Deciphering and overcoming epigenetic erosion at imprinted loci in mouse and human naive pluripotent stem cells

在naïve多能干细胞的常规培养皿培养中，发现这些干细胞在一组名为印迹的基因中获得错误。Naïve多能干细胞是代表胚胎发育最早阶段的细胞，具有完全自我复制或转变为体内任何细胞类型的能力。印记是一种导致基因以特定方式表达的现象。这个过程涉及修饰，DNA甲基化和组蛋白甲基化，在影响其表达的印迹基因的调控区域。这些印记会在父母的种系中自然地消除和建立，并随后在受精卵的所有其他细胞（体细胞）中保持到成年。异常地，这些印迹在naïve多能干细胞的常规培养皿培养中被擦除，阻碍了这些细胞的潜能。我的研究小组最近发现了一种名为Impera的基因，它可能会指导观察到的印迹在印迹基因上的去除。现在我们想要从机制上定义Impera是如何工作的，并定义防止Impera诱导印记清除的策略。实现这些目标将导致发育生物学的根本性发现，第一个确定的能够介导印记消除的基因，以及在小鼠和人类naïve多能干细胞培养中防止印记消除。在医学背景下，适当的印记对正常发育很重要。涉及不适当/缺乏印记的人类疾病包括Angelman综合征和Prader-Willi综合征。因此，了解印记消除发生的机制可能会发现途径和过程，从而增强我们对病理过程的理解。Naïve多能干细胞也被广泛用作早期发育研究的平台，许多投资都集中在了解细胞如何维持其naïve状态以及它们如何致力于成为特定的细胞类型。拟议的研究将为我们如何能够将干细胞稳定在naïve状态提供见解，为用作研究工具、药物发现计划和再生医学应用提供更稳定的干细胞平台。我们的研究小组在干细胞生物学方面有着良好的记录，并且有必要的专业知识来成功完成这个重要的项目。此外，与包括Kevin Chalut博士在内的密切合作伙伴在剑桥干细胞研究所的工作使我们处于战略优势地位。多能性和分化的物理生物学专家，与我的实验室一起开发了一种新的水凝胶方案（手稿准备中），支持naïve多能干细胞自我复制。这可能被证明是一个非常有用的工具，在我们的目标，以产生naïve多能干细胞自由的印记错误

During the routine petri dish culture of naïve pluripotent stem cells it was found that these acquire errors in a group of genes named imprinted. Naïve pluripotent stem cells are cells that represent the very earliest stage of embryonic development, and have the ability to self-replicate exactly or to change into any cell type in the body. Imprinting is a phenomenon that leads to genes being expressed in a parent of origin specific manner. This process involves modifications, DNA methylation and histone methylation, at regulatory regions of imprinted genes that affects their expression. These imprint marks are naturally erased and established in the germline of the parents and are subsequently maintained in all the other cells (soma) from the fertilized egg till adulthood. Abnormally, these imprints are erased in the routine petri dish culture of naïve pluripotent stem cells hindering the potential of these cells. My group has recently discovered a gene, Impera, that potentially directs the observed removal of imprints at imprinted genes. Now we want to define mechanistically how Impera works and to define strategies that prevent Impera from inducing imprint erasure. Achieving these goals will lead to a fundamental discovery in developmental biology, the first identified gene capable of mediating imprint erasure, and to the prevention of imprint erasure in the cultures of both mouse and human naïve pluripotent stem cells. In the medical context, appropriate imprinting is important for normal development. Human diseases involving inappropriate/lack of imprinting include Angelman syndrome and Prader-Willi syndrome. Thus, understanding mechanistic how imprint erasure occurs may uncover pathways and processes that will enhance our understanding of pathological processes. Naïve pluripotent stem cells are also widely used as a platform for early developmental research, with much investment being focussed on both understanding how cells maintain their naïve state and how they commit to becoming specific cell types. The proposed research will provide insights into how we might be able to stabilise stem cells in this naïve state, providing more stable stem cell platforms for use as a research tool, in drug discovery programmes and in regenerative medicine applications. Our research group has a strong track record in stem cell biology and has the necessary expertise to successfully complete this important project. Further, working within the Cambridge Stem Cell Institute puts us in a strategically strong position, with close collaborators including Dr Kevin Chalut. An expert in the physical biology of pluripotency and differentiation which together with my lab developed a novel hydrogel protocol (manuscript in preparation) which supports naïve pluripotent stem cell self-replication. This may prove a very useful tool in our aim to generate naïve pluripotent stem cells free of imprint errors