Cited by 317CRISPR and Genetic EngineeringRNA regulation and diseaseGenomics and Rare Diseases

Massively parallel assessment of human variants with base editor screens

Open access
Hanna, Ruth E., Hegde, Mudra, Fagre, Christian R., DeWeirdt, Peter C., Sangree, Annabel K., Szegletes, Zsofia, Griffith, Audrey, Feeley, Marissa N. +6 more·2021·Cell

Published version of a preprint

Abstract

Understanding the functional consequences of single-nucleotide variants is critical to uncovering the genetic underpinnings of diseases, but technologies to characterize variants are limiting. Here, we leverage CRISPR-Cas9 cytosine base editors in pooled screens to scalably assay variants at endogenous loci in mammalian cells. We benchmark the performance of base editors in positive and negative selection screens, identifying known loss-of-function mutations in BRCA1 and BRCA2 with high precision. To demonstrate the utility of base editor screens to probe small molecule-protein interactions, we screen against BH3 mimetics and PARP inhibitors, identifying point mutations that confer drug sensitivity or resistance. We also create a library of single guide RNAs (sgRNAs) predicted to generate 52,034 ClinVar variants in 3,584 genes and conduct screens in the presence of cellular stressors, identifying loss-of-function variants in numerous DNA damage repair genes. We anticipate that this screening approach will be broadly useful to readily and scalably functionalize genetic variants.

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Crossref · 10.1016/j.cell.2021.01.012

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  • Base editing has been utilized for single point mutations and targeted mutation induction for therapeutic corrections of diseases, mutational screening, and gene knockout.

    “It has been utilized for several applications, including single point mutations and targeted mutation induction for therapeutic corrections of diseases (Musunuru et al., 2021), mutational screening, and gene knockout among the genomes (Hanna et al., 2021; Que et al., 2010).”

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Authors
Hanna, R. E., Hegde, M., Fagre, C. R., DeWeirdt, P. C., Sangree, A. K., Szegletes, Z., Griffith, A., Feeley, M. N., Sanson, K. R., Baidi, Y., Koblan, L. W., Liu, D. R., Neal, J. T., & Doench, J. G.
Affiliations
Broad Institute (US)
Corresponding author
Doench, John G.
Venue
Cell
Publisher
Elsevier BV
Published
2021-02
Volume
184
Issue
4
Pages
1064-1080.e20
ISSN
0092-8674 (ISSN-L)
OpenAlex
W3132864675
Cited by
316
References
81
Citations by year
2026: 41 · 2025: 65 · 2024: 57 · 2023: 62 · 2022: 57 · 2021: 29 · 2020: 5
FWCI
16.24
Type
journal article
Language
en
Open access
Locations
  • Cell (publishedVersion)
  • bioRxiv (Cold Spring Harbor Laboratory) (acceptedVersion, cc-by-nc-nd)
  • PubMed (publishedVersion)
Funders
  • National Science Foundation
  • National Institutes of Health
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › CRISPR and Genetic Engineering
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA regulation and disease
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Genomics and Rare Diseases
Keywords
single nucleotide variants, loss-of-function mutations, BRCA1, BRCA2, BH3 mimetics, PARP inhibitors, drug resistance, drug sensitivity, DNA repair genes, cellular stress, single guide RNA library, ClinVar variants
MeSH terms
Gene Editing, Loss of Function Mutation, Alleles, Base Sequence, Humans, Poly(ADP-ribose) Polymerases, Selection, Genetic, Genetic Variation, Reproducibility of Results, Mutagenesis, Point Mutation, BRCA1 Protein, Catalytic Domain, BRCA2 Protein, Cell Line, Tumor, bcl-X Protein, High-Throughput Nucleotide Sequencing, Myeloid Cell Leukemia Sequence 1 Protein

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