FACTORS MODULATING GAMMA GLOBIN GENE EXPRESSION

胎儿血红蛋白持续表达的镰状细胞性疾病的临床改善应该为这种疾病探索。基因转移一种能够逆转成年红系细胞中伽马到β珠蛋白转换的转录因子是治疗这种毁灭性疾病的一种方法。与转移编码珠蛋白基因的顺式序列相比，这种策略的优势是可以预期的伴随的β/S表达的减少。该项目的长期目标是确定一个因素(S)，该因素可用于这种基因治疗方法。在目前的筹资期间，我们确定了三个可能符合这一标准的因素。首先，我们鉴定了阶段选择器蛋白复合体的胎儿和红系成分。在胎儿红细胞生成过程中，这个复合体参与了基因位点控制区对γ启动子的募集。在我们的第一个特定目标中提出了这个复合体的研究，试图确定这个因子C106在SSP复合体的形成中的作用，并确定它在胎儿红系细胞中的作用。我们将在一种新的原始和确定的红细胞生成模型、转人β珠蛋白基因的小鼠和人类红系祖细胞中评估该因子激活胎儿血红蛋白表达的能力。最终，如果这些研究表明C106能够重新激活胎儿血红蛋白的表达，它在镰刀状红细胞生成和非人类灵长类动物的NOD/SCID模型中调节伽马珠蛋白基因表达的能力将与项目4和项目5合作确定。此外，我们还开发了一种新的筛选策略，利用新的化学探针来寻找更多的胎儿珠蛋白调控基因。通过这一策略，正如在特定目标2中提出的工作所概述的那样，我们已经确定了两个因子，HLH蛋白Id2和一个新的CCAAT盒结合蛋白同源物Hap5h。有趣的是，Id2在胎儿红系细胞中的强制表达导致伽玛珠蛋白基因表达的9倍诱导。我们建议进一步确定这些因素的特征，以确定它们在伽马到β转换中的作用，以及它们潜在的治疗作用。最后，在具体目标3中，我们描述了更多具有不同结构的新型化学探针的使用，这些探针使我们能够识别更多的胎儿调节因子。综上所述，这些研究将加深我们对伽马珠蛋白基因表达的分子调控的理解，并可能为镰状细胞疾病的治疗提供新的治疗策略。

The clinical amelioration in sickle cell disease seen with persistent fetal hemoglobin expression should be explored for this disease. Gene transfer of a transcription factor capable of reversing the gamma to beta globin switch in adult erythroid cells represents one approach to the treatment of this devastating disorder. n advantage of this strategy over the transfer of cis-sequences encoding the globin genes is the concomitant reduction of beta/S expression that would be anticipated. The long-term goal of this project is to identify a factor(s) which may be utilized in this approach to gene therapy. Over the current funding period we have identified three factors which may fulfill this criteria. Firstly, we have identified a fetal and erythroid component of the stage selector protein complex. This complex has been implicated in the recruitment of the locus control region to the gamma promoter during fetal erythropoiesis. This complex has studies proposed in our first specific aim seek to define the role of this factor, c106, in the formation of the SSP complex and determine its role in fetal erythroid cells. We will evaluate the ability of the factor to activate fetal hemoglobin expression in a novel model of primitive and definitive erythropoiesis, mice transgenic for the human beta globin locus and human erythroid progenitors. Ultimately, if these studies demonstrate that c106 is capable of reactivating fetal hemoglobin expression, its ability to modulate gamma-globin gene expression in a NOD/SCID model of sickle erythropoiesis and non-human primates will be determined in collaboration with Projects 4 and 5. In addition, we have also developed a novel screening strategy to identify further fetal globin regulatory genes utilizing novel chemical probes. With this strategy, as outlined in the work proposed in specific aim 2, we have identified two factors, the HLH protein Id2 and a novel CCAAT box binding protein homologue Hap5h. Interestingly, enforced expression of Id2 in fetal erythroid cells results in a 9 fold induction of gamma globin gene expression. We propose to characterize these factors further, to determine their role in the gamma to beta switch, as well as their potential therapeutic role. Finally, in specific aim 3 we describe the use of further novel chemical probes with diverse structures that allow us to identify further fetal regulatory factors. In summary, these studies will enhance our understanding of the molecular regulation of gamma globin gene expression and may provide new therapeutic strategies for the treatment of sickle cell disease.