Figure 5. Intrinsic IFNγ signaling initiates the lung-BRM differentiation program in GC B cells.

(A-C) WT and IfngR1^−/− GC B cells were sorted from WT/IfngR1^−/− BM chimeras on day 12 after PR8 infection, and RNA-seq was performed. (A) PCA of normalized gene expression in WT and IfngR1^−/− GC B cells (B) GSEA for the memory B cell gene signature (Table S2) in WT vs. IfngR1^−/− GC B cells. (C) Volcano plot highlighting genes differentially expressed in WT vs. IfngR1^−/− GC B cells. Three replicates for each cell type were obtained from three independent experiments. (D) Frequency of CXCR3^hi cells within WT and IfngR1^−/− GC B cells from PR8-infected WT/IfngR1^−/− BM chimeras on day 12 after infection. Data are representative of three independent experiments (n=5-6 mice/time point). P values were determined using a two-tailed Student’s t-test. *p < 0.05, **p < 0.01, ***p < 0.001; ns, not significant. (E-F) CXCR3^hi and CXCR3^hi GC B cells were sorted from the med-LN of day 12 PR8-infected B6 mice, and RNA-seq was performed. GSEA for the GC IFNγ-induced program (Table S1) (E), the memory B cell gene signature (Table S2) (F), and the lung-BRM gene signature ⁴ (G) in CXCR3^hi vs CXCR3^hi GC B cells. Three replicates for each cell type were obtained from three independent experiments. (H-K) Class-switched GC B cells from the med-LN of PR8-infected B6 mice and paired lung-BRMs (CXCR3⁺class-swicthed memory B cells) were sorted on day 30 and RNA-seq was performed. (H) IPA showing the activation Z-score for the top -ranked upstream regulators in BRM vs. GC B cells. (I) Top hallmark pathways significantly enriched in the BRM transcriptome. GSEA for the hallmark IFNγ signatures (J) and the IFNγ-induced program (Table S1) in BRM vs. GC B cells. Three replicates for each cell type were obtained from three independent experiments.