Locus-linked Regulatory Motifs of Globin Gene Switching

[脊椎动物基因调控的分析长期以来一直受益于从β-珠蛋白开始的研究， 基因座发育阶段和组织特异性转录控制的基本原则起源于 研究这个复杂的基因座。我研究的总体目标是 控制人类P样珠蛋白基因转换的基本调节机制，在这种情况下，自主Y珠蛋白 基因沉默在成熟阶段的决定性红细胞生成。我们最近发现了一种新的艾珠蛋白 与第一个新的HPFH点连接的基因沉默子基序和相关的阻遏物复合物 10年内描述的突变。该沉默子位于相对于mRNA CAP位点的-570处， 一个加塔结合基序和阻遏是由在该位点的加塔-1结合介导的。在我们的这个网站的突变 （3-YAC转基因小鼠模型导致HPFH表型。这种表型得到了 在一个家族中发现了HPFH，该家族在戈伊-9 ′ c-bin基因中具有类似突变（Luo等，2004年）。的 一项拟议中的研究将检验Y-珠蛋白基因沉默部分通过加塔-1介导的假设 辅阻遏物的募集。我们将评估-570沉默器在体内的功能，表征沉默器的特性， 加塔-1-Mi2阻遏物复合物，结合在该位点并鉴定其组分蛋白。几个HPFH 突变与加塔-1结合的丧失有关。我们将测试这种机制的普遍性， vivo.最后，我们将产生和确定新的阻遏物结合位点内的Ay-globin基因启动子 使用基于细胞的报告基因测定来选择HPFH突变。这些目标将扩展我们的研究 从上一个资助期到功能研究中确定各种c/s调节基序的作用 其将辨别由反式作用因子引起的γ-珠蛋白基因沉默的分子机制，所述反式作用因子包括 阻遏复合物。这些蛋白质可能被开发成增加HbF的潜在治疗靶点。 镰状细胞病（SCO），β-地中海贫血和库利贫血是常见的遗传性疾病， 影响着全世界数百万人仅上合组织就影响了每年出生的500名非洲裔美国人中的一人。 了解控制珠蛋白基因转换的分子机制可能有助于发展 治疗这些疾病的靶向疗法或疗法，特别是旨在启动胎儿的研究， γ-珠蛋白基因，这已被证明是有效的治疗SCO。

[Analysis of gene regulation in vertebrates has long benefited from studies initiated with the p-globin locus. Fundamental principles of developmental stage- and tissue-specific control of transcription originated with the study of this complexly regulated locus. The overall goal of my research has been to understand basic regulatory mechanisms governing human p-like globin gene switching, in this instance, autonomous yglobin gene silencing during the adult stage of definitive erythropoiesis. We recently identified a novel Ayglobin gene silencer motif and an associated represser complex that are linked to the first new HPFH point mutation to be described in over 10 years. This silencer is located at -570 relative to the mRNA CAP site in a GATA binding motif and repression is mediated by GATA-1 binding at this site. Mutation of this site in our (3-YAC transgenic mouse model resulted in a HPFH phenotype. This phenotype is corroborated by the discovery of HPFH in a family with the analogous mutation in the Gy-9'c-bin gene (Luo et al., 2004). The proposed study will test the hypothesis that y-globin gene silencing is mediated, in part, through GATA-1 recruitment of co-repressors. We will assess the function of the -570 silencer in vivo, characterize the GATA-1-Mi2 represser complex that binds at this site and identify its component proteins. Several HPFH mutations are associated with a loss of GATA-1 binding. We will test the universality of this mechanism in vivo. Finally, we will generate and identify novel represser binding sites within the Ay-globin gene promoter using a cell-based reporter assay to select for HPFH mutations. These aims will extend our studies identifying the role of various c/s-regulatory motifs from the previous funding period into functional studies that will discern molecular mechanisms of y-globin gene silencing by trans-acting factors comprising represser complexes. These proteins may be developed into potential therapeutic targets to increase HbF.] Sickle cell disease (SCO), p-thalassemias and Cooley's anemia are common genetic diseases that affect millions of people worldwide. SCO alone impacts one of 500 African Americans born each year. Understanding the molecular mechanisms controlling globin gene switching may aid in the development of targeted therapies or therapeutics to treat these diseases, particularly research aimed at turning on the fetal y-globin genes, which has been shown to be effective for the treatment of SCO.