3D Chromatin Studies in Pediatric B Cells To Study the Genetics of Autoimmunity

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Abstract – The purpose of this small project is to gather preliminary data we need to support a range of projects relating to autoimmunity in children, and particularly to pediatric systemic lupus erythematosus (pSLE). Specifically, we need to generate 3D chromatin maps on B cells from children in order to carry our work forward. The project also addresses the serious need for reference genomic data sets that can be used in a broad spectrum of research aimed at understanding childhood-onset, immune-driven diseases. The problem we face is a common one in pediatrics and limits our ability to rigorously pursue genetic and genomic translational studies: the uncertainty as to whether publicly available genomic data sets (e.g., from Roadmap Epigenomics) can be used as comparison data to interpret similar data generated in children. The limitations in the use public data sets therefore represent a “tax” on the research of pediatric investigators that is not always required of investigators studying adult or adult-onset diseases, where a broad variety of genomic data sets are already available, including those from diseased tissues. In this project, we will produce an reference data set that will be useful to a range of investigations into immune-based diseases that occur in children. Our work in pediatric autoimmune diseases is aimed at identifying causal variants on genetic risk haplotypes as well as the genes impacted by those variants. These target genes may not be those nearest the causal variant, or even genes on the risk haplotype. They will almost invariably be genes within the same chromatin loop or topologically associated domains, chromatin loop structures that are anchored by the CCCT binding factor (CTCF) and cohesin. Identifying these loop structures is therefore a key step in identifying target genes. This project is aimed to identify these structures, and the genes expressed within them, in pediatric samples. Our approach is straightforward. We will use CTCF Cut-and-Run and HiChIP to identify CTCF-anchored loop structures in the B cells of healthy children. We will also perform RNA sequencing on these same samples in order to identify the genes expressed within loops. At the completion of this 2-year project, we will have obtained the essential data that we need to rigorously interpret the studies we already have under way in pSLE and will have provided the field with an essential tool for future investigations into autoimmunity in children.