Systematic mapping of enhancers, silencers, and topological elements with prime editing deletion screens

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Abstract
Noncoding regulatory elements in the human genome can control gene expression through distinct mechanisms, including acting as enhancers, silencers, or topological elements. However, we lack tools to identify all such classes systematically and quantify their effects on gene expression in an endogenous genomic context. Here we develop Swap-seq, a method that combines pooled twin prime editing with flow-sorting to delete hundreds of endogenous regulatory sequences and measure their quantitative effects on the expression of a nearby gene. We apply Swap-seq to dissect the regulatory elements that control PPIF expression in THP-1 monocytes. We quantify the effects of distal and intronic enhancers, characterize CTCF insulators, and discover a silencer that regulates distal genes through histone deacetylase (HDAC)-mediated repression of neighboring enhancers. Hundreds of other genomic elements share its chromatin signatures and transcription factor binding sites, suggesting that this silencer represents a broader class of repressive elements. We leverage Swap-seq data to benchmark sequence-to-function models and show that they often capture the local activity of regulatory elements but fail to link them to distal target genes. Swap-seq thus provides a generalizable tool to discover regulatory elements across mechanistic classes, characterize their effects on gene expression, and generate large-scale datasets for evaluating sequence-to-function models.
Funding Information
  • NIH-NHLBI (R01 HL171609)
  • NIH-NHLBI (R01 HL184893)
  • the Applebaum Foundation
  • The Novo Nordisk Foundation Center for Genomic Mechanisms of Disease (Novo Nordisk Fonden NNF21SA0072102)
  • BASE Research Initiative at Lucile Packard Children’s Hospital Stanford
  • NIH-NIGMS (R35 GM128947)
  • NIH-NIGMS (5T32GM007790)
  • NIH-NHGRI (R01 HG011866)
  • NIH-NHGRI (R01HG014500)
  • NSF Graduate Research Fellowship (DGE-1656518)
  • Knight-Hennessy Scholars
  • Gabilan Stanford Graduate Fellowship
  • American Heart Association (26PRE1565731)
  • National Institute of Biomedical Imaging and Bioengineering (R01EB035127)