Cross-species analysis of regulatory function using synthetic regulatory genomics

Understanding the evolution of regulatory DNA has been impeded both by the lack of functional genomics data available outside of a select group of organisms, as well as by the barriers to implementing genome engineering in diverse organisms for functional analysis. Given the low conservation of the genomic regulatory landscape, this presents a serious impediment to studying genome evolution and limits the value of a phylogenetic approach to understanding human genomic function. We recently developed the Big-IN genome engineering technology to rewrite large genomic loci in-place through delivery of DNA payloads upwards of 160 kb. Here we propose a new synthetic regulatory genomics approach for characterization of the regulatory function of genomic sequence across a phylogenetic tree. We will analyze the activity of a set of model loci in mouse embryonic stem cells by delivering orthologous sequences from 5 vertebrate species. We will employ two strategies to: (i) deliver larger regions (up to 160 kb) including the full gene replacing full orthologous mouse locus; and (ii) combinatorially assess pairs of shorter (~1 kb) candidate enhancer sequences. These data will open a novel approach to understanding genomic regulatory syntax through phylogenetic analysis, as well as a bridge to direct functional assessment of human disease-relevant loci. Project Number: 1R21HG014313-01A1 | Fiscal Year: 2026 | NIH Institute/Center: National Human Genome Research Institute (NHGRI) | Principal Investigator: Matthew Maurano | Institution: NEW YORK UNIVERSITY SCHOOL OF MEDICINE, NEW YORK, NY | Award Amount: $466,125 | Activity Code: R21 | Study Section: Genetic Variation and Evolution Study Section[GVE] View on NIH RePORTER: https://reporter.nih.gov/project-details/11303817