NPM1 regulation of 2'-O-methylation in hematopoiesis and bone marrow failure disorder

1 R 01 DK 115536 -01修订版 遗传性骨髓衰竭疾病（BMFD）代表了由基因突变驱动的造血系统疾病的一个独特类别。在这些疾病中，已经确定了一组独特的基因，这些基因导致了称为核糖体病的BMFD子集。核糖体病对在核糖体加工和核糖体生物发生中起关键作用的基因具有突变。我们以前建立了DKC 1基因作为核糖体病基因，通过其调节适当翻译的能力，作为其功能的结果，将核糖体RNA（rRNA）上的尿苷残基转化为假尿苷。令人兴奋的是，我们实验室的初步数据现在表明，rRNA的另一种转录后修饰，2 '-O-甲基化（2'-O-Me）也有助于核糖体功能的适当调节。我们的数据显示，特定的C/D盒小核仁RNA（snoRNA）控制适当的IRES依赖的翻译的主要细胞周期和凋亡基因。在体内，成年小鼠造血干细胞（HSC）区室中2 '-O-Me的破坏导致核糖体病的特征，例如由于退出静止、凋亡和骨髓增生异常骨髓衰竭而导致的干细胞维持缺陷。重要的是，我们确定了与C/D盒snoRNA功能相关的先天性角化不良（DC）患者的新生殖系突变。因此，我们的初步研究结果提供了直接的遗传证据，核糖体专业化通过rRNA 2 '-O-Me，以及在多种疾病状态的发病机制，通过异常的HSC和核糖体功能的关键作用。为了更好地了解2 '-O-Me的作用和功能，我们建议（1）研究受2'-O-Me影响并导致骨髓衰竭过程的分子和细胞途径，（2）确定正常核糖体功能对造血的体内意义，以及通过开发基因工程小鼠模型识别的新型DC突变，和（3）评估参与催化2 ′-O-Me的生物化学复合物的基因在BMFD中可能改变的程度，并确定BMFD中是否存在靶向rRNA甲基化过程的其他新的致病性突变，包括snoRNA。这些数据将进一步促进我们对核糖体病的理解，并有助于揭示核糖体的重要调控如何通过rRNA修饰促进正常和异常的造血。

1R01 DK115536-01 revised Inherited bone marrow failure disorders (BMFDs) represent a distinct category of hematopoietic disorders that are driven by genetic mutation. Within these disorders, a distinct set of genes has already been identified that contribute to a sub-set of BMFDs known as ribosomopathies. Ribosomopathies harbor mutation to genes playing a critical role in ribosomal processing and ribosome biogenesis. We previously established the DKC1 gene as a ribosomopathy gene through its ability to regulate proper translation as a result of its function to convert uridine residues on ribosomal RNAs (rRNAs) to pseudouridine. Exciting, preliminary data from our lab now demonstrates that another post-transcriptional modification of rRNAs, 2’-O-methylation (2’-O-Me), also contribute to proper regulation of ribosome function. Our data reveal that specific C/D-box small nucleolar RNAs (snoRNAs) controls proper IRES-dependent translation of major cell cycle and apoptosis genes. In vivo, disruption of 2’-O-Me in adult mouse hematopoietic stem cell (HSC) compartment results in features characteristic of ribosomopathies such as defects in stem cell maintenance due to exit from quiescence, apoptosis, and myelodysplastic bone marrow failure. Importantly, we identify novel germ line mutations dyskeratosis congenita (DC) patients related to function of C/D box snoRNAs. Thus, our preliminary findings provide direct genetic evidence for the critical role ribosome specialization through rRNA 2’-O-Me, as well as in the pathogenesis of multiple disease states through aberrant HSC and ribosome function. In order to better understand the role and function 2’-O-Me, we propose to (1) study the molecular and cellular pathways that are impacted by 2’-O-Me and which contribute to the process of bone marrow failure, (2) determine the in vivo significance of normal ribosome function for hematopoiesis, and of novel DC mutations identified through development of genetically engineered mouse models, and (3) evaluate the extent to which genes involved in the biochemical complexes that catalyzes 2’-O-Me may be altered in BMFDs, and to determine if additional novel pathogenic mutations targeting the process of rRNA methylation, including snoRNAs, exist in BMFDs. Together these data will further facilitate our understanding of ribosomopathies, and help uncover how essential regulation of the ribosome, through rRNA modification, contributes to normal and aberrant hematopoiesis.