Fig. 2: CAF-1 depletion induces rapid and partial differentiation of iGMPs.
From: Regulation of chromatin accessibility by the histone chaperone CAF-1 sustains lineage fidelity
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.
