它们的活性。除了在全基因组水平上研究ES细胞中的P53信号外，我们还在单转录水平上进行了研究。我们最近发现了一个ES特异性的p53靶点，即无毛(HR)基因。HR蛋白含有一个JmjC结构域，这是一个组蛋白去甲基化结构域。HR基因敲除可减少P53介导的ES细胞凋亡，提示HR参与了ES细胞的P53基因网络。其他人的早期工作表明，HR可能在淋巴肿大中发挥作用。我们目前的研究集中在研究HR调节P53介导的ES细胞凋亡的机制。这项研究将揭示胚胎干细胞中P53信号的表观遗传调控以及在癌症中的作用。只有2%的基因组表达蛋白质编码基因。基因组的其余部分积极表达非编码RNA，这些RNA在各种生物过程中发挥着重要作用。然而，这些非编码RNA在ES细胞中的作用在很大程度上还没有被探索。利用全基因组的方法，我们已经确定了几个可能参与ES细胞中P53信号转导的非编码RNA。我们目前正在研究这些非编码RNA在ES细胞中的作用。其中一些非编码RNA与染色质结合蛋白结合。根据这些新的观察结果，我们将转录本定义为蛋白质编码和非编码RNA。3)人ES细胞是一种比小鼠ES细胞更好的模型。在2012财年，我们从康涅狄格大学的干细胞设施中获得了两个人类胚胎干细胞系。这两个人类胚胎干细胞系是经过批准的人类胚胎干细胞系，可以使用联邦资金进行研究。我们将使用这两个人类ES细胞系来研究人类ES细胞中的P53信号。然后，我们将比较小鼠ES细胞和人ES细胞中的P53信号，以期获得保守的P53基因网络。

During fiscal year of 2012, we have made significant progress of mapping genome-wide p53 signaling in embryonic stem cells. 1) It is known that p53 is a potent differentiation inducer of ES cells. However, the mechanisms of how p53 regulates the differentiation of ES cells are not fully appreciated. Some studies including ours suggest that p53 induces differentiation by repressing the transcription of ES-enriched genes, such as Nanog, a master regulator of ES cells. However, Nanog is not strictly required for the maintenance of ES cells, suggesting other mechanisms exist. To explore these other mechanisms, we introduced the powerful ChIP-seq technology into our study. Using this technology, we surprisingly found that p53 not only down-regulates ES-enriched genes but also up-regulates differentiation-associated genes. 2) In addition, p53 regulates the expression of ES-enriched genes and differentiation-associated genes using different mechanisms. For differentiation-associated genes, p53 tend to up-regulate their expression through acting in the promoter region. For ES-enriched genes, p53 generally binds to the distal enhancers and interferes with their activity. In addition to studying p53 signaling in ES cells at a genome-wide level, we also investigate this at a single-transcript level. We recently identified a ES-specific p53 target, the hairless (Hr) gene. The Hr protein contains a JmjC domain, which is a histone demethylation domain. Knocking down of Hr decreases p53-mediated apoptosis in ES cells, suggesting that Hr participates in p53 gene network in ES cells. Earlier work by others have shown that Hr may have a role in lymphomagenesis. Our currently study focuses on investigating the mechanism by which Hr regulates p53-mediated apoptosis in ES cells. This study will shed light on the epigenetic regulation of p53 signaling in ES cells as well in cancer. Only 2 percentage of the genome expresses protein-coding genes. The rest of the genome actively expresses non-coding RNAs, which play important roles in various biological processes. However, the roles of these non-coding RNAs in ES cells are largely unexplored. Using genome-wide approaches, we have identified several non-coding RNAs that may be involved in p53 signaling in ES cells. We are currently studying the roles of these non-coding RNAs in ES cells. Some of these non-coding RNAs bind to chromatin binding protein. In light of these new observations, we define transcripts as protein-coding and non-coding RNAs. 3) Human ES cells are a better model than mouse ES cells. In the fiscal year 2012, we have obtained two human ES cell lines from the Stem Cell Facility at the University of Connecticut. These two human ES cell lines are approved human ES cell lines that can be studied using federal funding. We will use study p53 signaling in human ES cells using these two human ES cell lines. Then, we will compared p53 signaling in mouse ES cells to that in human ES cells with the hope to derived conserved p53 gene network.