Fig. 1: Comparative inducible CRISPRi screens identify essential components of the mRNA translation machinery in human cells.

a, Schematic of inducible CRISPRi cell line generation and screening. Right, gene counts for major functional groups of the human translation machinery included in the sgRNA library. Neo, neomycin resistance gene; mCh, mCherry; Puro, puromycin resistance gene; KRAB, Krüppel associated box; rtTA, reverse tetracycline-controlled transactivator; mKate, monomeric Kate2 protein. b, Schematic of the workflow of inducible hiPS cell differentiation into NPCs, neurons and CMs and the different types and durations of inducible CRISPRi screens. c, Principal component analysis of variance stabilizing-transformed count data for sgRNAs from DESeq2 (n = 2 biological replicates for CM screens; n = 3 biological replicates for inducible hiPS cell, inducible HEK293 cell, NPC and neuron screens). PC, principal component; diff, differentiation; sur, survival. d, Volcano plots of gene-level sgRNA log₂ fold change (mean of the top three sgRNAs per gene by magnitude) and P values (from comparisons of all sgRNAs targeting a given gene to all negative control sgRNAs) for each screening condition relative to matched uninduced (−Dox) controls. Genes with significant (two-sided Mann–Whitney test, P ≤ 0.1) positive or negative enrichment scores are shown in green and purple, respectively. e,f, UpSet plots showing overlap of genes with significant (two-sided Mann–Whitney test, P ≤ 0.1) negative enrichment scores in inducible hiPS cell, inducible HEK293 cell and NPC screens in comparison to common essential genes in cancer cell lines (DepMap¹⁴) and genome-wide CRISPRi in the WTC11 hiPS cell line³⁵ (e) and neuron differentiation or survival screens compared to a genome-wide CRISPRi screen in WTC11-derived i³Neurons³⁶ (f). hiPSCi, inducible hiPS cell; HEK293i, inducible HEK293 cell.

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