A Multiplexed Single-Cell CRISPR Screening Platform Enables Systematic Dissection of the Unfolded Protein Response

Britt Adamson, Thomas M. Norman, Marco Jost, Min Y. Cho, James K. Nuñez, Yuwen Chen, Jacqueline E. Villalta, Luke A. Gilbert, Max A. Horlbeck, Marco Y. Hein, Ryan A. Pak, Andrew N. Gray, Carol A. Gross, Atray Dixit, Oren Parnas, Aviv Regev, Jonathan S. Weissman

Research output: Contribution to journalArticle

267 Scopus citations

Abstract

Functional genomics efforts face tradeoffs between number of perturbations examined and complexity of phenotypes measured. We bridge this gap with Perturb-seq, which combines droplet-based single-cell RNA-seq with a strategy for barcoding CRISPR-mediated perturbations, allowing many perturbations to be profiled in pooled format. We applied Perturb-seq to dissect the mammalian unfolded protein response (UPR) using single and combinatorial CRISPR perturbations. Two genome-scale CRISPR interference (CRISPRi) screens identified genes whose repression perturbs ER homeostasis. Subjecting ∼100 hits to Perturb-seq enabled high-precision functional clustering of genes. Single-cell analyses decoupled the three UPR branches, revealed bifurcated UPR branch activation among cells subject to the same perturbation, and uncovered differential activation of the branches across hits, including an isolated feedback loop between the translocon and IRE1α. These studies provide insight into how the three sensors of ER homeostasis monitor distinct types of stress and highlight the ability of Perturb-seq to dissect complex cellular responses.

Original languageEnglish (US)
Pages (from-to)1867-1882.e21
JournalCell
Volume167
Issue number7
DOIs
StatePublished - Dec 15 2016

All Science Journal Classification (ASJC) codes

  • Biochemistry, Genetics and Molecular Biology(all)

Keywords

  • CRIPSRi
  • CRISPR
  • Single-cell RNA-seq
  • cell-to-cell heterogeneity
  • genome-scale screening
  • single-cell genomics
  • unfolded protein response

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    Adamson, B., Norman, T. M., Jost, M., Cho, M. Y., Nuñez, J. K., Chen, Y., Villalta, J. E., Gilbert, L. A., Horlbeck, M. A., Hein, M. Y., Pak, R. A., Gray, A. N., Gross, C. A., Dixit, A., Parnas, O., Regev, A., & Weissman, J. S. (2016). A Multiplexed Single-Cell CRISPR Screening Platform Enables Systematic Dissection of the Unfolded Protein Response. Cell, 167(7), 1867-1882.e21. https://doi.org/10.1016/j.cell.2016.11.048