MECHANISMS OF ERYTHROID GENE EXPRESSION

这项建议的广泛的长期目标是阐明 珠蛋白和其他红细胞的转录调节机制 细胞特异性基因 更好地理解这些过程是可能的 提出操纵珠蛋白调节的方法， 在血红蛋白病如镰状细胞病的治疗中 β地中海贫血 具体目标是分析三个关键的DNA 负责珠蛋白的元件（此处称为加塔、CACC和AP-1/NFE-2） 基因调控 就加塔元素而言，这将涉及研究 其在启动子功能中的作用; AP-1/NFE-2元件将在 为了更好地理解多种转录因子， 在他们身上。 我们将构建携带最小启动子的转基因小鼠， 加塔、CACC和AP-1/NFE-2的组合驱动β- 半乳糖苷酶，以测试这些元素对组织和发育的影响 阶段特异性 这些研究可能对设计 基因治疗载体。 此外，我们会研究规管加塔 瞬时和稳定转染的红系细胞系中的顺式元件 为了发现加塔-1是否增加了 启动子或者更确切地说是随机作用以增加 保持活跃而不影响其活动水平。 最近的工作从 这个实验室和其他实验室表明，加塔元素（和推测的 加塔-1）使红细胞启动子和增强子更易接近 而不是直接激活它们。 最后 将探索AP-1/NFE-2元件的复杂调节。 虽然NFE-2 已被证明可以结合这些元素，并且对于它们的 激活，该因子（与AP-1不同）不结合所有AP-1/NFE-2 元素 将研究AP-1在AP-1/NFE-2调节中的作用 通过评估红系细胞中AP-1的组成， 这个组成。 此外，由于AP-1/NFE-2元件类似于核 激素反应元件，这些元件将被研究的调节， 类固醇激素受体家族的成员。

The broad long-term objective of this proposal is to elucidate the mechanisms of transcriptional regulation of globins and other red blood cell-specific genes. A better understanding of these processes is likely to suggest means of manipulating globin regulation, which would be of use in the treatment of hemoglobinopathies such as sickle cell disease and beta-thalassemia. The specific aims are to analyze the three key DNA elements (here termed GATA, CACC and AP-1/NFE-2) responsible for globin gene regulation. In the case of the GATA element, this will involve study of its role in promoter function; AP-1/NFE-2 elements will be studied in order to better comprehend the multiple transcription factors that may act on them. We will construct transgenic mice carrying minimal promoters consisting of combinations of GATA, CACC, and AP-1/NFE-2 driving expression of beta- galactosidase to test effects of these elements on tissue and developmental stage specificity. These studies may have implications for the design of gene therapy vectors. In addition, we will study the regulation of GATA cis-elements in transiently and stably transfected erythroid cell lines in order to discover whether GATA-1 increases the transcriptional activity of the promoter or rather acts stochastically to increase the likelihood of being active without affecting its level of activity. Recent work from this and other laboratories suggests that GATA elements (and presumable GATA-1) function to make erythroid promoters and enhancers more accessible to other factors rather than activating them directly. Finally, the complex regulation of AP-1/NFE-2 elements will be explored. While NFE-2 has been shown to bind these elements and be necessary for their activation, this factor (unlike AP-1) does not bind all AP-1/NFE-2 elements. The role of AP-1 in the regulation of AP-1/NFE-2 will be studied by assessing the composition of AP-1 in erythroid cells and manipulating that composition. In addition, since AP-1/NFE-2 elements resemble nuclear hormone response elements, these elements will be studied for regulation by members of the steroid hormone receptor family.