(A) Gating strategy of subpopulations of G1-ILCs (CD3−NK1.1+NKp46+) in the E18.5 fetal liver (FL), adult liver (AL), and bone marrow (BM). iILC1, immature ILC1. Data represent three independent experiments (FL, n=15; AL, n=8; BM, n=6). (B) Heatmap representing log2 transformed mean fluorescence intensity (MFI) of indicated protein expression normalized by z-score transformations (n=3 for all subsets). SP, spleen. (C) Expression of IL-7Rα on G1-ILCs except for CD49a−CD49b+ NK cells (non-NK G1-ILCs) in FL, AL, and BM. Data represent three independent experiments (FL, n=15; AL, n=8; BM, n=6). (D) The percentages of IL-7R− fractions in non-NK G1-ILCs in the indicated tissues. LPL, small intestinal lamina propria lymphocytes; PC, peritoneal cavity; mLN, mesenteric lymph node; SG, salivary gland. Data are pooled from three independent experiments (FL, n=15; AL, n=8; BM, n=6; LPL, n=6; PC, n=6; SP, n=6; mLN, n=6; SG, n=6). (E) Scatter plot showing relative gene expression of FL G1-ILCs and AL 7 R− ILC1s compared to AL NK cells in RNA-seq. Genes differentially expressed by FL G1-ILCs, AL 7 R− ILC1s, AL 7R+ ILC1s, and BM iILC1s (orange) or only by FL G1-ILCs and AL 7 R− ILC1s (blue) compared to AL NK cells are highlighted. FC, fold change. (F) First three principal components in PCA of top 3,000 variant genes. (G) Expression of T-bet (upper) and Eomes (lower) on FL G1-ILCs as well as NK cells, 7 R− ILC1s, and 7R+ ILC1s in AL. Shaded histograms (grey) indicate isotype controls. Data represent two independent experiments. (H and I) Fate-mapping analysis of adult chimeric mice reconstituted with BM YFP−Lin−Sca1+c-Kit+ (LSK) cells from PLZFGFPcre/+ Rosa26-YFP (PLZF-fm) mice. Representative histograms of YFP expression (H) and the percentages of YFP+ cells in indicated cell populations (I) are shown. Data represent or are pooled from two independent experiments (n=6 for AL T, AL NK, AL 7 R− ILC1, AL 7R+ ILC1, and SP ILC1; n=3 for BM NK and BM iILC1). (J) The percentage of YFP+ cells in FL G1-ILCs in E18.5 straight PLZF-fm mice. Data are from one experiment (n=5). RNA-seq data are from two (AL NK cells and SG G1-ILCs), three (AL 7R+ ILC1s, BM iILC1s, and BM NK cells), and four (FL G1-ILCs, AL 7 R− ILC1s, and LPL ILC1s) biological replicates (E and F). Data are presented as mean ± SD.