High-throughput interrogation of long-range enhancers

The National Human Genome Research Institute (NHGRI) awarded a $2.8 million Project Grant to The Washington University (Office of Sponsored Research Services) effective September 1, 2025, through August 31, 2029, under the Human Genome Research program (CFDA 93.172). This award supports the development and implementation of a high-throughput, genome-integrated reporter gene system designed to systematically measure enhancer activities at multiple fixed distances from target promoters. The project addresses a critical gap in understanding sequence features that enable enhancers to function across long genomic ranges, which is essential for interpreting how non-coding genetic variants contribute to heritable diseases.

The deliverables include comprehensive training datasets quantifying the effects of variables contributing to long-range enhancer function, including intrinsic enhancer strength, transcription factor binding site composition and arrangement, and chromatin environment. A unique strength of the system is its capacity to directly compare identical enhancer sequences at both long and short ranges, enabling identification of sequence features specifically required for long-distance regulatory activity. This research directly supports NHGRI's mission to accelerate scientific breakthroughs through fundamental genomics research and the development of genomic resources and methods broadly applicable to biomedical research.

Abstract

A majority of heritable disease-causing variation resides in the non-coding portions of the genome. The leading hypothesis is that these variants perturb the function of CIS-regulatory elements, such as enhancers. A remarkable property of enhancers is that they often reside at long distances from their targets. Thus, any framework for interpreting genetic variation must account for the long-range functions of enhancers. There have been many genome-wide studies of enhancers, yet we still lack an understanding of the sequence features that allow enhancers to function over long distances. This lack of understanding is due to the absence of systematic training data that measures the activities of enhancers as they are moved further and further from their target promoters. We propose to address this GAP by systematically measuring the activities of enhancers at multiple fixed distances from target promoters. To collect this data, we will introduce a high-throughput, genome-integrated reporter gene system that measures the activities of enhancers at long distances from their target promoters. We propose to quantify the effects of variables that contribute to long-range enhancer function. These variables include the intrinsic strength of an enhancer, the number and arrangement of transcription factor binding sites in an enhancer, and the chromatin environment in which an enhancer resides. A unique strength of our system is that we directly compare the same enhancer sequences at long- and short-ranges, which allows US to identify features required specifically at long distances.