Journal: bioRxiv
Article Title: Nucleosome stability safeguards cell identity, stress resilience and healthy aging
doi: 10.1101/2025.09.17.676776
Figure Lengend Snippet: a, Structural location of histone H2B residues D68 and E76 mutated in this study. H2B is shown in magenta, H4 in cyan. b, Nucleosome stability assessed by FRAP in IMR-90 SV40-T cells expressing EGFP-tagged H2B WT or mutants. Scale bar, 10 μm. Relative intensity was calculated by comparing the bleached site to the unbleached site. Data represent mean ± s.d. from >15 nuclei. The plateau value ( Ymax ) was calculated from the fitted recovery curve. P -values were calculated using unpaired, two-tailed Student’s t -tests comparing each mutant to WT at the final time point. c, Histone turnover in live cells assessed using SNAP-tagged H2B WT or mutants labeled with SNAP-TMR. Data represent mean ± s.d. of total field fluorescence from >2600 cells across three independent wells per condition. Half-life ( t 1/2 ) was calculated from the fitted decay curve. P -values were calculated using unpaired, two-tailed Student’s t -tests comparing each mutant to WT at the final time point. d, PCA of bulk mRNA-seq from skeletal muscle expressing H2B WT, mutants or empty vector. Each dot represents an individual mouse (n = 6, Empty; 5, WT; 4, D68N; 5, E76K; 5, E76R). e, GSEA of hallmark pathways in transduced skeletal muscle comparing each mutant to Empty and WT. (See for full pathways). f, IPA upstream regulator analysis showing suppression of Myod1- or Six1-associated genes and activation of Tnf- or Nfkb-associated genes across mutant-expressing skeletal muscle compared to Empty and WT. g, GSEA of Tabula Muris Senis aging signatures in transduced skeletal muscle comparing all mutants to Empty and WT. h, PCA of bulk mRNA-seq from heart expressing H2B WT, mutants or an empty vector. Each dot represents an individual mouse (n = 6, Empty; 5, WT; 4, D68N; 5, E76K; 5, E76R). i, GSEA of hallmark pathways in transduced heart comparing each mutant to Empty and WT. (See for full pathways). j, IPA upstream regulator analysis showing suppression of Ppara- or Ppargc1a-associated genes and activation of Tgfb1- or Smad3-associated genes across mutant-expressing heart compared to Empty and WT. k, GSEA of Tabula Muris Senis aging signatures in transduced heart comparing all mutants to Empty and WT. l, Schematic of experimental design. MSCV-based retroviral vectors expressing H2B WT or mutants were introduced into HSPCs (Lin⁻Sca-1⁺c-Kit⁺), and myeloid or megakaryocyte/erythroid differentiation was assessed. m, GSEA of hallmark pathways in transduced HSPCs comparing each mutant to Empty and WT. (See for full pathways). n, IPA upstream regulator analysis showing suppression of Cebpa- or Cebpb-associated genes (myeloid regulators) and activation of Gata1- or Gata2-associated genes (megakaryocyte/erythroid regulators) in mutant-expressing HSPCs 9 days after differentiation. o, GSEA of cell type signatures indicating increased megakaryocyte and erythroid lineage features and decreased myeloid (neutrophil, monocyte and macrophage) features in mutant-expressing cells compared to Empty and WT.
Article Snippet: Cell type signature profiles using the mouse aging cell atlas, Tabula Muris Senis , revealed that mutant-expressing skeletal muscle recapitulated aging-like transcriptional signatures across macrophages, T cells, muscle satellite cells, mesenchymal stem cells and B cells ( ).
Techniques: Expressing, Two Tailed Test, Mutagenesis, Labeling, Fluorescence, Plasmid Preparation, Activation Assay, Retroviral
