Reactivation of Gamma Globin Expression in β-Hemoglobinopathies

摘要 氧气输送到身体的组织和器官是依赖于定性和定量的 血红蛋白是由两个α和两个β样亚基组成的四聚体蛋白质。β-的 血红蛋白病是世界范围内最常见的单基因疾病， 无论患者是否存在β-地中海贫血的定量（β-地中海贫血）或定性（镰状细胞病（SCD））缺陷， 珠蛋白合成不幸的是，β-血红蛋白病的治疗选择很少。但 已经表明，另一种β-样亚基γ-珠蛋白具有补偿β-珠蛋白的能力， SCD缺陷和β-地中海贫血。迄今为止，很少有γ-珠蛋白表达的潜在激活剂被发现。 发现了此外，最近的研究表明，γ-珠蛋白启动子（~ 150 bp）处的空间位阻 转录起始位点上游）导致γ-珠蛋白表达的减少-这意味着 γ-珠蛋白激活剂结合区的一部分与这些发现一致，我做了一个混合基因组- 广泛的CRISPR激活筛选，确定了几个可能激活γ-珠蛋白的新候选基因 表情在这个提案中，我的目标是验证我在我的论文中发现的这些候选γ-珠蛋白激活剂。 筛选并进行研究，以确定这些候选基因增加γ- 珠蛋白表达此外，在我最近的工作中，我发现， 与具有正常循环的红系细胞相比，人红系细胞表达γ-珠蛋白（称为F-细胞 速度.在这个建议中，我还旨在剖析细胞周期速度调节对γ-珠蛋白的影响 表情从这项工作中，我希望发现γ-珠蛋白的关键调节因子。这些新发现的调节器 可能导致β-血红蛋白病的新疗法的发展。重要的是，这项提案 将使我能够发展技能，知识和经验，在学术事业取得成功 research.

ABSTRACT Oxygen delivery to the body’s tissues and organs is dependent on the qualitative and quantitative features of hemoglobin, a tetrameric protein made up of two α- and two β-like subunits. β- hemoglobinopathies are the most common monogenic disorders worldwide, and are defined based on whether patients have quantitative (β-thalassemia) or qualitative (Sickle Cell Disease (SCD)) defects in β- globin synthesis. Unfortunately, there are few treatment options for β-hemoglobinopathies. However, it has been shown that an alternative β-like subunit, γ-globin, has the ability to compensate for the β-globin defect in SCD and β-thalassemia. To date, few potential activators of γ-globin expression have been discovered. Additionally, a recent study showed that steric hindrance at the γ-globin promoter (~150bp upstream of the transcription start site) led to a decrease in γ-globin expression- implying the presence of a γ-globin activator-binding region. Consistent with these findings, I have performed a pooled genome- wide CRISPR activation screen that identified several novel candidate genes that may activate γ-globin expression. In this proposal, I aim to validate these candidate γ-globin activators that I uncovered in my screen and perform studies to determine the mechanism by which these candidate genes increase γ- globin expression. Additionally, in my recent work I have found that a high percentage of slow cycling human erythroid cells express γ-globin (termed F-cells) compared to erythroid cells with normal cycling speeds. In this proposal, I also aim to dissect the impact of cell cycle speed regulation on γ-globin expression. From this work I expect to uncover critical regulators of γ-globin. These newfound regulators may lead to the development of novel therapeutics for β-hemoglobinopathies. Importantly, this proposal will allow me to develop the skills, knowledge, and experience for a successful career in academic research.