Fig. 6: IRF4-mediated transcriptional programming of human LCs.

Knockdown (KD) of IRF4 using CRISPR-Cas9 editing. An IRF4 guide–CAS9 complexes were introduced by nucleofection into migrated LCs. Indicated analyses were performed 48 h after nucleofection. a Intranuclear expression levels of IRF in CRISPR-Cas9 edited (orange) and control (blue) migrated LCs measured using flow cytometry in CD207+ CD1a+ live LCs. A representative example of n = 3 independent donors. b Quantification of the intranuclear expression levels of IRF4 in CRISPR-Cas9 edited (orange) and control (blue) migrated LCs. Mean fluorescence intensity (MFI) of IRF4-expressing CD207+ CD1a+ live LCs shown, n = 3 independent donors, paired t test, box and whiskers show min and max value, line at median. Source data are provided as a Source Data file. c scRNA-seq analysis of WT and IRF4 KD migrated LCs. UMAP plot of 1484 LCs (WT:617 cells, blue, KD, 867 cells, orange) (filtering setting gene count 500–10,000 per cell, expression of mitochondrial genes <0.2, Leiden r = 0.2). d GO processes and pathways differentially enriched in control LC (WT, left) versus IRF4 CRISPR-Cas9 edited LCs (IRF4 KD, right). DEG analysis was performed using Single TK package, scDiffExlimma algorithm, scnorm data, FDR-corrected P-value used as a significance measure. Gene ontology enrichment: ToppGene tool, Biological Processes and Molecular Function. Source data are provided as a Source Data file. e Barplots of genes expressed at higher levels in WT LCs. Each bar shows CPTT normalised expression level of indicated gene in a given WT LC cell (blue) or IRF4 CRISPR-Cas9 edited LC (orange). IRF4 plus top five genes by FDR-corrected P-value and five genes representative for biological processes enriched in WT are displayed. f Barplots of genes expressed at higher levels in IRF4 KD LCs. Each bar shows CPTT normalised expression level of indicated gene in a given WT LC cell (blue) or IRF4 CRISPR-Cas9 edited LC (orange). Top five genes by FDR-corrected P-value and five genes representative for biological processes enriched in KO are displayed.