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. Author manuscript; available in PMC: 2018 Sep 19.
Published in final edited form as: Immunity. 2017 Sep 19;47(3):435–449.e8. doi: 10.1016/j.immuni.2017.08.012

Figure 4. Rroid promotes group 1 ILC maturation and lineage identity through Id2.

Figure 4

A. Volcano plot of RNA-seq results in sorted splenic DP (CD27+, CD11b+) Rroid+/+ and Rroid−/− NK cells. Genes with FDR<0.05 are depicted in blue (n=3 mice per group).

B. Id2 expression in maturing NK cell populations sorted from spleen and analyzed by qPCR (n=3 mice per group). Normalized to Hprt expression. Data are representative of two independent experiments.

C. qPCR of Id2 expression in sorted liver NK cells and ILC1s (n=2 of 5 pooled mice per group). Normalized to Hprt expression.

D. Id2 expression in sorted ILC3s from small intestine (left) or IL-33 expanded ILC2s from lung (right) analyzed by qPCR (For ILC3s, n=4 mice per group pooled from two experiments; for ILC2s, n=5 mice per group from one experiment). ILC2s were expanded by intranasal administration of 250ng IL-33 daily for 4 days and sorted 24 hours following the last injection. Normalized to Hprt expression.

E. Gene Set Enrichment Analysis (GSEA) depicting enrichment of known Id2-regulated genes in Rroid−/− NK cells.

F. GSEA depicting enrichment of KEGG T cell receptor signaling pathway genes in Rroid−/− NK cells.

G. Venn diagram depicting unique ATAC-seq peaks called in sorted splenic Rroid+/+ and Rroid−/− NK cells (n=3 mice per group).

H. Heatmap of transcription factor footprinting analysis. Rroid+/+ and Rroid−/−-specific ATAC-seq peaks were subjected to transcription factor footprinting analysis by the Protein Interaction Quantitation (PIQ) algorithm.

I. Enrichment of cellular pathways associated with open chromatin elements in Rroid+/+ and Rroid−/− NK cells.

J. Schematic of Id2 transduction experiment. Rroid−/− bone marrow was transduced in vitro with retrovirus bearing Id2 or empty vector control and used to reconstitute lethally irradiated CD45.1+ hosts. Transduced cells were identified by expression of GFP.

K. NK cells as a proportion of GFP+ cells in bone marrow chimeras reconstituted with either empty retrovirus or Id2 retrovirus (n=4 mice per group).

L. Frequency of maturing splenic NK cells in bone marrow chimeras reconstituted with either empty retrovirus or Id2 retrovirus (n=4 mice per group).

*p≤0.05, **p≤0.01, ***p≤0.001. Two-tailed t-test. All error bars represent SEM. See also Figure S4 and Table S1.