Higher-order chromatin structure and regulatory sequence variation in human induced pluripotent stem cell (iPSC) self-renewal and differentiation

人类诱导多能干细胞（iPSC）自我更新和分化的高阶染色质结构和调控序列变异

• 批准号: MR/T016787/1
• 负责人: Stefan Schoenfelder
• 金额: $ 48.5万
• 依托单位: Babraham Institute
• 依托单位国家: 英国
• 项目类别: Fellowship
• 财政年份: 2019
• 资助国家: 英国
• 起止时间: 2019 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=MR%2FT016787%2F1
• 关键词: Higher; order; chromatin; structure; regulatory

The human body contains over 200 different cell types which fulfil highly specialized functions in different organs such as the liver, brain, heart, and skin. Pluripotent stem cells (PSCs) are fascinating because they can give rise to each of these specialized cell types, in a process called cell differentiation. This unique property (pluripotency) makes PSCs not only immensely valuable to study development, but also endows them with great promise for medical applications. A discovery honoured with the 2012 Nobel prize found that the process of differentiating PSCs into specialized cells can be reversed: mature skin cells can be reprogrammed to become so-called induced PSCs (iPSCs). In the clinic, this reprogramming makes it possible to derive an entire palette of bespoke cells from a patient's skin cells (via iPSCs), for example to replace damaged cells or even entire organs. This is a key aim of regenerative medicine, and it has the potential to solve the problems of the shortage of organs available for donation, and of organ transplant rejection.All cells contain the same genes; what makes cell types different is which sets of genes are 'on' and 'off' in a particular cell type. This determines which proteins (the cell's workhorses) are present in a cell; only 'on' genes are expressed and translated into proteins (for example, in liver cells a different set of genes is 'on' than in PSCs). Interestingly, although genes control cell function, they make up only a small amount of the DNA in our cells (~2%). The vast majority of our DNA (~98%) is non-coding, i.e. does not carry the information to make proteins. Once thought to be 'junk' DNA without a function, we now know that this non-coding DNA contains a plethora of so-called regulatory elements. These regulatory elements function like molecular switches by controlling which genes are 'on' and when. Regulatory elements make physical contacts with their target genes to switch them on. This is achieved by DNA looping; the genome folds in three dimensions (3D) in a cell-type specific manner to bring the appropriate switches and genes together.We often talk of THE human genome, but this is misleading as there are billions of human genomes - we are all different genetically (apart from identical twins). The sequence differences in our genomes are largely confined to the non-coding genome, and can explain not only why we all look different, but also why the cells in our bodies function in a slightly different way, for example in the context of disease. In iPS cells, this naturally occurring sequence variation is thought to be the reason for functional heterogeneity: some iPS cell lines can be differentiated more efficiently into liver cells, whereas other iPS cell lines can be differentiated more efficiently into brain cells, and yet others show a generally poor differentiation efficiency towards all more specialised cell types. Understanding this phenomenon mechanistically is important for the clinic, as it will in the future help to tailor treatment options in regenerative medicine specifically to the requirements of individual patients; this is a key aim of personalised medicine.Because of their immense potential for basic research and regenerative medicine, it is crucial that we understand the molecular mechanisms that endow iPSCs with their unique properties. The aim of this project therefore is to i) map the 3D genome folding in iPSCs to uncover which regulatory elements they use, ii) elucidate how this repertoire of regulatory elements changes as iPSCs differentiate, and iii) identify the genetic variants in these regulatory elements that cause iPS cell lines to differentiate more/less efficiently into specialised cell types.This knowledge is also important in research outside of PSCs, as aberrant 3D genome folding can bring together the wrong pairs of regulatory elements and genes, which can lead to disease including developmental malformations and cancer.

人体包含200多种不同的细胞类型，它们在不同的器官中发挥高度专业化的功能，如肝脏，大脑，心脏和皮肤。多能干细胞（PSC）是迷人的，因为它们可以在称为细胞分化的过程中产生这些专门的细胞类型中的每一种。这种独特的特性（多能性）使PSC不仅对研究发育非常有价值，而且还赋予它们在医学应用方面的巨大前景。2012年诺贝尔奖的一项发现，PSC分化为特化细胞的过程可以逆转：成熟的皮肤细胞可以重新编程成为所谓的诱导PSC（iPSC）。在临床上，这种重新编程使得从患者的皮肤细胞（通过iPSC）中获得整个定制细胞成为可能，例如替换受损细胞甚至整个器官。这是再生医学的一个关键目标，它有可能解决可供捐赠的器官短缺和器官移植排斥的问题。所有细胞都含有相同的基因;使细胞类型不同的是哪组基因在特定细胞类型中是“打开”和“关闭”的。这决定了哪些蛋白质（细胞的主力）存在于细胞中;只有“on”基因表达并翻译成蛋白质（例如，在肝细胞中，与PSC中不同的一组基因是“on”）。有趣的是，虽然基因控制细胞功能，但它们只占我们细胞中DNA的一小部分（约2%）。我们的绝大多数DNA（约98%）是非编码的，即不携带制造蛋白质的信息。曾经被认为是没有功能的“垃圾”DNA，我们现在知道这种非编码DNA含有过多的所谓调控元件。这些调控元件就像分子开关一样，通过控制哪些基因“打开”以及何时“打开”来发挥作用。调控元件与靶基因发生物理接触，从而开启它们。这是通过DNA循环实现的;基因组以细胞类型特异性的方式在三维（3D）中折叠，将适当的开关和基因结合在一起。我们经常谈论人类基因组，但这是误导性的，因为有数十亿个人类基因组-我们在遗传上都是不同的（除了同卵双胞胎）。我们基因组中的序列差异主要局限于非编码基因组，不仅可以解释为什么我们看起来都不同，而且还可以解释为什么我们体内的细胞以略微不同的方式发挥作用，例如在疾病的背景下。在iPS细胞中，这种天然存在的序列变异被认为是功能异质性的原因：一些iPS细胞系可以更有效地分化为肝细胞，而其他iPS细胞系可以更有效地分化为脑细胞，而其他iPS细胞系对所有更特化的细胞类型表现出普遍较差的分化效率。从机理上理解这一现象对临床非常重要，因为它将有助于在未来根据个体患者的需求定制再生医学的治疗方案;这是个性化医疗的关键目标。由于其在基础研究和再生医学方面的巨大潜力，我们了解赋予iPSC独特特性的分子机制至关重要。因此，该项目的目的是i）绘制iPSC中的3D基因组折叠图，以揭示它们使用哪些调节元件，ii）阐明随着iPSC分化，调节元件的库如何变化，以及iii）鉴定这些调节元件中的遗传变异导致iPS细胞系更有效/更低效地分化为专门的细胞类型。这些知识在PSC之外的研究中也很重要，因为异常的3D基因组折叠可以将错误的调控元件和基因对聚集在一起，这可能导致包括发育畸形和癌症在内的疾病。

项目成果

3D genome organization links non-coding disease-associated variants to genes.

• DOI: 10.3389/fcell.2022.995388
• 发表时间: 2022
• 期刊: Frontiers in cell and developmental biology
• 影响因子: 5.5

High-resolution three-dimensional chromatin profiling of the Chinese hamster ovary cell genome.

• DOI: 10.1002/bit.27607
• 发表时间: 2021-03
• 期刊: Biotechnology and bioengineering
• 影响因子: 3.8
• 作者: Bevan S;Schoenfelder S;Young RJ;Zhang L;Andrews S;Fraser P;O'Callaghan PM
• 通讯作者: O'Callaghan PM

Transcription-dependent cohesin repositioning rewires chromatin loops in cellular senescence.

• DOI: 10.1038/s41467-020-19878-4
• 发表时间: 2020-11-27
• 期刊: Nature communications
• 影响因子: 16.6
• 作者: Olan I;Parry AJ;Schoenfelder S;Narita M;Ito Y;Chan ASL;Slater GSC;Bihary D;Bando M;Shirahige K;Kimura H;Samarajiwa SA;Fraser P;Narita M
• 通讯作者: Narita M

Widespread reorganisation of pluripotent factor binding and gene regulatory interactions between human pluripotent states.

• DOI: 10.1038/s41467-021-22201-4
• 发表时间: 2021-04-07
• 期刊: Nature communications
• 影响因子: 16.6
• 作者: Chovanec P;Collier AJ;Krueger C;Várnai C;Semprich CI;Schoenfelder S;Corcoran AE;Rugg-Gunn PJ
• 通讯作者: Rugg-Gunn PJ

Dynamic DNA methylation turnover at the exit of pluripotency epigenetically primes gene regulatory elements for hematopoietic lineage specification

• DOI: 10.1101/2023.01.11.523441
• 发表时间: 2023-01
• 期刊: bioRxiv
• 作者: Aled J. Parry;C. Krueger;T. Lohoff;S. Wingett;S. Schoenfelder;W. Reik