Uncovering the Role of the DEAD Box Helicase Ddx41 in Hematopoiesis

摘要 骨髓增生异常综合征（MDS），一组恶性前骨髓衰竭综合征， 造血干细胞（HSC）的缺陷是人类最常见的血液恶性肿瘤之一。 约30%的老年人进展为急性髓性白血病（AML）。最近，功能丧失的生殖细胞和 已经在MDS患者中鉴定了DEAD盒解旋酶41基因（DDX 41）的体细胞突变， AML，并被认为有助于疾病的发病机制。生殖系和体细胞DDX 41突变都是 被认为促进造血失调和白血病发生。虽然有很强的临床相关性， 虽然在DDX 41和MDS中的突变之间发现了DDX 41，但DDX 41在造血中的体内作用尚未被证实。 阐明。为了解决这个问题，我们将描述斑马鱼ddx 41功能丧失的造血作用。 变种人我们的初步研究表明ddx 41突变体发生中性粒细胞减少和贫血。这种动物 该模型将使我们能够阐明Ddx 41在正常造血中的体内作用和潜在的造血功能。 任何缺陷的机制。DDX 41与剪接有关，并显示与组件相关。 包括剪接因子3B，亚基1（SF 3B 1），最常见的突变剪接因子 在MDS。此外，具有DDX 41突变的MDS/AML患者样本显示mRNA剪接错误， 通常发生在剪接体的组分有缺陷时。尽管这些数据表明 对于DDX 41的剪接，尚不清楚剪接畸变是否导致了观察到的异常， DDX 41突变的血液病。使用ddx 41缺陷的斑马鱼将使我们能够描绘出 当ddx 41突变时，功能相关的早期分子事件导致血细胞缺陷， 有望为MDS的起源提供新的见解。在这个提案中，我们将在体内使用我们的小说 Ddx 41缺乏模型，以检验Ddx 41通过调节对正常造血至关重要的假设 剪接，这一过程的故障有助于DDX 41- 突变的MDS/AML。在目标1中，我们将确定哪些血液群体需要Ddx 41，哪些功能性 Ddx 41的结构域是健康造血所必需的。在目标2中，我们将阐明 DDX 41和SF 3B 1的蛋白质和遗传相互作用研究。结合这些研究， DDX 41突变的MDS/AML的病理生理学。

Abstract Myelodysplastic syndromes (MDS), a group of pre-malignant bone marrow failure syndromes arising from defects in hematopoietic stem cells (HSCs), are amongst the most common hematological malignancies of the elderly with ~30% progressing to acute myeloid leukemia (AML). Recently, loss-of-function germline and somatic mutations in the DEAD-box Helicase 41 gene (DDX41) have been identified in patients with MDS and AML, and are thought to contribute to disease pathogenesis. Both germline and somatic DDX41 mutations are thought to promote hematopoietic deregulation and leukemogenesis. Although a strong clinical correlation is found between mutations in DDX41 and MDS, the in vivo role of DDX41 in hematopoiesis has not been elucidated. To address this question, we will characterize hematopoiesis in a zebrafish ddx41 loss-of-function mutant. Our preliminary studies indicate that ddx41 mutants develop neutropenia and anemia. This animal model will allow us to elucidate the in vivo role of Ddx41 in normal hematopoiesis and the underlying mechanism for any defects. DDX41 has been implicated in splicing and shown to associate with components of the spliceosome including Splicing Factor 3B, subunit 1 (SF3B1), the most commonly mutated splicing factor in MDS. Additionally, MDS/AML patient samples with DDX41 mutations displayed errors in mRNA splicing that typically occur when components of the spliceosome are defective. Although these data point towards a role for DDX41 in splicing, it is still unknown if splicing aberrations contribute to the observed abnormalities in DDX41-mutated hematologic diseases. Using ddx41-deficient zebrafish will permit us to delineate the functionally relevant early molecular events leading to blood cell defects when ddx41 is mutated, which is anticipated to provide novel insight into the origins of MDS. In this proposal, we will use our novel in vivo model of Ddx41-deficiency to test the hypothesis that Ddx41 is critical for normal hematopoiesis via regulation of splicing, and that malfunctioning of this process contributes to the hematological defects seen in DDX41- mutated MDS/AML. In Aim 1, we will determine which blood populations require Ddx41 and which functional domains of Ddx41 are required for healthy hematopoiesis. In Aim 2, we will elucidate connections between DDX41 and SF3B1 using protein and genetic interaction studies. Combined these studies will reveal insights into the pathophysiology of DDX41-mutated MDS/AML.