Extended Data Fig. 1: Design and validation of the iTreg whole-genome CRISPR screen in primary human T cells.
From: Genome-wide CRISPR screen in human T cells reveals regulators of FOXP3
a, Detailed timeline schematic of the CRISPR screening pipeline. The schematic of the cells was adapted from ref. 51, Springer Nature, and the graphic of the next-generation sequencing was created using BioRender (https://biorender.com). b, Gating strategy used for the FOXP3 screen in a. c, Scatter plot of gene-level LFC (between FOXP3hi and FOXP3lo sorting bins), comparing screens between two replicates. d, Gene-set enrichment analysis of top-ranked FOXP3 negative (left) and positive (right) screen hits in a curated list from a previous study13. FDR = False discovery rate, permutation test. Representative unique and shared hits are shown in the text-box on the right. e, Western Blot analysis of protein expression of RBPJ, HDAC3 and NCOR2 in cells treated with the indicated perturbations. Data is representative of two independent donors (n = 4 donors). f, Representative histograms depicting FOXP3 expression in iTregs with indicated genetic perturbations, related to Fig. 1e,f. Negative (red) and positive (blue) regulators are overlaid on NTC (gray). MFI, median fluorescence intensity.
