: This proposal seeks to characterize the epigenetic programming that normally occurs at the IL-4/IL13 locus during the process of T helper lineage specification. This will form a basis for understanding the role environmental factors may play in perturbing normal epigenetic control of cytokine gene expression and - by extension - immune function. We have previously shown that, during Th2 lineage specification, the alleles of the IL-4 gene are subject to independent imprinting, leading to the development of epigenetically distinct Th2 sub-lineages, differing in the choice and number of IL-4 alleles they express. Similar findings have been reported for IL-13, IL-5, IL-3 and GM-CSF which- together with IL4 - are clustered within a 500kb interval on mouse chromosome 11. In order to approach the epigenetic compartmentalization established throughout this cytokine-rich multi-genic locus, we have comparatively mapped Thl, Th2 and naive CD4 T cells for the occurrence of intergenic RNA transcription at discrete sites distributed throughout a 50 kilobase (kb) interval spanning the IL-4 and IL-l3 genes. Additionally, over the same 50kb interval, we have constructed maps of distinct chromatin structural states in Thl and Th2 cells. The feature distributions on these maps reveal signatures that can distinguish among Thl, Th2 and naive CD4 T cells. Significantly, the Th2 signatures in the chromatin structure and transcriptional activity maps are almost identical, suggesting functional links between them and to the developmentally programmed specification of IL4 and IL 13 transcription inducibility. Here, we propose to extend our transcript and chromatin structure maps to the allelic level. This will allow us to determine the relationship of discrete elements in the Th2-1ineage-specific signatures comprising both transcript and chromatin structure features to the epigenetic imprints specifying active and silent IL4 and ILl3 alleles in distinct Th2 sub-lineages. Since the programming of cytokine gene expression inducibility is critical for the proper coordination of immune responses, a mechanistic understanding of the normal process that leads cells to specify particular allelic patterns of cytokine gene expression inducibility is a pre-requisite to understanding how (and whether) environmental factors can perturb the epigenetic regulation of immune responses.
