Extended Data Fig. 8: Identification of H4K77ac-related pathways and binding proteins.
From: Acetylation profiling by Iseq-Kac reveals insights into HSC aging and lineage decision
a, Fraction of reads falling within peak regions (FRiP, SEACR FDR ≤ 0.05) of H4K77ac CUT&Tag data in young and aged HSCs (n = 20000 HSCs/experiment). b,c, Overlap between genes differentially expressed (downregulated/upregulated) in aged HSCs and genes showing differentially H4K77ac enrichment (TSS ± 3 kb) at their promoters (n = 20000 HSCs/experiment). b, Venn diagrams of overlapping genes. c, Pathway enrichment from Over-Representation Analysis (ORA) of overlapping genes (Fisher’s precision probability tests. p value < 0.01). Blue, downregulated in aged HSCs; Orange, upregulated in aged HSCs. d, Relative mRNA expression levels of signature genes in young (blue) and aged (orange) HSCs, as determined by transcriptome. e, Distribution of ATAC-seq reads (OCRs) at transcription start sites (TSS ± 3 kb) of genes annotated by common peaks (Fig. 4g) associated with lymphocyte differentiation (GO:0030217 for T cell differentiation, GO:0030183 for B cell differentiation) or myeloid cell development (GO:0030099 for myeloid cell differentiation) between young and aged HSCs promoters (n = 20000 HSCs/experiment). f, Distribution of H4K77ac CUT&Tag reads at the transcription start site (TSS ± 3 kb) of genes associated with lymphocytes (GO:0030217 and GO:0030183 gene sets for T and B cell differentiation, respectively) or myeloid cell development (GO:0030099 gene sets for myeloid cell differentiation) in young (left) or aged HSCs (right) promoters (n = 20000 HSCs/experiment). g, Comparative analysis of proteins pulled down by acetylated H4K77 peptide or its unmodified form. h, Immunoblotting showing H4K77ac levels in HCT116 cells treated with HDAC3 inhibitor (RGFP966) or SIRT2 inhibitor (Thiomyristoyl) i, Representative data showing the frequencies of T lymphocytes (CD4-CD8-, CD4+CD8+, CD4+ and CD8+) derived from aged HSCs, determined at 2 weeks post-RGFP966 or DMSO treatment. j, ShRNA-Hdac3-mediated gene knockdown efficiency and H4K77ac levels in K562 cells, assessed by immunoblotting. Data are presented as the median ± 25% (a,e,f) or mean ± s.e.m. (d). All data are derived from three independently biological replicates (a,d,e,f). Representative blots from three independent experiments (h,j). Statistical significance was assessed using the two-side Wilcoxon rank sum test (d) or two-side Kruskal-Wallis rank sum test (e,f).
