Mechanism of action of Prdm16 in hematopoietic stem cells

描述(申请人提供)：造血干细胞(HSCs)可以自我更新并生成所有造血系统的谱系。尽管我们对涉及造血干细胞自我更新、分化和静止的机制的了解取得了重大进展，但关于这些机制如何协同作用来调节体内造血系统的动态平衡反应的一致图景尚未出现。作为白血病融合蛋白的合作伙伴，几种转录调控因子在HSC生物学中发挥着关键作用。我们利用Prdm16-/-小鼠发现，Prdm16是一种140 kDa的锌指蛋白，最初被发现是急性髓细胞白血病(AML)某些易位的融合伙伴，在发育过程中和移植后对HSCs的建立和维持是必不可少的。在这项研究中，我们希望进一步分析Prdm16在造血干细胞生物学中的功能，并确定其作用机制。我们的初步数据强烈表明Prdm16在维持线粒体功能和完整性方面发挥作用，特别是在HSCs中。PRDM16以两种剪接形式出现。Short(S)Prdm16缺乏N端PR结构域。这种剪接变异体通常通过易位、启动子低甲基化或逆转录病毒插入PR结构域而在白血病中过度表达。我们的初步数据表明，sPrdm16主要是Prdm16缺陷小鼠的造血表型的原因。具体目标如下：目标1：确定HSC功能所需Prdm16的关键发育窗口；目标2：检测Prdm16-/-HSCs和MEF的代谢、线粒体功能和动力学；目标3：比较sPrdm16和flPrdm16在HSC维持中的作用。

DESCRIPTION (provided by applicant): Hematopoietic stem cells (HSCs) can self renew and generate all lineages of the hematopoietic system. Despite significant progress in our understanding of mechanisms involved in self-renewal, differentiation and quiescence of HSCs, a coherent picture of how these mechanisms act in concert to regulate homeostatic responses of the hematopoietic system in vivo has not emerged yet. Several transcriptional regulators involved as partners of leukemogenic fusion proteins play a critical role in HSC biology. We found, using germline Prdm16-/- mice, that Prdm16, a 140kDa zinc finger protein that was originally discovered as a fusion partner in some translocations in acute myeloblastic leukemia (AML), is essential for the establishment and maintenance of HSCs during development and after transplantation. In this proposal we want to further analyze the function of Prdm16 in the biology of HSCs, and determine its mechanism of action. Our preliminary data strongly suggest a role for Prdm16 in maintaining mitochondrial function and integrity specifically in HSCs. Prdm16 occurs in two splice forms. Short (s) Prdm16 lacks the N-terminal PR domain. This splice variant is typically overexpressed in leukemias through translocation, promoter hypomethylation, or retroviral insertion into the PR domain. Our preliminary data indicate that primarily sPrdm16 is responsible for the hematopoietic phenotype of Prdm16-deficient mice. The specific aims are the following: Aim 1: To determine the critical developmental window of Prdm16 requirement for HSC function; Aim 2: To examine metabolism, mitochondrial function, and dynamics in Prdm16-/- HSCs and MEFs; Aim 3: To examine the roles of sPrdm16 vs. flPrdm16 in HSC maintenance.