The functional role of snoRNAs in NPM1 wildtype and mutant AML

NPM1突变是急性髓性白血病（AML）中最常见的突变之一。NPM1是一种参与核糖体生物发生和基因组稳定性的多功能核仁伴侣。NPM1的突变在c端区域产生一个新的核输出信号，并导致突变的NPM1 （NPM1c）的异常胞质错位。基于其多效性功能，NPM1基因改变在白血病发生中的作用机制已被提出。然而，npm1突变AML的病理机制尚不完全清楚。包括microRNA （miRNA）在内的小非编码rna （ncrna）已被证明对各种生理和细胞过程产生影响。我们之前发现C/D盒小核核rna （snoRNA）是白血病干细胞活性的关键调节因子和AML1-ETO诱导的白血病发生的重要介质。为了评估C/D盒snoRNA在其他形式AML中的潜在作用，我们在两个独立的患者队列中通过高通量小RNA-Seq分析了snoRNA的表达。我们的初步数据显示，在两组C/D盒中，snoRNAs在NPM1野生型AML中高表达，而在NPM1突变的AML标本中表达低得多。值得注意的是，我们的质谱数据显示野生型NPM1与snoRNA表达调控因子DDX21和AES之间存在直接的蛋白蛋白相互作用。因此，我们的初步研究结果为NPM1在snoRNA表达中的调节作用提供了证据。为了进一步探讨snoRNA在NPM1野生型和突变型AML中的功能作用，我们建议：1)明确NPM1调控snoRNA的机制，并鉴定野生型和突变型NPM1的蛋白和RNA相互作用组；2)在携带NPM1突变与野生型的AML细胞中，研究snoRNA解除管制对核糖体生物发生和功能的功能影响；3)评估snoRNA下调在NPM1c AML对化疗的有利反应中的潜在作用，以及snoRNA抑制在AML对化疗增敏中的作用。这些数据将提高我们对npm1突变的AML病理的理解，并进一步揭示C/D盒snoRNA通路在异常造血中的重要作用。

NPM1 mutations rank among the most frequent mutations in acute myeloid leukemia (AML). NPM1 is a multifunctional nucleolar chaperone involved in ribosome biogenesis and genomic stability. Mutations in NPM1 generate a novel nuclear export signal in the C-terminal domain and lead to aberrant cytoplasmic dislocation of mutated NPM1 (NPM1c). Based on its pleiotropic functions, several mechanisms have been proposed for the contribution of NPM1 gene alterations to leukemogenesis. However, the pathology of NPM1-mutated AML remains incompletely understood.Small non-coding RNAs (ncRNAs) including microRNA (miRNA) have been shown to impact onto various physiological and cellular processes. We previously identified C/D box small nucleolar RNAs (snoRNA) as crucial regulators of leukemia stem cell activity and essential mediator of AML1-ETO induced leukemogenesis. To evaluate a potential role of C/D box snoRNAs in other forms of AML, we profiled snoRNA expression by high throughput small RNA-Seq in two independent cohorts of patients. Our preliminary data shows that in both sets C/D box snoRNAs are highly expressed in NPM1 wildtype AML whereas expression are much lower in AML specimens with NPM1 mutations. Of note, our mass spectrometry data demonstrates a direct protein-protein interaction between wildtype NPM1 and the snoRNA expression regulators DDX21 and AES. Thus, our preliminary findings provide evidence for a regulatory role of NPM1 in snoRNA expression. To further explore the functional role of snoRNAs in NPM1 wildtype and mutant AML, we propose to 1) define the mechanistic role of NPM1 for snoRNA regulation and to identify the protein and RNA interactome of wildtype and mutant NPM1; 2) investigate the functional consequences of snoRNA deregulation on ribosome biogenesis and function in AML cells harboring NPM1 mutation versus wildtype; 3) evaluate the potential role of snoRNA downregulation in the favorable response of NPM1c AML to chemotherapy and the utility of snoRNA suppression in sensitizing AML to chemotherapy. Together these data will improve our understanding of pathology of NPM1-mutated AML, and further uncover the essential role of C/D box snoRNA pathway in aberrant hematopoiesis.