Chromatin dynamics during the differentiation of long-term hematopoietic stem cells to multipotent progenitors

Abstract

ATAC-seq provides genome-wide chromatin state in 3 cell types of hematopoietic stem/progenitor cells.Transcription factor cohorts are associated with dynamic changes of open chromatin during the differentiation of LT/ST-HSCs to MPPs.

Graphical abstract

Figure 1.

ATAC-seq provides the open chromatin landscape of LT-HSCs, ST-HSCs, and MPPs. (A) Representative flow cytometry results to assess the Sca1⁺c-Kit⁺ population of cells, then to further distinguish LT-HSCs, ST-HSCs, and MPPs using the CD150 and CD48 combination of markers. (B) PCA for read counts of bins genome-wide among ATAC-seq samples. (C) Scatterplot and Pearson correlation of ATAC-seq data between 2 biological replicates for each of the 3 HSC cell types (LT-HSCs, ST-HSCs, and MPPs). (D-F) Browser track plot of ATAC-seq signal (RPM) in the genomic regions around CD34, Flt3/Flk2, and Hoxb5, respectively. (G) Gene expression activity of CD34, Flt3/Flk2, and Hoxb5 in different types of blood cells. Expression data collected from Gene Expression Commons (https://gexc.stanford.edu). Gene expression activity ranged from −100 (most inactive) to 100 (most active). MEP, megakaryocyte-erythroid progenitor.

Figure 2.

The open chromatin state of protein coding genes and miRNAs changes dynamically in HSCs. (A) Classification of the genomic features in an open chromatin configuration as determined by our ATAC-seq analysis. TTS, transcription termination site. (B) The overlap of genes with an open chromatin configuration in their promoter-TSS region among LT-HSCs, ST-HSCs, and MPPs. (C) Heat map showing enrichment (-log₁₀ [FDR]) of GO terms in gene set with ATAC-seq peaks found in their promoter regions. (D) Heat map of ATAC-seq peak scores ranking by LT-HSCs for the promoter-TSS regions of miRNAs. FDR, false discovery rate.

Figure 3.

Chromatin remodeling and enrichment of DNA cis-elements in the promoter regions upon HSC differentiation to MPPs. (A) Heat map of reads density around gene promoters displaying an open chromatin state (from −1000 bp to 1000 bp) in LT-HSCs, ST-HSCs and MPPs, respectively. (B) The average reads density (smoothed) around gene promoters displaying an open chromatin state (from −1000 bp to 1000 bp) in LT-HSCs, ST-HSCs, and MPPs, respectively. (C) Heat map of significant enrichment value (-log [P value]) of transcription factor–binding motifs in gene promoters displaying an open chromatin state. (D) PCA for the enriched DNA motifs in open promoters within the 3 cell types. The first 2 components account for 90.67% and 9.33% of the total variance in the dataset, respectively. Dots represent the PCA loadings for the 166 transcription factors. The different colored dots represent 3 different cohorts. The motif of Krüppel-like factors (Klf9/Klf10) is displayed (top left). (E) Gene expression activity of Klf9 and Klf10 in LT-HSCs, ST-HSCs, and MPPs. Gene Expression Commons (https://gexc.stanford.edu). Gene expression activity ranged from −100 (most inactive) to 100 (most active).

Figure 4.

Enrichment of DNA cis-elements in enhancers that display an open chromatin configuration in hematopoietic cells. (A) The overlap of enhancers displaying an open chromatin state in LT-HSCs, ST-HSCs, and MPPs. (B) The overlap of enriched transcription factor–binding motifs found in both promoters and enhancers that display an open chromatin state in our ATAC-seq analysis. (C) PCA for the enriched DNA motifs in open enhancers within the 3 cell types. The first 2 components account for 99.21% and 0.79% of the total variance in the dataset, respectively. Dots represent the PCA loadings for the 153 transcription factors. The different colored dots represent 3 different cohorts. (D-H) Metaplot of ATAC-seq signal around the flanking 1 kb of PU.1, SPIB, ELF5, CCCTC-binding factor (CTCF), and BORIS binding motifs. (I) Venn diagram showing the overlap of poised enhancers in LT-HSCs and ST-HSCs and active enhancers in MPPs.