Fig 2: Loss of Cemip Enhances Local Skin Inflammation in Response to S. aureus(A–F) Flow cytometry analysis of single-cell suspensions from the skin showing expression of CD11c/MHCII, Ly6G/CD11b, and LyG-C cells isolated from control, Cemip-/- control infection, and Cemip-/- infection. Cells were gated on CD3-negative. Numbers represent the percentages of the cells in the indicated gate.(G) Representative sections of skin from control and Cemip-/- mice at 3 days after S. aureus infection. Tissues are stained with red with Gr-1 antibody and blue with DAPI. Scale bar, 20 µm.(H) qRT-PCR of the relative abundance of transcripts for IL-6 as normalized to ß-actin (n = 4 for normal condition, n = 8 for infection mice/group).All error bars indicate mean ± SEM. *p < 0.05, **p < 0.01, and ***p < 0.001 (t test).

Fig 3: CAF-1 depletion induces rapid and partial differentiation of iGMPs.a Western blot time-course analysis of CHAF1A and CHAF1B in CAF-1 OFF and HOXA9 OFF conditions showing a closely matching downregulation trend of both CAF-1 subunits. Two independent experiments were performed with similar results, but one representative experiment is shown. b Quantification of the western blot time course in a. c Flow cytometric time-course analysis showing activation of three neutrophil markers (GFP, CD11b, and Gr1) in CAF-1 OFF and HOXA9-OFF conditions. Mean and standard deviation of marker positive percent population is plotted for n = 3 experimental replicates. See corresponding representative histograms in Supplementary Fig. 2b. d ATP growth assay time course of uninduced, CAF-1 OFF and HOXA9 OFF cells. Mean and standard deviation of log10 transformed relative luminescence units (RLU) is plotted for n = 3 experimental replicates. Two-way ANOVA with Dunnett’s test. e Top: Schematic of cellular differentiation commitment assay in either CAF-1 OFF or HOXA9 OFF conditions. Shown are CAF-1 OFF (+IPTG) or HOXA9 OFF (-E2) pulses of 24 h increments followed by chase periods of no induction. Bottom: Mean and standard deviation of % GFP+ population from n = 3 experimental replicates. Each bar represents a “pulse+chase” condition analyzed 6 days post-induction. Two-way ANOVA with Dunnett’s test. *p < 0.0001. f Morphological analysis of clones expressing shCTRL or shChaf1b in uninduced or 96 h post-induction of each treatment. Scale bar = 10 µm. Three independent experiments were performed with similar results, but one representative experiment is shown. g Phagocytosis flow cytometric assay. Density plots of shChaf1b expressing clone in uninduced and 96 h post-induction of each treatment showing GFP signal versus fluorescently labeled bacterial particles (pHrodo). pHrodo positive cells are gated. h Quantification of % pHrodo+ populations shown in g for two independent subclones. Two independent experiments were performed with similar results, but one representative experiment is shown. Source data are provided as a Source Data file.

Fig 4: Administration of IL-13 or rIL-4 increases the infiltration of macrophages at 3 days after SCI. (A) Representative flow cytometry density plots showing CD45+ and CD11b+ cells in the spinal cord after treatment with PBS, rIL-4, and rIL-13. Note that rIL-4 and rIL-13 led to the appearance of resolution-phase macrophages (square) absent in PBS. (B) Plot showing the counts of microglia, macrophages, and granulocytes. (C) Plot showing the counts of resolution phase-macrophages. Data are presented as mean ± SEM. N = 4 per group. * p < 0.05, *** p < 0.001 against PBS. & p < 0.05 against rIL-4. One-way ANOVA with Tukey's post hoc test.

Fig 5: Contribution of Lyl-1-expressing MF-progenitors to the fetal liver and brain.A All MF-progenitors from E10-Brain expressed Lyl-1, contrary to the corresponding YS (Fig. 1b) which harbor both FDG+/Lyl-1+ and FDG-/Lyl-1- subsets. MF-progenitors from E10 Lyl-1LacZ/LacZ FL harbored both FDG+/Lyl-1+ and FDG-/Lyl-1 MF-progenitor subsets. in E10-brain, mature MFs (CD11b+F4/80+ gate) were all FDG+/Lyl-1+. The contour plots in WT samples indicate the level of non-specific background ß-Gal activity/FDG labeling in WT samples. Representative profiles of 3 independent samples, each consisting of 3-4 E10-Brain or 8-12 E10-FL. B Quantification of Kit+CD45+CD11b+ MF-progenitors in E10-FL (plots show mean ± s.e.m.; Unpaired, two-tailed t-Test). C Lyl-1 marks the entire F4/80 + microglia/BAM population from the onset of brain colonization. FDG/Lyl-1 expression in F4/80+ microglia/BAM from the brain of Lyl-1WT/LacZ and Lyl-1LacZ/LacZ embryos at E9 to E11. The rare CD11b+ F4/80low-neg cells present in the brain at E9 are FDG+/Lyl-1+. Gray histograms indicate non-specific background ß-Gal activity/FDG levels in WT samples. Representative profiles of 3–5 independent samples, each consisting of 3-5 brains. D MF-progenitors from E10-brain express Myb levels similar to E9-YS MFPrim progenitors. RT-qPCR quantification of Myb expression levels in Kit+CD45+CD11b+ MF-progenitors sorted from WT E9-YS and from WT and Lyl-1WT/LacZ brain at E10. Lin-Sca+cKit+ (LSK) progenitors from WT E12-FL were used as positive control. Myb expression levels, shown on a Log2 scale, were normalized to the mean expression value obtained for WT E10-YS, considered as 1 (error bars show mean ± s.e.m.; Unpaired, two-tailed t-Test). E RT-qPCR analyses of Lyl-1 expression in A1 to A3 MF subsets isolated from Cx3cr1WT/GFP brain at E10. Lyl-1 is expressed by the 3 subsets, with levels decreasing with differentiation. Expression levels were normalized to the mean value obtained for Cx3cr1WT/GFP YS A1 progenitors (n = 2; Error bars show mean ± s.e.m). F Defective differentiation of brain MF-progenitor in Lyl-1 mutant embryos. Distribution of A1-A2 and A3 MF subsets in E10 brain from Cx3cr1WT/GFP:Lyl-1WT/WT, Cx3cr1WT/GFP:Lyl-1WT/LacZ and Cx3cr1WT/GFP:Lyl-1LacZ/LacZ embryos. The size of the whole MF population was similar in the three genotypes, but Lyl-1 deficiency modified the distribution of the MF subsets with an increased size of the A1 subset and a reduced A3 pool (5-12 independent analyses, 6–8 brains per sample. Error bars show mean ± s.e.m.; Unpaired, two-tailed t Test). G Heatmap expression profile of the genes that mark the development of tissue resident-MFs in WT E9 and E10 MF-progenitors (Heatmap displays transformed log2-expression values; Unpaired, two-tailed t-test).