Regulation of Gene Expression in the Th2 Cytokine Locus

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DESCRIPTION (provided by applicant): During development into Th2 T cell effectors, the IL-4 locus undergoes significant changes in chromatin structure that facilitate coordinated expression of the genes within the locus. These changes include, but are not limited to, changes in DNase l sensitivity, histone acetylation, and DNA methylation. In a number of identified loci, these changes are coordinated, and possibly regulated by cis-acting elements, such as Locus Control Regions (LCR) and insulators or boundary elements. No such elements in the IL-4 locus have been identified to date. We have identified a previously unrecognized region of the RAD50 gene, which becomes DNase l hypersensitive (HS) specifically in Th2 cells and contains the properties of a Locus Control Region. We hypothesize that this region comprises elements of a Locus Control Region and possibly an insulator. The principal goal of this project is to delineate and functionally characterize elements within this region that possess LCR and/or insulator activity. The first specific aim is to characterize potential enhancer activity of HS elements within the region, as LCRs possess strong lineage-specific enhancer activity. We will use a transgenic mouse system to fulfill the goals of this aim. The second specific aim is to assess the chromatin character across the entire locus, focusing on transitions in acetylation and DNasel sensitivity patterns. These experiments are designed to identify potential boundary elements within the locus. Subsequent experiments will be performed to characterize these elements in terms of their ability to block enhancer function and/or prevent the encroachment of condensed chromatin. Inappropriate Th2 differentiation has been implicated in a number of pathological states, most notably allergic diseases. The information gleaned from these studies will contribute to our understanding of the process of Th2 differentiation, and will hopefully enable the ultimate goal, which is to intervene in this process, and enhance our ability to avoid or correct these pathological manifestations.