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Compatibility logic of human enhancer and promoter sequences

ABSTRACT: We transfected K562 cells with a STARR-seq plasmid library of paired promoter and enhancer sequences to characterize activation and compatibility between promoters and enhancers.

ORGANISM(S): Homo sapiens

PROVIDER: GSE184426 | GEO | 2021/10/25

REPOSITORIES: GEO

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Compatibility logic of human enhancer and promoter sequences

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Project description:The identification of transcriptional enhancers in the human genome is a prime goal in biology. Enhancers are typically predicted via chromatin marks, yet their function is primarily assessed with plasmid-based reporter assays. Here, we show that two previous observations relating to plasmid-transfection into human cells render such assays unreliable: (1) the function of the bacterial plasmid origin-of-replication (ORI) as a conflicting core-promoter and (2) the activation of a type I interferon (IFN-I) response. These problems cause false-positives and -negatives in luciferase assays and are prohibitive to genome-wide STARR-seq screens, which are dominated by INF-I-related false-positives, suffer from poor signal-to-noise, and miss a majority of enhancers. We overcome both problems by directly employing the ORI as a core-promoter and by inhibiting two kinases central to IFN-I induction. This corrects luciferase assays and enables genome-wide STARR-seq screens in human cells. Comprehensive enhancer activity profiles in HeLa-S3 cells uncover strong enhancers, IFN-I-induced enhancers, and enhancers endogenously silenced at the chromatin level. Our findings apply to all episomal enhancer activity assays in mammalian cells, and are key to the characterization of human enhancers.

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