Reconstituting gene regulatory landscapes to identify mechanisms of transcriptional control and dysregulation

The National Institute of General Medical Sciences awarded the University of Southern California $426,145 on August 1, 2026, under the Biomedical Research and Research Training program (CFDA 93.859) to investigate molecular mechanisms controlling enhancer-promoter compatibility in mammalian gene expression.

The research develops scalable experimental tools to identify how transcription factors cooperate to modulate enhancer activity and establish enhancer-promoter and enhancer-enhancer communication. The work includes creating a reduced-complexity, genome-integrated reporter assay to quantify how distinct transcription factor binding site combinations, their affinities, and copy number affect enhancer activity across different cellular contexts, and establishing how a transcription factor combinatorial code governs regulatory element communication. This insight is intended to aid interpretation and identification of disease-linked non-coding mutations and inform therapies targeting transcriptional dysregulation in cancer and rare genetic disorders.

The University of Southern California's Department of Contracts and Grants serves as the recipient. Performance occurs in Los Angeles, California. The award period runs from August 1, 2026, through April 30, 2031. The assistance type is a Project Grant, which under NIGMS typically carries a review-to-award timeline of approximately nine months.

Abstract

Mammalian gene expression is governed by a 'separation of function' principle, by which gene regulatory activity is split between promoters (p) and enhancers (e). Conceptually, enhancers work by recruiting a combination of transcription factors (TFS), which bind DNA in a sequence-specific manner. Once bound, TFS interact with coregulators to induce epigenetic changes. While many examples indicate a sequence-dependent e-p compatibility code, little is known about the exact molecular interactions that mediate communication. Shedding light on this process is critical to advance our understanding of how enhancer-centric mutations fuel transcriptional dysregulation and disease - from cancer to rare genetic disorders. The goal of this research program is to investigate the molecular mechanisms that control mammalian e-p compatibility. Our focus is on the development of scalable experimental tools that can break down regulatory complexity to a human-interpretable level. Such insight is needed to design therapies that directly target 'transcriptional dysregulation'. We and others have shown that TFS cooperate to boost enhancer activity and communication. However, all these observations are based on average effects across unique genomic contexts, precluding insight into the contributions of individual TFS and TF pairs. Our first goal is therefore to develop a reduced-complexity, genome-integrated reporter assay that can quantify how distinct TF binding site combinations, their affinities and copy number, modulate enhancer activity in different cellular contexts. This work will provide a catalogue of TF combinatorial function, which will aid the interpretation and identification of disease- linked non-coding mutations. Our second goal is to identify how a TF combinatorial code establishes e-p and e- e communication. Specifically, we will probe how enhancer sequence alters epigenetic landscapes and how IT is interpreted by different promoter architectures. We will optimize the extra-SEQ tool i previously developed to reconstitute thousands of kilobase-scale e-p and e-e-p landscapes in a single experiment. We will track the transcriptional and epigenetic output for each DE novo created genotype in different cellular contexts to identify the minimally required set of TF and promoter motifs that mediate e-p and e-e communication. While perturbing DNA is an important first step to decode regulatory grammar, IT only provides correlative insight into the protein interactions that mediate e-p communication. Our third goal is therefore to directly map the protein domains involved in this process. This research will shed light on the molecular layers involved in translating enhancer sequence to transcriptional output, insight we lack, and which is necessary to advance personalized therapies whose aim IT is to disrupt transcriptional dysregulation at the source - perturbing regulatory interactions that drive disease states without affecting 'normal' gene regulation.