The role of ALKBH5-mediated RNA demethylation in the maintenance of genomic stability in HSPCs

ALKBH5 介导的 RNA 去甲基化在维持 HSPC 基因组稳定性中的作用

• 批准号: 10669161
• 负责人: Zhijian Qian
• 金额: $ 47.85万
• 依托单位: UNIVERSITY OF FLORIDA
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-01 至 2026-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10669161
• 关键词: Aging; Apoptosis; Biological Process; CD34 gene; Cell Death; Cell Maintenance; Cells; Chromatin; DNA Damage; DNA Repair; DNA lesion; Development; Dysmyelopoietic Syndromes; Functional disorder; Gene Expression; Gene Expression Regulation; Genome Stability; Genomic Instability; Goals; Growth; Hematological Disease; Hematopoietic; Hematopoietic stem cells; Heterozygote; Histone Deacetylase; Human; Human Cell Line; In Vitro; Induction of Apoptosis; Long-Term Effects; Maintenance; Malignant - descriptor; Malignant Neoplasms; Mediating; Mediator; Messenger RNA; Methylation; Modeling; Modification; Molecular; Mus; Mutagenesis; Oncogene Activation; Oxidative Stress; Pathway interactions; Patients; Phenocopy; Physiological; Play; Polyribosomes; Post-Transcriptional Regulation; Post-Translational Protein Processing; Preleukemia; Process; RNA; RNA methylation; Reactive Inhibition; Reactive Oxygen Species; Ribosomes; Role; Source; Speed; Stress; System; Testing; Transgenic Mice; Transgenic Organisms; Up-Regulation; Xenograft procedure; cell injury; crosslinking and immunoprecipitation sequencing; demethylation; epigenetic regulation; genome integrity; hematopoietic stem cell expansion; hematopoietic stem cell quiescence; hematopoietic stem cell self-renewal; in vivo; insight; knock-down; leukemic stem cell; leukemic transformation; novel; novel therapeutic intervention; overexpression; posttranscriptional; prevent; public database; repaired; response; self-renewal; stem cell function; stem cells; transcriptome sequencing; tumorigenesis

Abstract Myelodysplastic syndromes (MDS) are a group of diverse malignant hematological disorders that originate from hematopoietic stem cells (HSCs). Increased levels of reactive oxygen species (ROS) and DNA damage are commonly detected in hematopoietic cells from MDS patients. An elevated level of ROS, generated from either endogenous or exogenous sources including oncogene activation, leads to loss of quiescence and self-renewal of HSCs. ROS-induced DNA damage speeds up the aging process of stem cells and contributes to the mutagenesis associated with cancer development. m6A RNA methylation plays a significant role in multiple biological processes by introducing another layer of post-transcriptional regulation of gene expression within cells. The goal of this project is to elucidate the significant role of ALKBH5-mediated epigenetic regulation in the maintenance of genomic stability in hematopoietic stem/progenitor cell (HSPCs) during oxidative stress, and how deregulation of ALKBH5 contributes to promotion of leukemic transformation of HSPCs in the initiation and development of MDS. We found that ROS significantly increased global m6A RNA methylation in human cell lines, and that the elevation of m6A mRNA methylation is required for rapidly repairing ROS-induced DNA lesions and preventing cell death. Interestingly, we found that ALKBH5, the m6A RNA demethylase, is responsible for ROS-induced elevation of m6A mRNA methylation. ROS induced post- translational modification of ALKBH5, and inhibited the demethylase activity of ALKBH5. We showed that forced expression of ALKBH5 inhibited ROS-induced m6A mRNA methylation and significantly delayed repair of ROS-induced DNA damage. Thus, we hypothesize that aberrant expression of ALKBH5 disrupts HSPC functions by negatively influencing genome integrity and survival of HSPCs, thereby contributing to leukemic transformation of HSPCs during the initiation and development of MDS. In this proposal, we will determine 1) the role and underlying mechanism of ALKBH5 in the maintenance of genomic stability in HSPCs in response to oxidative stress; 2) the effects of ALKBH5/Alkbh5 overexpression on the maintenance of mouse and human primary HSPCs during ROS stress in vivo; and 3) whether ALKBH5/Alkbh5 is required for the maintenance of pre-leukemic stem cells (pre-LSCs) in MDS. Our study will provide new insights into novel mechanisms of MDS development and epitranscriptional regulation of gene expression in HSPCs in response to oxidative stress. Additionally, our study will provide the first set of evidence to support a significant role of ALKBH5- mediated m6A mRNA demethylation in the maintenance of normal HSPCs and pre-leukemic stem cell (pre- LSCs).

抽象的 骨髓增生综合征（MDS）是一组多种恶性血液学疾病 起源于造血干细胞（HSC）。活性氧（ROS）和DNA的水平增加 MDS患者的造血细胞中通常检测到损伤。升高的ROS， 由内源性或外源性来源（包括癌基因激活）产生的，导致 HSC的静止和自我更新。 ROS诱导的DNA损伤加快了干细胞的衰老过程 并有助于与癌症发展相关的诱变。 M6a RNA甲基化播放A 通过引入另一层转录后调节，在多个生物过程中的重要作用 细胞内的基因表达。该项目的目的是阐明ALKBH5介导的重要作用 造血茎/祖细胞（HSPCS）维持基因组稳定性的表观遗传调节 在氧化应激期间，以及ALKBH5的放松管制如何促进白血病转化 HSPC在MDS的启动和开发中。我们发现ROS大大增加了全球M6A 人类细胞系中的RNA甲基化，并且需要快速需要M6a mRNA甲基化的升高 修复ROS诱导的DNA病变并防止细胞死亡。有趣的是，我们发现AlkBH5，M6A RNA脱甲基酶负责ROS诱导的M6a mRNA甲基化升高。 ROS引起的后 ALKBH5的翻译修饰，并抑制ALKBH5的脱甲基酶活性。我们表明了这一点 ALKBH5强迫表达抑制ROS诱导的M6a mRNA甲基化，并显着延迟修复 ROS诱导的DNA损伤。因此，我们假设ALKBH5的异常表达破坏了HSPC 通过负面影响HSPC的基因组完整性和生存的功能，从而有助于白血病 HSPC在MDS的启动和开发过程中的转化。在此提案中，我们将确定1） ALKBH5在HSPC中维持基因组稳定性中的作用和潜在机制。 氧化应激； 2）ALKBH5/ALKBH5过表达对维持小鼠和人的影响 在体内ROS胁迫期间的一级HSPC； 3）维护是否需要ALKBH5/ALKBH5 MDS中的白细胞前干细胞（前LSC）。我们的研究将提供有关新型机制的新见解 氧化作用的MDS发育和HSPC中基因表达的表达调节 压力。此外，我们的研究将提供第一组证据，以支持ALKBH5-的重要作用。 介导的M6a mRNA脱甲基在维持正常HSPC和白细胞前干细胞（前 - LSC）。