Direct characterization of cis-regulatory elements and functional dissection of complex genetic associations using HCR-FlowFISH

Reilly SK, Gosai SJ, Gutierrez A, Mackay-Smith A, Ulirsch JC, Kanai M, Mouri K, Berenzy D, Kales S, Butler GM, Gladden-Young A, Bhuiyan RM, Stitzel ML, Finucane HK, Sabeti PC, Tewhey R (2021) Direct characterization of cis-regulatory elements and functional dissection of complex genetic associations using HCR-FlowFISH. Nature Genetics 53, 1166-1176 https://doi.org/10.1038/s41588-021-00900-4.

Effective interpretation of genome function and genetic variation requires a shift from epigenetic mapping of cis-regulatory elements (CREs) to characterization of endogenous function. We developed hybridization chain reaction fluorescence in situ hybridization coupled with flow cytometry (HCR–FlowFISH), a broadly applicable approach to characterize CRISPR-perturbed CREs via accurate quantification of native transcripts, alongside CRISPR activity screen analysis (CASA), a hierarchical Bayesian model to quantify CRE activity. Across >325,000 perturbations, we provide evidence that CREs can regulate multiple genes, skip over the nearest gene and display activating and/or silencing effects. At the cholesterol-level-associated FADS locus, we combine endogenous screens with reporter assays to exhaustively characterize multiple genome-wide association signals, functionally nominate causal variants and, importantly, identify their target genes.

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