MicroRNA Regulation of Stem Cell Self-renewal

描述(由申请人提供)： 细胞内存在调节造血干细胞(HSC)自我更新的机制。仅限于正常干细胞的自我更新能力对维持组织完整性是必不可少的，并负责损伤后的再生。然而，当被异常细胞获得时，它可能会产生一种无法治疗的自我更新的癌症干细胞。了解自我更新的机制对于造血干细胞的治疗扩增以及根除血癌的恶性根源具有重要意义。 与信使核糖核酸的表达相比，微核糖核酸的表达可以准确地标志发育的历史和/或阶段。造血干细胞的发育经历了一个特征明确的分步分化过程，并逐渐丧失了自我更新能力。破坏microRNA生物发生关键酶Dester，完全消融了干细胞的活性。因此，我们假设在调节这些关键步骤时需要特定的microRNAs，一些microRNAs的表达可能与自我更新的能力有关。使用了两种互补的方法来识别这种特定的microRNAs。通过扫描已知对造血起重要作用的基因的3‘非编码区，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不更新的直系后代细胞进行了microRNA表达谱分析。根据microRNA表达谱，我们鉴定了miR-99b-let-7e-miR-125a簇，该簇在体内长期移植后显著扩大了造血室。 我建议通过两个特定的目的来详细了解这些microRNA物种在调节HSC自我更新方面的作用。1.利用miR-150基因敲除小鼠确定miR-150在HSC功能中的作用。2.确定miR-99b-let-7e-125a簇在促进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斯卡登博士的实验室随时可用的适当的体内和体外测试来实现。 与公共健康相关：这项拟议的研究承诺发现有助于HSC自我更新的microRNA调节器。这类microRNAs将为HSC的调控提供新的范例。他们还将作为研究工具，发现更多的分子途径和网络，赋予自我更新的表型，不仅是正常的造血干细胞，也是白血病干细胞。

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.