Characterisation of a novel NANOG / KDM4B complex to regulate heterochromatin function and chromosome stability in pluripotent stem cells

多能干细胞（PSC）是一种非特化的细胞，可以形成身体的任何细胞类型。目前，人们非常希望PSC可以用于基于细胞的疗法，用于治疗疾病，替代衰老的组织，以及更好地了解人类发育，但在实现这些目标之前，仍有几个障碍必须克服。其中一个障碍是遗传不稳定的出现，特别是过多染色体的积累，这可能会影响PSC。这些不必要的变化是如何产生的仍然知之甚少，但科学家们正试图阻止这些变化的发生，以生产更安全、质量更好的PSC。我们选择研究基因组中令人兴奋而神秘的组成异染色质。在许多不同的细胞类型中，异染色质对关键的细胞过程很重要，包括遗传稳定性的维持和染色体数量的控制，尽管迄今为止在PSC中的研究相对较少。在我们最近的工作中，我们已经确定了PSC中异染色质控制的新途径。出乎意料的是，这个新途径使用了几个众所周知的干细胞因子，但我们现在能够赋予它们新的功能。重要的是，当我们使用遗传技巧来阻止这些因子在PSC中发挥作用时，它会导致异染色质组织的缺陷，导致遗传稳定性的相关丧失，并导致细胞中额外染色体的积累。干细胞因子、异染色质组织和染色体稳定性控制之间的这种意想不到的联系很重要，因为它可以解释PSC中遗传变化是如何出现的，并可能使研究人员能够防止这种情况发生。我们的研究使我们形成了异染色质直接参与PSC遗传不稳定性的假设。因此，本研究计划的总体目标是确定PSC中的异染色质是如何被控制的，以及当这种控制水平出错时会发生什么。我们仔细规划了三个主要目标来验证我们的假设。第一个目标是确定干细胞因子如何控制PSC中的异染色质。我们的研究强烈地暗示了一个额外的因素，KDM4B的参与，所以我们想更详细地检查这个因素。我们将通过询问KDM4B是否定位于PSC中的异染色质来实现这一点，然后测量当我们从PSC中去除KDM4B时异染色质发生了什么。我们预测异染色质在PSC中会出现缺陷，并可能出现染色体不稳定。第二个目的是研究异染色质缺陷如何导致PSC染色体不稳定。先前许多实验室的研究表明，标记和定义异染色质的特定信号是防止遗传不稳定所必需的，我们希望通过研究PSC中如何控制这些信号来推进这一研究，特别是考虑到我们现在已经确定的异染色质调节的新模式。第三个目标是利用我们产生的知识来提高PSC的质量和遗传稳定性，这样我们就可以消除目前对未来应用的障碍之一。我们预计，如果我们能够理解异染色质组织是如何与PSC中的染色体不稳定性联系在一起的，那么我们就可以在未来设计出防止这种情况发生的方法。了解这种情况发生的详细机制可能会改善干细胞在再生医学中的应用。这种知识在PSC以外的研究中也很重要，特别是在衰老和癌症等异染色质正常调节过程被破坏的领域。通过更好地了解异染色质是如何被控制的，我们可能能够制定出检测早期变化的策略，并防止它们的发生。

Pluripotent stem cells (PSC) are unspecialised cells that can form any cell type of the body. There is currently much hope that PSC could be used for cell-based therapies for the treatment of diseases, replacement for worn out tissues as we age, and for better understanding of human development, but there are still several hurdles that must be overcome before these goals are achieved. One of the hurdles is the appearance of genetic instability, and in particular the accumulation of too many chromosomes, that can affect PSC. How these unwanted changes arise remains poorly understood, but scientists are trying to prevent the changes from occurring in order to produce safer and better quality PSC.We have chosen to study an exciting and mysterious part of our genome called constitutive heterochromatin. In many different cell types, heterochromatin is important for key cellular processes, including the maintenance of genetic stability and control of chromosome number, although it has been relatively poorly studied in PSC so far. In our recent work, we have identified a new pathway through which heterochromatin is controlled in PSC. Unexpectedly, this new pathway uses several well-known stem cell factors but we are now able to assign new functions to them. Importantly, we when use genetic tricks to prevent these factors from functioning in PSC, it leads to defects in heterochromatin organisation, to the associated loss of genetic stability, and to the accumulation of additional chromosomes in the cells. This unanticipated connection between stem cell factors, heterochromatin organisation and the control of chromosome stability is important because it could provide an explanation for how genetic changes appear in PSC and would potentially allow researchers to prevent this from occurring. Our research has led us to form the hypothesis that heterochromatin is involved directly in the genetic instability of PSC. The overall aim in this research proposal, therefore, is to determine how heterochromatin is controlled in PSC, and what happens when this level of control goes wrong. We have carefully planned three main objectives to test our hypothesis.The first objective is to define how the stem cell factors control heterochromatin in PSC. Our research strongly suggests the involvement of an additional factor, KDM4B, and so we would like to examine this factor in more detail. We will achieve this by asking whether KDM4B localises to heterochromatin in PSC, and then measure what happens to heterochromatin when we remove Kdm4b from PSC. We predict that heterochromatin will show defects and possibly the appearance of chromosome instability in the PSC.The second objective is to investigate how defects in heterochromatin lead to chromosome instability in PSC. Previous research from many laboratories has shown that the particular signals that mark and define heterochromatin are required to prevent genetic instability, and we want to take this forward by investigating how these signals are controlled in PSC, especially in light of the new mode of heterochromatin regulation that we have now identified.The third objective is to use the knowledge that we generate to improve the quality and genetic stability of PSC so that we can remove one of the current hurdles to future applications. We anticipate that if we can understand how heterochromatin organisation is connected to chromosome instability in PSC, then we can, in future, devise ways to prevent this from happening. Understanding the detailed mechanism of how this occurs may lead to improved use of stem cells for regenerative medicine. This knowledge is also important in research outside of PSC, especially in ageing and cancer for example, where the normal process of heterochromatin regulation is disrupted. By better understanding how heterochromatin is controlled in general, we may be able to develop strategies to detect early changes and also to prevent them from happening.