Defining regulators of hematopoietic stem cell self-renewal and lineage potential

建议书摘要 我的建议的目的是了解调节人的自我更新和血统潜力的机制 造血干细胞(HSCs)以及如何通过操纵HSCs来控制HSC的命运和功能 表观遗传参数。我的研究背景是独一无二的，因为我们发现了一个胎儿， 发育受限的HSC(DrHSC)具有不寻常的特性：drHSC能够长期、 连续移植后的多系重建(LTMR)，但在正常情况下不会持续到成年期 发展。因此，drHSCs的自我更新和持续能力是在移植后诱导的。 此外，尽管能够产生迄今为止研究过的所有“传统”造血细胞类型，但 DRHSC偏重于淋巴，与共存的B1a细胞相比，具有更好的B1a细胞重建能力 胎肝(FL)HSC。令人惊讶的是，淋巴偏向和B1a容量连续几个月保持不变 移植实验。定义功能性HSC的两个核心属性-自我更新和 血统潜力-因此在drHSCs中是唯一受调控的。我将利用这些独特的特性来 了解控制HSC命运和功能的分子和表观遗传机制，以及这些机制是如何 机制既是稳定的(如drHSC谱系潜力的情况)，又是动态的(如 诱导持久性)。我还将利用我的能力分离出三种不同的HSC群体： 胎儿HSCs(FHSCs)和成人HSCs(AHSCs)。总之，这将使我能够问 从独特的角度和新的策略研究HSC生物学的基本问题：长期是如何 移植后诱导的drHSCs的持久性，而它们的谱系偏见仍被保留？是血统吗？ 只有一种潜在的功能丧失现象，或者成年的HSCs是否有分化能力 造血干细胞缺乏？如果是这样的话，这些是如何获得的？如何将一组HSC重新编程为另一组HSC 亚型？我建议通过评估表观遗传学和转录组动力学来探讨这些问题(目标1)， 并通过使用CRISPRi小鼠模型在功能上操纵HSC电位(目标2)。由此产生的结果 建议为HSC的长期操作提供基本的概念验证实验 目的是应用这些工具来提高干细胞移植疗法的疗效。

PROPOSAL SUMMARY The goal of my proposal is to understand the mechanisms regulating the self-renewal and lineage potential of hematopoietic stem cells (HSCs) and how HSC fate and function can be controlled by manipulation of epigenetic parameters. The context of my study is unique, because we have discovered a fetal, developmentally restricted HSC (drHSC) with unusual properties: the drHSCs are capable of long-term, multilineage reconstitution (LTMR) upon serial transplantation, but do not persist into adulthood during normal development. The ability of drHSCs to self-renew and persist is therefore induced upon transplantation. Further, while capable of generating all the “traditional” hematopoietic cell types investigated to date, the drHSCs are lymphoid biased and have superior B1a cell reconstitution capacity compared to the co-existing fetal liver (FL) HSCs. Amazingly, the lymphoid bias and B1a capacity are retained over many months in serial transplantation experiments. The two core properties that define functional HSCs – self-renewal and lineage potential – are therefore uniquely regulated in drHSCs. I will leverage these unique properties to understand the molecular and epigenetic mechanisms that govern HSC fate and function, and how these mechanisms are both stable (as in the case of drHSC lineage potential) and dynamic (as in the case of induced persistence). I will also take advantage of my ability to isolate three distinct populations of HSCs: the drHSCs, co-existing fetal HSCs (fHSCs), and adult HSCs (aHSCs). Together, this will enable me to ask fundamental questions in HSC biology from a unique perspective and with novel strategies: How is long-term persistence of drHSCs induced upon transplantation, whereas their lineage bias is retained? Is lineage potential exclusively a loss-of-function phenomenon or do adult HSCs have differentiation capabilities that fetal HSCs lack? If so, how are these gained? How can one HSC population be “reprogrammed” into another HSC subtype? I propose to pursue these questions by assessing epigenetic and transcriptome dynamics (Aim 1), and by functionally manipulating HSC potential using CRISPRi mouse models (Aim 2). Outcomes from this proposal will be provide fundamental proof-of-concept experiments for HSC manipulation with the long-term goal of applying these tools to increase the efficacy of stem cell transplant therapies.