Multi-omics characterization of partial chemical reprogramming reveals evidence of cell rejuvenation
- Division of Genetics, Department of Medicine, Brigham and Women’s Hospital, Harvard Medical School, United States
- Department of Cell Biology, Harvard Medical School, United States
- Broad Institute of MIT and Harvard, United States
Figures
Figure 1
Functional effects of partial chemical reprogramming.
(A) Overview of the study. Tail and ear fibroblasts were isolated from young (4-month-old) and old (20-month-old) male C57BL/6 mice, and cryo-stocks were prepared once reaching ~80–90% confluency …
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Figure 1—source data 1
Normalized change in TMRM fluorescence.
- https://cdn.elifesciences.org/articles/90579/elife-90579-fig1-data1-v1.xlsx
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Figure 1—source data 2
Oxygen consumption rates.
- https://cdn.elifesciences.org/articles/90579/elife-90579-fig1-data2-v1.xlsx
Figure 2 with 2 supplements
Effect of partial chemical reprogramming on gene expression.
(A) PCA of bulk RNA-seq samples. Fibroblasts were treated with 2c, 7c, or control for 6 days followed by RNA-seq analyses. (B) Differentially expressed genes. Differentially expressed genes were …
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Figure 2—source data 1
Expression of pluripotency markers.
- https://cdn.elifesciences.org/articles/90579/elife-90579-fig2-data1-v1.xlsx
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Figure 2—source data 2
Splicing damage values.
- https://cdn.elifesciences.org/articles/90579/elife-90579-fig2-data2-v1.xlsx
Figure 2—figure supplement 1
Effect of partial chemical reprogramming on aging-related changes in fibroblast gene and protein expression.
(A) Effect of fibroblast age on gene expression changes. Left panel: association of aging-related changes in fibroblast gene expression with signatures of aging and OSKM reprogramming. Signatures of …
Figure 2—figure supplement 2
Characterization of alternative splicing events.
(A) Observed alternative splicing events. Observed alternative splicing events in control samples (n=4 independent biological replicates) grouped by different types of alternative splicing (A3SS = …
Figure 3 with 2 supplements
Effect of partial chemical reprogramming on protein expression.
(A) PCA of log2 normalized protein abundances. Fibroblasts were treated for 6 days with 2c, 7c, or control followed by mass spectrometry-based proteomic analyses. (B) Global effects of partial …
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Figure 3—source data 1
Differential expression analysis of mRNA-seq experiments following data normalization in edgeR.
- https://cdn.elifesciences.org/articles/90579/elife-90579-fig3-data1-v1.xlsx
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Figure 3—source data 2
Differential expression analysis of TMT proteomics experiments following data normalization.
- https://cdn.elifesciences.org/articles/90579/elife-90579-fig3-data2-v1.xlsx
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Figure 3—source data 3
Gene set enrichment analysis (GSEA) of mRNA-seq experiments.
- https://cdn.elifesciences.org/articles/90579/elife-90579-fig3-data3-v1.xlsx
Figure 3—figure supplement 1
Additional correlation analyses.
(A) Comparisons between protein differential abundance and differential gene expression. Shown here for all treatment conditions (top: 2c vs. control, bottom: 7c vs. control. Left panels: young …
Figure 3—figure supplement 2
GSEA of protein abundances.
(A) GSEA (GObp, HALLMARK, KEGG, and REACTOME) of functions and pathways most significantly altered by partial chemical reprogramming. Shown are the top 25 most significant ontologies in each …
Figure 4
Correlation of gene expression and protein abundance changes with signatures of aging and OSKM reprogramming.
Spearman correlation of gene expression and protein abundance changes induced by partial chemical reprogramming (blue: 7c, green: 2c) with the signatures of OSKM reprogramming (cyan) and aging …
Figure 5 with 1 supplement
Effect of partial chemical reprogramming on the phosphoproteome.
(A) Targeted GSEA of phosphorylation targets for four selected gene ontologies following partial chemical reprogramming. Bar lengths depict normalized enrichment scores. **Benjamini-Hochberg FDR < …
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Figure 5—source data 1
Differential expression analysis of TMT phosphoproteomics experiments following data normalization.
- https://cdn.elifesciences.org/articles/90579/elife-90579-fig5-data1-v1.xlsx
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Figure 5—source data 2
Normalized change in TMRM fluorescence.
- https://cdn.elifesciences.org/articles/90579/elife-90579-fig5-data2-v1.xlsx
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Figure 5—source data 3
Uncropped blots (image files) against GAPDH and Prkaca.
- https://cdn.elifesciences.org/articles/90579/elife-90579-fig5-data3-v1.zip
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Figure 5—source data 4
Uncropped blots (labeled PDF file) against GAPDH and Prkaca.
- https://cdn.elifesciences.org/articles/90579/elife-90579-fig5-data4-v1.zip
Figure 5—figure supplement 1
Effect of partial chemical reprogramming on cellular signaling.
(A) GSEA of phosphopeptide abundances. GSEA of phosphorylated pathways significantly altered by partial chemical reprogramming. We show all ontologies with Benjamini-Hochberg FDR < 0.05 in at least …
Figure 6 with 1 supplement
Effect of partial chemical reprogramming on the metabolome.
(A) Hierarchical clustering (Ward’s method, Euclidean distance) of polar metabolite samples. Fibroblasts were treated for 6 days with 2c, 7c, or control followed by cell scraping and collection in …
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Figure 6—source data 1
HILIC-neg and HILIC-pos normalized metabolomics data.
- https://cdn.elifesciences.org/articles/90579/elife-90579-fig6-data1-v1.xlsx
Figure 6—figure supplement 1
Effect of age on fibroblast metabolome.
(A) PCA of metabolite samples. PCA was performed following normalization and log-transformation of metabolite peak areas. (B) PCA of untreated fibroblasts. PCA was performed following normalization …
Figure 7 with 3 supplements
Effects of partial chemical reprogramming on biological age.
(A) Effects of partial chemical reprogramming on transcriptomic age (tAge). tAge was assessed by using mouse multi-tissue transcriptomic clocks to analyze the bulk RNA-seq data presented in Figure 2 …
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Figure 7—source data 1
Transcriptomic age predictions.
- https://cdn.elifesciences.org/articles/90579/elife-90579-fig7-data1-v1.xlsx
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Figure 7—source data 2
Epigenetic age predictions.
- https://cdn.elifesciences.org/articles/90579/elife-90579-fig7-data2-v1.xlsx
Figure 7—figure supplement 1
Expression of pluripotency markers following OSKM reprogramming of fibroblasts.
Following doxycycline-induced expression of OSKM in young and old fibroblasts, fully-reprogrammed iPSC colonies were stained for pluripotency markers Sox2 (top), Oct4 (middle), and Nanog (bottom). …
Figure 7—figure supplement 2
Aging-related changes in the fibroblast epigenome.
(A) Number of fibroblast CpG sites significantly differentially methylated with age. Shown in cyan and red are sites that experience an increase and decrease in mean methylation with age, …
Figure 7—figure supplement 3
Effect of partial chemical reprogramming on the epigenome.
(A) PCA of DNAm microarray samples. PCA was performed after normalization of array data and filtering of probes. (B) Number of fibroblast CpG sites significantly differentially methylated with 2c …
Author response image 1
Additional files
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Supplementary file 1
Raw genecounts.
Raw genecounts from mRNA-seq experiments obtained following STAR mapping.
- https://cdn.elifesciences.org/articles/90579/elife-90579-supp1-v1.xls
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Supplementary file 2
Proteomics and phosphoproteomics datasets.
Normalized peptide and phosphopeptide abundances obtained from TMT 18plex experiments.
- https://cdn.elifesciences.org/articles/90579/elife-90579-supp2-v1.xlsx
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Supplementary file 3
Splicing raw data.
Observed alternative splicing events.
- https://cdn.elifesciences.org/articles/90579/elife-90579-supp3-v1.xlsx
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MDAR checklist
- https://cdn.elifesciences.org/articles/90579/elife-90579-mdarchecklist1-v1.docx
