Identifying chromosomal regions associated with glucocorticoid-regulated gene transcription

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PROJECT SUMMARY The way the genome is arranged in the nucleus plays an important role in gene regulation but several aspects remain unknown. Genomes are spatially organized into compartments further made up of megabase structures called topologically associating domains (TADs) that have similar chromatin modifications and histone marks. TAD boundaries are demarcated by the Cohesin complex and the CCCTC binding factor (CTCF). It is presumed that response elements, segments of DNA where transcription factors typically bind to increase or decrease gene transcription, interact with promoters of genes within the scope of a TAD. DNA response elements can be far apart on a linear genome but chromatin loops bring these elements in proximity of gene promoters for gene transcription in the nucleus. I aim to investigate the relationship between gene transcription and two higher-order genome structures, chromatin loops and TADs. It has proven difficult to identify response elements and their cognate promoters thus the relationship between metazoan transcriptional regulation and genomic arrangements has not been defined. The first objective of this project is to identify functional response elements and their cognate regulated promoters (Aim 1). I will exploit the properties of the glucocorticoid receptor (GR), a transcriptional regulatory factor whose genomic binding and regulatory activities require prior binding by glucocorticoid ligands. Glucocorticoid receptor response elements (GREs) are segments of DNA that confer a regulatory action in vivo but it is not known which GR binding site is a GRE and which specific gene it targets. I will then assess the formation and positions of chromatin loops relative to validated glucocorticoid response element (GRE) – promoter pairs, and test for loop degeneracy (Aim 2). Finally, I shall examine whether TAD boundaries limit looping and GRE action (Aim 3). This proposal will be the first to properly identify GREs and their target genes. This work will provide us with insight regarding gene regulation and TAD regulatory structure.