The Novel Role and Mechanism of RBM33 in Leukemogenesis

摘要 急性髓系白血病（acute myeloid leukemia，AML）是最常见和最致命的恶性血液病之一 是由基因突变和基因组重排引起的。AML患者的治愈率没有 几十年来有了很大的改善。AML发病机制的分子机制不是 完全理解通过使用新的机器学习方法分析公开的基因组数据， 结合基序蛋白33（RBM 33）是一种RNA结合蛋白，被鉴定为AML的必需基因。 然而，RBM 33的生物学功能尚不清楚。我们的初步研究提供了第一个引人注目的 证据表明RBM 33在调节m6 A RNA去甲基化中具有新功能。更重要的是我们 结果显示，RBM 33敲低显著抑制了人和小鼠白血病细胞的生长和存活， 细胞在分子水平上，我们已经确定了白血病细胞中RBM 33的潜在下游靶点。 ALKBH 5被称为m6 A mRNA去甲基化酶（Eraser），可从RNA中去除m6 A甲基化基团。 迄今为止，仍然不清楚是否需要另一种RNA结合蛋白来确保 将ALKBH 5募集至其mRNA靶点。我们最近已经证明ALKBH 5在以下方面具有关键作用： AML开发和维护。我们假设RBM 33在糖尿病的发病机制中起关键作用。 AML通过调节ALKBH 5介导的m6 A去甲基化。为了验证这一假设，我们将研究三个具体的问题。 目标。在Aim 1中，我们将确定RBM 33在调节动态m6 A RNA甲基化中的新作用， 白血病细胞在Aim 2中，我们将研究Rbm 33在AML发展和维持中的作用。在目标3中， 我们将确定在AML的白血病发生中介导RBM 33功能的下游通路。 我们将采用小鼠遗传模型以及人类患者来源的小鼠模型来阐明 RBM 33在体内正常造血和白血病发生中的作用，并将联合收割机转录组和 表位转录组分析，以确定关键的下游靶点和相关的下游途径， 介导RBM 33在白血病发生中的作用。我们的研究将揭示RBM 33在m6 A RNA中的新作用 修改，并定义RBM 33在AML发展中的重要性和潜在机制， LSC/LIC自我更新。因此，我们项目的成功将大大推动我们的 理解m6 A修饰介导的基因调控的复杂机制， 白血病细胞和m6 A RNA去甲基化在白血病发生中的关键作用。

Abstract Acute myeloid leukemia (AML) is one of the most common and fatal forms of hematological malignancies caused by gene mutations and genomic rearrangements. The cure rates for AML patients have not significantly improved for decades. The molecular mechanisms underlying the pathogenesis of AML are not fully understood. By analysis of publicly available genomic data using a new machine learning approach, RNA Binding Motif Protein 33 (RBM33), an RNA binding protein, is identified as an essential gene in AML. However, the biological function of RBM33 is unknown yet. Our preliminary studies provide the first compelling evidence suggesting a novel function of RBM33 in regulating m6A RNA demethylation. More importantly, we showed that RBM33 knockdown significantly inhibited growth and survival of human and mouse leukemia cells. At a molecular level, we have identified a potential downstream target of RBM33 in leukemia cells. ALKBH5 is known as an m6A mRNA demethylase (Eraser), which removes m6A methylated groups from RNA. To date, it remains unknown whether another member of RNA binding proteins is required for ensuring recruitment of ALKBH5 to its mRNA targets. We have recently demonstrated that ALKBH5 has a critical role in AML development and maintenance. We hypothesize that RBM33 plays a critical role in the pathogenesis of AML by regulating ALKBH5-mediated m6A demethylation. To test this hypothesis, we will pursue three specific aims. In Aim1, we will determine a novel role of RBM33 in regulation of dynamic m6A RNA methylation in leukemia cells. In Aim2, we will investigate the role of Rbm33 in AML development and maintenance. In Aim 3, we will determine the downstream pathway that mediates the function of RBM33 in leukemogenesis in AML. We will employ both mouse genetic models as well as human patient-derived mouse models to elucidate the role of RBM33 in normal hematopoiesis and leukemogenesis in vivo, and will combine transcriptome and epitranscriptome analysis to identify the key downstream targets and associated downstream pathways that mediate the role of RBM33 in leukemogenesis. Our studies will uncover a novel role of RBM33 in m6A RNA modification, and define the importance and underlying mechanisms of RBM33 in AML development and maintenance as well as LSC/LIC self-renewal. Thus, the success of our project will significantly advance our understanding of the complex mechanisms underlying the m6A modification-mediated gene regulation in leukemia cells and the critical role of m6A RNA demethylation in leukemogenesis.