Figure 5.
Deregulation of genes involved in cell cycle and HSC homeostasis in PROSER1-deficient LSK cells. (A) Schematic showing experimental setup to isolate wild-type and PROSER1-deficient LSK cells from BM chimeric animals. C57BL/6 recipients were lethally irradiated. (B) Principal component analysis plot of RNA seq data from biological replicate samples of wild-type (n = 4) and PROSER1 KO (n = 3) LSK cells. (C) MA plot showing differential expression analysis of wild-type and PROSER1 KO LSK cells. Red and blue symbols indicate significantly differentially upregulated and downregulated genes, respectively (threshold, P-adj < .05; FC > 1.5). The total number of differentially regulated genes in each category are indicated. See supplemental Table 3 for full list of differentially expressed genes. (D) Differential expression of selected genes with known roles in HSC homeostasis. Bars represent log2 fold change of expression in PROSER1 KO relative to wild-type LSK. Error bars represent the estimated standard error from DESeq2 output. Statistical significance was measured by Benjamini-Hochberg adjusted P value. ∗P-adj < .05; ∗∗P-adj < .01; ∗∗∗P-adj < .001; ∗∗∗∗P-adj < .001. (E) Enrichment scores (GSEA) of PROSER1 KO vs wild-type LSK cells in gene sets associated with cell cycle regulation and HSC quiescence. NES, gene ratios, and –log10 FDR values are displayed for each gene signature. (F) Illustration of the roles of PROSER1 and TET2 in regulation of DNA methylation and hematopoiesis. PROSER1 is a pan-TET interactor important for maintenance of DNA methylation homeostasis in hematopoietic cells. Loss of PROSER1 results in disruption of TOPD complexes and overlapping and distinct enhancer DNA methylation changes compared to changes observed in TET2 KO cells. These DNA methylation changes and the resulting deregulation of genes governing cell cycle and HSCs likely drive the observed exhaustion of HSC activity over time. GSEA, gene set enrichment analysis; FDR, false discovery rate; FC, fold change; NES, normalized enrichment scores; WT, wild-type.

Deregulation of genes involved in cell cycle and HSC homeostasis in PROSER1-deficient LSK cells. (A) Schematic showing experimental setup to isolate wild-type and PROSER1-deficient LSK cells from BM chimeric animals. C57BL/6 recipients were lethally irradiated. (B) Principal component analysis plot of RNA seq data from biological replicate samples of wild-type (n = 4) and PROSER1 KO (n = 3) LSK cells. (C) MA plot showing differential expression analysis of wild-type and PROSER1 KO LSK cells. Red and blue symbols indicate significantly differentially upregulated and downregulated genes, respectively (threshold, P-adj < .05; FC > 1.5). The total number of differentially regulated genes in each category are indicated. See supplemental Table 3 for full list of differentially expressed genes. (D) Differential expression of selected genes with known roles in HSC homeostasis. Bars represent log2 fold change of expression in PROSER1 KO relative to wild-type LSK. Error bars represent the estimated standard error from DESeq2 output. Statistical significance was measured by Benjamini-Hochberg adjusted P value. ∗P-adj < .05; ∗∗P-adj < .01; ∗∗∗P-adj < .001; ∗∗∗∗P-adj < .001. (E) Enrichment scores (GSEA) of PROSER1 KO vs wild-type LSK cells in gene sets associated with cell cycle regulation and HSC quiescence. NES, gene ratios, and –log10 FDR values are displayed for each gene signature. (F) Illustration of the roles of PROSER1 and TET2 in regulation of DNA methylation and hematopoiesis. PROSER1 is a pan-TET interactor important for maintenance of DNA methylation homeostasis in hematopoietic cells. Loss of PROSER1 results in disruption of TOPD complexes and overlapping and distinct enhancer DNA methylation changes compared to changes observed in TET2 KO cells. These DNA methylation changes and the resulting deregulation of genes governing cell cycle and HSCs likely drive the observed exhaustion of HSC activity over time. GSEA, gene set enrichment analysis; FDR, false discovery rate; FC, fold change; NES, normalized enrichment scores; WT, wild-type.

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