Deciphering the genetically interactive network of the DLK1-DIO3 ncRNA locus in the hematopoietic system and in infant leukemias

许多研究已经确定非编码RNA（ncRNA）-特别是microRNA（miRNAs）-的失调是白血病发生的致病因素。这些研究在很大程度上研究了单个miRNA或长非编码RNA（lncRNA）的功能。然而，由于42%的miRNA基因是以多顺反子转录本的形式组织的，因此研究它们的相互作用对于理解miRNA或miRNA网络如何控制基本的细胞功能和细胞命运决定至关重要。在我们的人类造血系统的ncRNA表达图谱中，我们发现了DLK 1-DIO 3位点的特异性和协调表达，该位点包含人类基因组中最大的miRNA簇（54个miRNA;称为miRNA megacluster），许多box C/D snoRNA和lncRNA-在巨核细胞和巨核细胞白血病中。后一种白血病类型最常发生在婴儿中，因此与发育有很强的联系。作为DFG资助的正在进行的项目的一部分，我们为询问巨核细胞生成和急性巨核细胞白血病（AMKL）发病机制中的DLK 1-DIO 3位点奠定了基础。我们首先阐明了巨核细胞和巨核母细胞的上游调控机制，使我们能够干扰基因座的表达。我们还鉴定了驱动巨核细胞分化的基因座内的miRNA，以及影响巨核细胞生成并控制基因座表达的新型顺式调节长基因间ncRNA（lincRNA; LINC 02285）。此外，我们建立了LNP包装和递送平台，以预期RNA为中心的治疗概念的临床前测试。 在本研究单元的后续项目中，我们将继续研究DLK 1-DIO 3基因座和其中的ncRNA。我们打算继续（1）描绘正常造血和AMKL过程中miRNA大簇内的相互作用网络，包括共表达的lncRNA，（2）解决单个参与者的特征和机制，和（3）涉及这些ncRNA的治疗干预的临床前测试。总的来说，该项目旨在阐明复杂的器官系统如何受到相互作用的遗传网络的调节，以及这些网络的失调如何影响肿瘤发生。

Numerous studies have established the dysregulation of non-coding RNAs (ncRNAs) – in particular of microRNAs (miRNAs) – as a causative factor in leukemogenesis. These studies have largely examined the functions of individual miRNAs or long non-coding RNAs (lncRNAs). However, since 42% of miRNA genes are organized in polycistronic transcripts, studying their interaction is crucial for understanding how miRNAs or miRNA networks control essential cellular functions and cell-fate decisions. In our ncRNA expression atlas of the human hematopoietic system, we uncovered specific and coordinated expression of the DLK1-DIO3 locus – which contains the largest miRNA cluster of the human genome (54 miRNAs; called a miRNA megacluster), numerous box C/D snoRNAs and lncRNAs – in megakaryocytes and megakaryoblastic leukemias. The latter leukemia type occurs most frequently in infants and thus has strong developmental ties. As a part of this ongoing DFG-funded project, we laid a foundation for interrogating the DLK1-DIO3 locus in megakaryopoiesis and acute megakaryoblastic leukemia (AMKL) pathogenesis. We first elucidated an upstream regulatory mechanism in megakaryocytes and megakaryoblasts, allowing us to interfere with expression from the locus. We also identified miRNAs within the locus that drive megakaryocytic differentiation, as well as a novel cis-regulatory long intergenic ncRNA (lincRNA; LINC02285) that affects megakaryopoiesis and controls expression of the locus. Further, we established an LNP packaging and delivery platform in anticipation of the preclinical testing of RNA-centered therapy concepts. In this proposed follow-up project for the Research Unit, we will continue our efforts to dissect the DLK1-DIO3 locus and the ncRNAs within it. We intend to move forward with (1) delineating the network of interactions within the miRNA megacluster – including co-expressed lncRNAs – during normal hematopoiesis and AMKL, (2) resolving the features and mechanisms of individual players, and (3) pre-clinical testing of therapeutic interventions involving these ncRNAs. Overall, this project aims to elucidate how complex organ systems are regulated by interactive genetic networks and how dysregulation of these networks impacts oncogenesis.