Defining the Niche-dependent Role of RNA Editing in Aged and MDS Hematopoietic Stem and Progenitor Cell Dysfunction

标题定义RNA编辑在老年和MDS造血干细胞和MDS中的生态位依赖作用 祖细胞功能障碍 项目摘要 在衰老过程中，由以下因素引起的造血干/祖细胞维持受损 克隆DNA突变以及利基驱动的RNA加工放松管制可以为 骨髓增生异常综合征(MDS)启动。最近，腺苷脱氨酶升高与 我们的团队和其他研究团队展示了RNA1(ADAR1)介导的A-to-I编辑 在广泛的恶性肿瘤中导致治疗耐药性。另外，我们发现， 慢病毒增强ADAR1在HSPC中的表达促进髓系分化 上调PU1，减少休眠。全转录组RNA测序(RNA-seq) 分析表明，炎性细胞因子信号通路和RNA编辑增加 在正常老年HSPC向MDS演变过程中。因此，我们假设生态位依赖的激活 由ADAR1进行的RNA编辑为MDS提供了比正常HSPC更具竞争力的优势。大多数人 ADAR1介导的腺苷到肌苷(A-to-I)RNA编辑事件在人类中发生在双 由灵长类特定的Alu序列产生的链状RNA(DsRNA)环，包括10% 从而强调了ADAR1在功能上存在重要差异 人HSPC与小鼠HSPC的比较。然而，有限的研究努力旨在 破译ADAR1介导的RNA编辑在HSPC维护中的作用 主要是在小鼠模型中，而不是高纯度的人HSPC。因为ADAR1是由 加速衰老和MDS启动的炎性细胞因子，我们的主要目标是定义利基- RNA编辑对人HSPC细胞衰老及细胞周期调控的依赖作用 MDS启动。我们将首先确定整个转录组和单细胞的RNA编辑图谱 RNA-seq、RESSqPCR和慢病毒RNA编辑记者。ADAR1在HSPC中的功能作用 老化和MDS的启动将在基质共培养中进行检测，无论是否添加 炎性细胞因子、FUCCI2BL细胞周期报告与人源化老年HSC和MDS 免疫功能低下的小鼠模型。我们还将研究RNA编辑对APOBEC3的影响 DNA脱氨酶家族在MDS启动过程中的作用。拟议的研究是对以下问题的独特回应 PAS-13-033：刺激血液学研究：新的努力(闪耀)，因为它将 鉴定ADAR1介导的调节mRNA和miRNA编辑在HSPC髓系中的作用 炎症骨中年龄依赖性MDS启动过程中的承诺和细胞周期失控 骨髓龛。这项研究的最终目标是确定生物学、诊断和预后 ADAR1介导的RNA编辑在HSPC老化和MDS启动中的意义。

Title Defining the Niche-dependent Role of RNA Editing in Aged and MDS Hematopoietic Stem and Progenitor Cell Dysfunction Project Summary During aging, impaired hematopoietic stem and progenitor cell (HSPC) maintenance induced by clonal DNA mutations as well as niche-driven RNA processing deregulation can set the stage for myelodysplastic syndrome (MDS) initiation. Recently, increased adenosine deaminase associated with RNA1 (ADAR1)-mediated A-to-I editing was shown by our group and other research teams to contribute to therapeutic resistance in a broad array of malignancies. Also, we discovered that lentivirally enforced ADAR1 expression in HSPC enhanced myeloid differentiation commensurate with upregulation of PU.1 and reduced dormancy. Whole transcriptome RNA sequencing (RNA-seq) analysis demonstrated that inflammatory cytokine signaling pathways and RNA editing increased during normal aged HSPC evolution to MDS. Thus, we hypothesized that niche dependent activation of RNA editing by ADAR1 provides a competitive advantage for MDS over normal HSPCs. The majority of ADAR1 mediated adenosine-to-inosine (A-to-I) RNA editing events in humans occur within double- stranded RNA (dsRNA) loops created by primate-specific Alu sequences, which comprise 10 percent of the human genome, thereby underscoring that important ADAR1 functional differences exist between human HSPCs compared with their murine counterparts. However, the limited research effort aimed at deciphering the role of ADAR1-mediated RNA editing in HSPC maintenance has been performed primarily in mouse models rather than highly purified human HSPCs. Because ADAR1 is activated by inflammatory cytokines that accelerate aging and MDS initiation, our main goal is to define the niche- dependent role of RNA editing on human HSPC cell fate and cell cycle regulation during aging and MDS initiation. We will first determine the RNA editing profile by whole transcriptome and single cell RNA-seq, RESSqPCR and lentiviral RNA editing reporters. The functional role of ADAR1 in HSPC aging and MDS initiation will be examined in stromal co-cultures with or without addition of inflammatory cytokines, FUCCI2BL cell cycle reporters, and humanized aged HSC and MDS immunocompromised mouse models. We will also examine the effect of RNA editing on APOBEC3 family of DNA deaminase function during MDS initiation. The proposed study is uniquely responsive to PAS-13-033: Stimulating Hematology Investigation: New Endeavors (SHINE) because it will identify the role of ADAR1-mediated regulatory mRNA and miRNA editing in HSPC myeloid lineage commitment and cell cycle deregulation during age-dependent MDS initiation in the inflammatory bone marrow niche. The ultimate goal of this study is to determine the biological, diagnostic and prognostic significance of ADAR1-mediated RNA editing in HSPC aging compared with MDS initiation.