Fig. 4: Transcriptome analysis of CNS Treg cells during different phases of EAE.
EAE was induced in Foxp3 (GFP) reporter mice. Tconv and Treg cells were sorted from the spleen and CNS during peak EAE or the recovery phase (rec) and were subjected to bulk RNA-seq. a, PC analysis. Each symbol represents an individual mouse. b,c, Volcano plots showing genes with annotation to the Gene Ontology (GO) terms ‘GO:0009986 cell surface’ (b) and ‘GO:0003700 DNA binding transcription factor activity’ (c) with differential expression (≥1.5-fold, adjusted P < 0.05) between recovery phase and peak CNS Treg cells. DEGs are highlighted in orange. Adjusted P values were derived from DESeq2 using a two-tailed Wald’s test with correction for multiple testing using the Benjamini and Hochberg method (CNS Treg peak n = 4, CNS Treg recovery phase n = 5 biological replicates); TF, transcription factor. d, Heat map with hierarchical clustering of DEGs between CNS recovery phase and peak Treg cells. DEGs with the same phase-dependent regulation in CNS Tconv cells as in CNS Treg cells were subtracted to yield a CNS Treg private gene signature. e, Upregulated DEGs from the recovery phase CNS Treg private signature were used to perform GO enrichment analysis (gprofiler 2g:GOSt). P values are derived from Fisher’s one-tailed test, and adjusted P values were determined by the g:GOSt implemented g:SCS method for multiple testing correction. The top five enriched terms are marked and summarized in the table on the right; MF, molecular function; CC, cellular component; BP, biological process; KEGG, Kyoto Encyclopedia of Genes and Genomes; REAC, Reactome; WP, WikiPathways; Reg., regulation; multicell., multicellular; org., organismal; proc., process; Pos., positive; biol., biological. f,g, Volcano plots displaying only genes that are part of both the GO terms ‘response to stress’ (GO:0006950) and ‘cell surface’ (GO:0009986; f) or ‘DNA-binding transcription factor activity’ (GO:0003700; g) between CNS recovery phase and peak Treg cells. DEGs (≥1.5-fold, adjusted P < 0.05) from the recovery phase CNS Treg private ‘up’ signature are highlighted in purple. Adjusted P values are derived from DESeq2 using a two-tailed Wald’s test with correction for multiple testing using the Benjamini and Hochberg method.
