Journal article
A Single-Cell Transcriptomics CRISPR-Activation Screen Identifies Epigenetic Regulators of the Zygotic Genome Activation Program
C Alda-Catalinas, D Bredikhin, I Hernando-Herraez, F Santos, O Kubinyecz, MA Eckersley-Maslin, O Stegle, W Reik
Cell Systems | CELL PRESS | Published : 2020
Open access
Abstract
Zygotic genome activation (ZGA) is an essential transcriptional event in embryonic development that coincides with extensive epigenetic reprogramming. Complex manipulation techniques and maternal stores of proteins preclude large-scale functional screens for ZGA regulators within early embryos. Here, we combined pooled CRISPR activation (CRISPRa) with single-cell transcriptomics to identify regulators of ZGA-like transcription in mouse embryonic stem cells, which serve as a tractable, in vitro proxy of early mouse embryos. Using multi-omics factor analysis (MOFA+) applied to ∼200,000 single-cell transcriptomes comprising 230 CRISPRa perturbations, we characterized molecular signatures of ZGA..
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Awarded by Bundesministerium für Bildung und Forschung
Funding Acknowledgements
The authors thank all members of the Reik and Stegle laboratories for helpful discussions. We also thank Laura Benson for technical assistance; Mario Iurlaro for helpful discussions on Smarca5; Paul Datlinger for advice on CROPseq lentiviral preparation; Felix Krueger and the bioinformatics facility from the Babraham Institute for processing sequencing data and assistance with repeat mapping; Lia Chappell for training in 10x Genomics library preparation; Anne Segonds-Pinchon for statistical advice; the sequencing facilities at Babraham Institute, Sanger Institute, and CRUK in Cambridge for high-throughput library preparation and sequencing; and the flow cytometry facility at the Babraham Institute for cell sorting. Smarca5 (Snf2h) KO ESCs were a kind gift from Dirk Sch_ubeler. Lenti sgRNA(MS2)_puro backbone (Addgene 73795), lenti dCAS-VP64_Blast (Addgene 61425), and lenti MS2-P65-HSF1_Hygro (Addgene 61426) were a gift from Feng Zhang. pMD2.G (Addgene 12259) and psPAX2 (Addgene 12260) were a gift from Didier Trono. C.A.-C. was supported by a postgraduate award by UK Research and Innovation (UKRI, 1645504). D.B is supported by a Darwin Trust fellowship. I.H.-H. is supported by a Marie Sklodowska-Curie Individual Fellowship (751439). M.A.E.-M. is supported by a BBSRC Discovery fellowship (BB/T009713/1). Research in the Reik lab is supported by BBSRC (BBS/E/B/000C0422) and Wellcome Trust (105031/Z/14/Z; 210754/Z/18/Z). Research in the Stegle lab is supported by core funding from EMBL, the BMBF, the Volkswagen Foundation, and the EU (ERC project DECODE 810296).