MicroRNA Regulation of Stem Cell Self-renewal

描述（由申请人提供）： 造血干细胞（HSC）的自我更新是由细胞内在机制调控的。局限于正常干细胞的自我更新能力对于维持组织完整性至关重要，并且负责损伤后的再生。然而，当被异常细胞获得时，它可以产生一种自我更新的癌症干细胞，这种干细胞对治疗很难。了解自我更新的机制对于HSC的治疗扩展以及根除血癌的恶性根源是重要的。 与mRNA表达相比，microRNA表达忠实地标记了发育历史和/或阶段。HSC通过充分表征的逐步分化过程发育，逐渐丧失自我更新能力。破坏microRNA生物发生的关键酶Dicer，完全消除了干细胞的活性。因此，我们假设特定的microRNA是调节这些关键步骤所必需的，并且一些microRNA的表达可能与自我更新的能力相关。两种互补的方法被用来鉴定这种特定的microRNA。通过扫描已知对造血重要的基因的3 'UTR区域，我们确定miR-150通过靶向c-Myb发挥其功能。此外，使用原代干细胞及其非更新的直接后代细胞进行microRNA表达谱分析。基于microRNA表达谱，我们鉴定了miR-99 b-let-7 e-miR-125 a簇，其在体内长期移植后显著扩增造血区室。 我建议通过两个特定的目标来详细了解这些microRNA在调节HSC自我更新中的作用。1.使用miR-150敲除小鼠确定miR-150在HSC功能中的作用。2.确定miR-99 b-let-7 e-125 a簇在增强HSC自我更新中的作用，并评估其候选分子靶点。这些目标将通过结合使用基因工程小鼠品系和适当的体内和体外试验来实现，这些试验在Scadden博士的实验室中很容易获得。 公共卫生相关性：拟议的研究有望发现HSC自我更新的microRNA调节剂。这些microRNA将为HSC调控提供新的范例。它们还将作为研究工具，揭示赋予自我更新表型的其他分子途径和网络，不仅对正常HSC，而且对白血病干细胞。

DESCRIPTION (provided by applicant): Cell intrinsic mechanisms exist to regulate hematopoietic stem cell (HSC) self-renewal. Self-renewal ability confined to the normal stem cells is essential for maintaining tissue integrity and is responsible for regeneration after injury. When acquired by aberrant cells, however, it could produce a self-renewing cancer stem cell that is refractory to treatments. Understanding the mechanisms of self-renewal is important for therapeutic expansion of HSCs as well as for eradicating the malignant roots of blood cancers. Contrasting to mRNA expression, microRNA expression faithfully marks developmental history and/or stage. HSCs develop through a well characterized step-wise differentiation process with a gradual loss of the self-renewing capacity. Disruption of the microRNA biogenesis key enzyme, Dicer, completely ablated stem cell activity. We thus hypothesized that specific microRNAs are required in regulating these key steps and the expression of some microRNAs may correlate with the ability to self-renew. Two complementary approaches were used to identify such specific microRNAs. By scanning the 3'UTR regions of genes known to be important for hematopoiesis, we identified that miR-150 exerted its function by targeting c-Myb. Additionally, microRNA expression profiling was performed using primary stem cells and their non-renewing immediate descendant cells. Based on the microRNA expression profile, we identified the miR-99b-let-7e-miR-125a cluster which dramatically expanded the hematopoietic compartment after long term transplantation in vivo. I propose to gain detailed understanding of these microRNA species in regulating HSC self-renewal through two specific aims. 1. Determine the role of miR-150 in HSC function using miR-150 knockout mice. 2. Define the role of the miR-99b-let-7e-125a cluster in enhancing HSC self-renewal and assess its candidate molecular targets. These aims will be achieved through the combinational use of genetically engineered mouse strains and the appropriate in vivo and in vitro assays readily available in Dr. Scadden's laboratory. PUBLIC HEALTH RELEVANCE: The proposed study promises the discovery of microRNA regulators for HSC self-renewal. Such microRNAs will provide new paradigms for HSC regulation. They will also serve as research tools to uncover additional molecular pathways and networks that confer the self-renewal phenotype, not only to normal HSCs, but also to leukemia stem cells.