The Role of DDX41 in Inherited Myelodysplastic Syndromes

项目摘要 骨髓增生异常综合征（MDS）是由造血干/祖细胞受损引起的遗传性疾病， 细胞（HSPC），其可转化为急性髓性白血病（AML）。MDS通常发生在老年人 然而，MDS也可以表现为年轻个体中遗传性生殖系突变的结果。 DEAD/H-box解旋酶基因DDX 41的突变是与糖尿病相关的最常见的改变之一。 遗传性MDS遗传性DDX 41突变是杂合的，并且通常是移码，这表明 这些突变导致DDX 41功能丧失。DDX 41突变也在新发MDS中观察到， AML，并且通常是错义突变，经常导致氨基酸取代R525 H。DDX 41是 水解ATP的RNA解旋酶，并且可以作为先天免疫传感器、RNA剪接因子发挥作用，以及 核糖体调节因子DDX 41突变改变HSPC功能并促进HSPC功能的确切机制 对MDS/AML的影响仍然未知。因此，拟议的项目将定义DDX 41突变在 MDS/AML的发病机制。模拟在人MDS中观察到的移码突变，并确定其作用 DDX 41在正常造血中的表达，我们产生了造血特异性和条件性Ddx 41缺陷 小鼠我们的初步数据显示，DDX 41缺陷（完全敲除）与 HSPC功能和造血，而DDX 41杂合性增加BM HPSC。另外通过 通过整合蛋白质组学和生物化学方法，我们在白血病细胞中鉴定了一种新的DDX 41相互作用蛋白， 细胞，SAMHD 1，一种dNTPs和宿主防御因子，控制dNTPs的细胞库。总体而言，我们 初步数据表明DDX 41在造血和HSPC功能中具有关键作用。我们假设 DDX 41表达和/或功能的降低导致无效的造血作用， 在MDS和AML的发病机制中，部分原因是SAMHD 1活性增加和细胞dNTP池改变。 因此，本提案的目的是在正常和非正常的人中建立体细胞和生殖系DDX 41突变模型， 恶性造血，并阐明HSPC功能所需的DDX 41的分子功能。 通过广泛的造血方法，我们将确定DDX 41缺陷和R525 H在造血干细胞中的作用。 表达在MDS发病率和向AML进展中的作用（Aim 1）。由于DNMT 3A和DDX 41突变 通常在进展为AML的MDS患者中同时发生，我们将确定DDX 41 杂合性或R525 H表达与DNMT 3A缺陷型小鼠组合将导致高风险MDS或 显性AML细胞dNTP池减少损害细胞周期进程并导致基因组不稳定; 因此，我们将确定SAMHD 1活性在DDX 41缺陷型HSPC或突变型MDS中的作用， 减少的细胞dNTPs库是否导致基因组不稳定性和AML的发展（Aim 2）。通过 阐明DDX 41的恶性造血功能，我们预计将发现新的机制， 潜在的遗传性和新发MDS。

Project Summary Myelodysplastic syndromes (MDS) are genetic disorders caused by impaired hematopoietic stem/progenitor cells (HSPC), which can transform to acute myeloid leukemia (AML). MDS typically occurs in the ageing population, however MDS can also manifest as a result of inherited germline mutations in younger individuals. Mutations in the DEAD/H-box helicase gene DDX41 are among the most common alterations associated with inherited MDS. Inherited DDX41 mutations are heterozygous and are typically frameshifts, suggesting that these mutations result in loss of DDX41 function. DDX41 mutations are also observed in de novo MDS and AML, and are typically missense mutations frequently resulting in the amino acid substitution R525H. DDX41 is an RNA helicase that hydrolyzes ATP, and can function as an innate immune sensor, RNA splicing factor, and ribosome regulator. The precise mechanism(s) by which DDX41 mutations alter HSPC function and contribute to MDS/AML remains unknown. As such, the proposed project will define the role of DDX41 mutations in the pathogenesis of MDS/AML. To mimic the frameshift mutations observed in human MDS and determine the role of DDX41 in normal hematopoiesis, we generated hematopoietic-specific and conditional Ddx41-deficient mice. Our preliminary data has revealed that DDX41-deficiency (complete knockout) is not compatible with HSPC function and hematopoiesis, whereas DDX41 heterozygosity increases BM HPSC. In addition, through integrative proteomic and biochemical approaches we identified a novel DDX41-interacting protein in leukemic cells, SAMHD1, a dNTPase and host defense factor, which controls cellular pools of dNTPs. Collectively, our preliminary data indicate that DDX41 has a critical role in hematopoiesis and HSPC function. We hypothesize that diminished DDX41 expression and/or function contributes to ineffective hematopoiesis and to the pathogenesis of MDS and AML, in part due to increased SAMHD1 activity and altered cellular dNTP pools. Therefore, the objectives of this proposal are to model somatic and germline DDX41 mutations in normal and malignant hematopoiesis, and to elucidate the molecular function of DDX41 required for HSPC function. Through extensive hematopoietic approaches, we will define the role of DDX41 deficiency and R525H expression in MDS incidence and progression to AML (Aim 1). Since DNMT3A and DDX41 mutations commonly co-occur in MDS patients that have progressed to AML, we will determine whether DDX41 heterozygosity or R525H expression combined with DNMT3A-deficient mice will result in high-risk MDS or overt AML. Reduced cellular dNTP pools impair cell cycle progression and result in genomic instability; therefore, we will determine the role of SAMHD1 activity in DDX41-deficient HSPC or mutant MDS, and whether diminished cellular dNTPs pools lead to genomic instability and development of AML (Aim 2). By elucidating the function of DDX41 malignant hematopoiesis, we predict to uncover novel mechanisms underlying inherited and de novo MDS.