The long term goal of the research is to understand the tissue and developmental-specific regulation of genes expressed in erythroid cells. The Investigators have concentrated on the regulation of genes in the b globin locus as a model of erythroid-specific gene expression. The regulation of these genes depends on transcription factors which are limited to erythroid cells or cells of closely related lineages, and on an organized chromatin structure dictated by the formation of hypersensitive sites within a region known as the LCR or locus control region. The investigations have led to the identification of a new cis element involved in the expression of the human b globin gene. This element confers a strong enhancer activity and appears to function in conjunction with hypersensitive site 2. Thus, the factor binding to this site is a candidate to mediate this critical interaction between the LCR and the globin promoter. In addition, it is known that transcription factors regulate the expression of other transcription factors and these interrelationships suggest a complex hierarchy of interaction that not only regulates specific gene products in a particular tissue, but also regulate each other to function in early development. It is thus likely that the study of the interrelationship among transcription factors will be important for understanding both the expression of these genes in adult tissue, and the differentiation of specific cells during early development. They have therefore also focused on the regulation of the EKLF erythroid-specific transcription factor gene.
The specific aims of our studies are as follows: 1) Identification and characterization of a new cis element in the distal 5' flanking sequence of the human b globin gene. We will characterize this site through footprinting and oligonucleotide competion studies and the factor that binds the element will be isolated and cloned with the use of the yeast one-hybrid system or conventional procedures. 2) Regulation of the EKLF gene. The expression of transcription factor genes is highly regulated and previous studies involving both erythroid and non-erythroid transcription factors are revealing interesting interrelationships. To understand this hierarchy, we plan to define the cis elements and transacting factors responsible for the tissue-specific expression of the EKLF gene. Together, these studies will provide new insights into mechanisms of gene regulation as well as the biology of hematopoietic systems.
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