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Muerdter, Felix; Boryń, Łukasz M; Woodfin, Ashley R; Neumayr, Christoph; Rath, Martina; Zabidi, Muhammad A; Pagani, Michaela; Haberle, Vanja; Kazmar, Tomáš; Catarino, Rui R; Schernhuber, Katharina; Arnold, Cosmas D; Stark, Alexander
Nature methods, 02/2018, Letnik: 15, Številka: 2Journal Article
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 such assays are rendered unreliable by two previously reported phenomena relating to plasmid transfection into human cells: (i) the bacterial plasmid origin of replication (ORI) functions as a conflicting core promoter and (ii) a type I interferon (IFN-I) response is activated. These cause confounding false positives and negatives in luciferase assays and STARR-seq screens. We overcome both problems by employing the ORI as core promoter and by inhibiting two IFN-I-inducing kinases, enabling genome-wide STARR-seq screens in human cells. In HeLa-S3 cells, we 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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in: SICRIS
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