Regulation of gene expression underlies the enormous diversity of cell types, and the responses of cells to myriad signals. Correct regulation of gene expression is thus critical, and defects in this process underlie many human disorders.
To a large extent, gene expression patterns are driven by regulatory DNA sequences that surround the genes and control their transcriptional activity. These regulatory elements include promoters (positioned at the start of each gene) and enhancers, which are often located tens to hundreds of kb away from the gene that they regulate. How such apparently haphazard linear arrangements can result in specific gene regulation is a major puzzle. To unravel this logic, it is necessary to systematically alter the positions of enhancers, promoters and other regulatory elements and study the effect on gene activity. So far, no efficient method has been available for this purpose. Our aim is to develop and apply RElocate, a scalable, broadly applicable technology to transplant selected DNA elements to hundreds of alternative positions within a ~2 Mb region, and track the functional consequences.
We will employ RElocate in combination with a high-throughput combinatorial reporter assay to systematically study how enhancers "choose" and activate the correct target promoter(s). Furthermore, we will adapt RElocate to precisely map how enhancers and promoters contact and communicate with each other. Finally, we will use the method to fine-map the repressive/activating chromatin landscape of selected regions at high resolution, and elucidate how enhancers and promoters may respond differently when inserted throughout this landscape.
This work will reveal how the ordering and spacing of regulatory elements along the genome contributes to optimal gene regulation, and will yield a powerful perturbation tool with many applications in genome biology and human genetics.
