Factors modulating gamma globin gene expression

调节伽马珠蛋白基因表达的因素

• 批准号: 7122108
• 负责人: John Michael Cunningham
• 金额: $ 34.11万
• 依托单位: ST. JUDE CHILDREN'S RESEARCH HOSPITAL
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7122108
• 关键词: binding proteins; biological models; embryo /fetus; erythropoiesis; gene expression; gene induction /repression; gene targeting; genetic promoter element; genetic regulatory element; genetically modified animals; globin; hemoglobin F; hemoprotein biosynthesis; laboratory mouse; regulatory gene; sickle cell anemia; transcription factor; transport proteins

Amelioration in sickle cell disease seen with persistent fetal hemoglobin expression suggests that therapies capable of reactivating gamma-globin gone expression should be explored. Gene transfer of a transcription factor capable of reversing the gamma- to beta-globin switch in adult erythroid cells represents one approach to treatment of this devastating disorder. We have identified a protein complex, the stage selector protein (SSP) that is a strong candidate for this approach. The SSP consists of the ubiquitous transcription factor CP2/LBP-1a, p22NF-E4, an erythroid-restricted component, and ALY, a chaperone protein that stabilizes the interaction between the other two. We have demonstrated that p22 NF-E4 functions as an activator of gamma-gene expression, and in the setting of fetal/adult gene competition promotes a concomitant down-regulation of adult beta-gene expression. Enforced expression of p22NF-E4 in betaYAC transgenic mice delayed the fetal/adult switch but was unable to override the intrinsic mechanisms that govern gamma-gene silencing. We have shown that the function of the SSP is mediated by direct protein/protein interactions with co-activators and general transcription factors. Based on these findings, we postulate a role for SSP in recruiting the locus control region (LCR) to the gamma-promoter through changes in factor binding to these elements. In Specific Aim 1, we will estabh'sh the pattern of transcription factor association with the LCR and gamma-promoter in cellular and animal models of fetal erythropoiesis and examine the effects of increased expression of p22 NF-E4 in these contexts. We will determine whether the LCR and gamma-promoter are in close physical proximity in fetal erythroid cells and if this is influencedby the SSP complex. In Specific Aim 2, we will dissect the relative contribution of the individual components of the SSP to the overall function of the complex, including their role in factor binding and LCR/promoter interaction. Specific Aim 3 will focus on LCR/gamma-promoter interactions and factor occupancy in the setting of a mutation that disrupts intrinsic gamma-gene silencing, and the effects of p22NF-E4 expression in this context. In summary, these studies will define the role of the SSP complex in modifying factor association with the LCR and gamma-promoter, and determine whether the predominant function of the SSP is to directly recruit the LCR to the promoter. Ultimately they may provide new therapeutic strategies for sickle cell disease.

持续胎儿血红蛋白表达所观察到的镰状细胞病的改善表明，应该探索能够重新激活γ-珠蛋白消失表达的疗法。能够逆转成人红细胞中γ-珠蛋白向β-珠蛋白转换的转录因子的基因转移代表了治疗这种破坏性疾病的一种方法。我们 已经鉴定出一种蛋白质复合物，即阶段选择蛋白（SSP），它是这种方法的有力候选者。 SSP 由普遍存在的转录因子 CP2/LBP-1a、p22NF-E4（一种红细胞限制性成分）和 ALY（一种稳定其他两者之间相互作用的伴侣蛋白）组成。我们已经证明 p22 NF-E4 作为 γ 基因表达的激活剂起作用，并且在胎儿/成人基因竞争的情况下促进成人 β 基因表达的伴随下调。 betaYAC 转基因小鼠中 p22NF-E4 的强制表达延迟了胎儿/成人的转换，但无法超越控制 γ 基因沉默的内在机制。我们已经证明，SSP 的功能是通过与共激活因子和一般转录因子的直接蛋白质/蛋白质相互作用介导的。基于这些发现，我们假设 SSP 通过改变与这些元件结合的因子，在将基因座控制区 (LCR) 招募到 γ 启动子中发挥作用。在具体目标 1 中，我们将在胎儿红细胞生成的细胞和动物模型中建立转录因子与 LCR 和 γ 启动子的关联模式，并检查 p22 NF-E4 表达增加在这些情况下的影响。我们将确定胎儿红细胞中 LCR 和 γ 启动子是否在物理上非常接近，以及这是否受到 SSP 复合物的影响。在具体目标 2 中，我们将剖析 SSP 各个组件对复合物整体功能的相对贡献，包括它们在因子结合和 LCR/启动子相互作用中的作用。具体目标 3 将重点关注破坏内在 γ 基因沉默的突变环境中的 LCR/γ 启动子相互作用和因子占据，以及在此背景下 p22NF-E4 表达的影响。总之，这些研究将明确 SSP 复合物在修饰与 LCR 和 γ 启动子相关的因子中的作用，并确定 SSP 的主要功能是否是直接将 LCR 募集到启动子上。最终，它们可能为镰状细胞病提供新的治疗策略。