Iron-sensitive RNA regulation during erythropoiesis

项目摘要 适当的红细胞生成需要干细胞和干细胞的基因表达和蛋白质合成的协调变化。 祖细胞分化并积累维持生命所需的大量含铁血红蛋白 在人类身上。贫血--循环红细胞不足--最常见的原因是缺铁， 仍然是世界上最常见的营养不足之一。除了血红蛋白的产生，铁在 红细胞生成过程中的其他细胞过程，如基因表达和RNA加工， 明白RNA加工事件是由RNA结合蛋白介导的，例如聚C结合蛋白 （多氯联苯）。PCBP 2通过驱动红细胞的基因转录物的选择性剪接调节红细胞生成 差异化方案。令人惊讶的是，PCBPs也被发现作为细胞内的铁伴侣， 这就提出了一个问题，即铁结合是独立于RNA结合还是与RNA结合协同发生。鉴于 铁水平密切调节红细胞分化，我们试图研究PCBP活性是否 在红细胞发育的背景下由铁调节。具体来说，我们假设铁调节了 PCBPs对靶RNA的亲和力或特异性，导致RNA剪接的变化，这对红色 血细胞分化我们将通过i）确定铁对PCBP-RNA的影响来解决这一假设 使用无偏的高通量体外测定，ii）评估铁对PCBP调节的 细胞中的RNA剪接事件，以及iii）确定原发性肿瘤中的总体基因表达和RNA剪接变化。 缺铁和不缺铁的人类和小鼠样本。从这些实验中我们将发现铁- 决定红细胞生成的敏感RNA剪接程序。

PROJECT SUMMARY Proper erythropoiesis requires coordinated changes in gene expression and protein synthesis for stem and progenitor cells to differentiate and amass the striking amount of iron-laden hemoglobin required to sustain life in humans. Anemia—insufficient circulating erythrocytes—is most frequently caused by iron deficiency which remains one the most common nutritional deficit in the world. Beyond hemoglobin production, the role of iron in other cellular processes during erythropoiesis, such as gene expression and RNA processing is poorly understood. RNA processing events are mediated by RNA binding proteins, such as the poly C binding proteins (PCBPs). PCBP2 regulates erythropoiesis via alternative splicing of gene transcripts that drive erythroid differentiation programs. Surprisingly, PCBPs were also discovered to function as intracellular iron chaperones, raising the question of whether iron-binding occurs independently of or in concert with RNA-binding. Given that iron levels intimately modulate red blood cell differentiation, we sought to investigate whether PCBP activity is modulated by iron in the context of red cell development. Specifically, we hypothesize that iron modulates the affinity or specificity of PCBPs towards target RNAs causing changes in RNA splicing that are important for red blood cell differentiation. We will address this hypothesis by i) determining the effect of iron on PCBP-RNA interactions using unbiased high throughput in vitro assays, ii) evaluating the effect of iron on PCBP-regulated RNA splicing events in cells, and iii) determine global gene expression and RNA splicing changes in primary human and mouse samples with and without iron deficiency. From these experiments we will uncover iron- sensitive RNA splicing programs that dictate erythropoiesis.