Unravelling the role of beta-catenin in ground state pluripotency

干细胞由两种特性定义：它们可以无限增殖，产生与自身相同的细胞（自我更新特性），并且可以特化（分化）为成熟细胞类型（多能性特性）。在成年人中，干细胞，例如在骨髓中发现的干细胞，主要在损伤情况下具有修复功能。成体干细胞目前也用于医学治疗;一个典型的例子是骨髓移植治疗白血病。将成体干细胞用于医疗目的的主要限制是低可用性，以及难以在培养中扩增它们以保持其特征完整。此外，成体干细胞不能分化为所有需要的细胞类型，因此再生范围很差。这些问题被胚胎干细胞（ESCs）的发现所克服：从囊胚中分离的ESCs可以在体外无限期培养，并且在适当的条件下可以产生所有类型的细胞。此外，2006年诺贝尔奖赢家山中伸弥做出了惊人的发现：体细胞可以重新编程回到干细胞样状态，获得所谓的诱导多能干细胞（iPSC）。ESCs和iPSCs被认为是一个巨大的希望：许多人认为，它们的发现将在短期内推动再生医学，基因治疗和个性化医学领域的令人印象深刻的扩展，最终导致许多健康相关问题的解决。到目前为止，这一承诺只有一部分得到了实现，主要是因为我们对使这些细胞如此特殊的基本生物过程知之甚少。在这项研究中，我们的目标是提高我们的理解的分子过程，协调多能性集中在β-连环蛋白（β-连环蛋白），一个关键的蛋白质在干性维护。事实上，我们和其他人报道了β-连环蛋白在体内体细胞重编程、分化、多能性和组织再生中的核心作用。尽管如此，b-连环蛋白控制胚胎干细胞多能性的机制仍存在很大争议，我们将用跨学科的方法来揭示这一复杂性。使用尖端技术，我们将工程小鼠胚胎干细胞（mESCs），其中b-连环蛋白可以以多种方式调节（蛋白质量，时间动态，调节其他基因的能力）。随后，我们将进行实验，将特定的b-连环蛋白扰动行为与mESCs的自我更新和分化能力联系起来。最后，我们将使用有效的计算方法来了解b-连环蛋白如何与其他对多能性很重要的基因相互作用。该项目将为胚胎干细胞生物学领域提供创新和知识的重要一步：结果将揭示β-连环蛋白如何调节多能性和分化之间的平衡，并指示如何在体外驱动特定的细胞命运。在未来，所提出的项目的结果可以扩展到控制人ESC和iPSC的b-连环蛋白依赖的自我更新和分化，缩短多能干细胞与其应用靶点之间的距离。此外，开发的技术可以用于其他生物过程中，其中b-连环蛋白参与，包括发展和癌症。

Stem cells are defined by two properties: they can proliferate indefinitely producing cells identical to themselves (self-renewal property), and can specialise (differentiate) into mature cells types (pluripotency property). In adults, stem cells, found for example in the bone marrow, have mainly a repair function in case of injury. Adult stem cells are currently used also in medical therapy; a typical example is bone marrow transplant for leukemia treatment. The major limit of using adult stem cells for medical purposes is the low availability, and the difficulty to expand them in culture maintaining their features intact. Moreover, adult stem cells have a poor regeneration range, as they are unable to differentiate into all desired cell types.Such issues were overcome thanks to the discovery of embryonic stem cells (ESCs): isolated from blastocysts, ESCs can be cultured in vitro indefinitely, and can give rise, under the appropriate conditions, to every cell type. Also, in 2006 the Nobel Prize winner Yamanaka made an astonishing discovery: somatic cells can be reprogrammed back to a stem-like state, obtaining the so-called induced pluripotent stem cells (iPSCs). ESCs and iPSCs were thought to be a great promise: many believed that their discovery would have fuelled an impressive expansion of regenerative medicine, gene therapy and personalised medicine fields in the short term, ultimately leading to the solution of many health-related problems. So far, only a part of this promise has been fulfilled, mainly because of the scarce knowledge we have about the basic biological processes that make these cells so special. In this research, we aim at improving our understanding of the molecular processes that orchestrate pluripotency focusing on beta-catenin (b-catenin), a pivotal protein in stemness maintenance. Indeed, we and others reported a central role of b-catenin in somatic cell reprogramming, differentiation, pluripotency and tissue regeneration in vivo. Still, the mechanisms through which b-catenin controls pluripotency of embryonic stem cells are highly debated.We will unfold this complexity using an interdisciplinary approach. Using cutting-edge technology, we will engineer mouse ESCs (mESCs) in which b-catenin can be modulated in a number of way (amount of protein, temporal dynamics, ability to regulate other genes). Subsequently, we will perform experiments to link specific b-catenin perturbed behaviours to mESCs self-renewal and differentiation ability. Finally, we will use potent computational approaches to understand how b-catenin interacts with other genes important for pluripotency.The project will provide a vital step of innovation and knowledge to the embryonic stem cells biology field: the results will reveal how b-catenin regulates the balance between pluripotency and differentiation, and indicate how to drive specific cell-fates in vitro.In the future, the output of the proposed project could be extended to control b-catenin dependent self-renewal and differentiation of human ESCs and iPSCs, shortening the distances between pluripotent stem cells and their applicative targets. Also, the developed techniques could be used in other biological processes in which b-catenin is involved, including development and cancer.