Fig. 5

α-DR3 stimulates GM-CSF production from ILC3s through p38 signaling. a, b Rag1^–/–Rorc^gfp/+ mice were treated with 1 µg of α-DR3 once, and large intestinal LPLs were isolated 3 days later. a Duplicates mRNA of FACS purified ILC3s (Lin^–GFP⁺) cells were exacted and subjected to genome-wide analysis (RNA-seq). Gene ontology enrichment analyses were performed on upregulated genes (red) and downregulated genes (green) of more than 1.5-fold in the α-DR3-treated group, and representative enriched biological processes are shown. b Relative expression of indicated genes in purified ILC3 (Lin^–GFP⁺) was analyzed by real-time RT-PCR and normalized to the PBS group. c ILC3s (Thy1.2^highCD45^intermediate) cells were purified from Rag1^–/– mice and treated with PBS or α-DR3 (250 ng/ml) for 18 h. Expression of indicated genes was analyzed by real-time RT-PCR. d–f Large intestinal LPLs were isolated from Rag1^–/– mice. Cells were treated with PBS, TL1A (100 ng/ml), and α-DR3 (250 ng/ml) in the presence or absence (DMSO) of inhibitors for NF-kB (Bay11-7082), JNK (SP600125), ERK (SCH772984), and p38 (SB203580) for 18 h. Brefeldin A was added for the last 2 h before analysis. Expression of GM-CSF in ILC3s (Lin^–RORγt⁺) was analyzed by flow cytometry and are shown. g Human tonsil lymphocytes were treated with PBS or human recombinant TL1A (100 ng/ml) in the presence or absence (DMSO) of inhibitors for p38 (SB203580) for 18 h. Brefeldin A was added for the last 2 h before analysis. Percentages of GM-CSF expression in ILC3s (Lin^–CD127⁺CD117⁺) was analyzed and are shown. h, i Large intestinal LPLs from Rag1^–/– mice were treated with α-DR3 or PMA plus ionomycin for indicated time interval. h Expression of phosphorylated p38 (p-p38) gated on ILC3s (Lin^–RORγt⁺) was analyzed and are shown in histogram. i MFI (mean fluorescence intensity) of p-p38 gated on ILC3s was normalized to the control group (time point 0), and fold change is shown. Statistical analyses were performed with paired t test (b) and Student’s t test (c). e–g, i The data are means ± SEM. b–i The data are representative of at least two independent experiments. Source data are provided as a Source Data File