Differences in cell cycle status underlie transcriptional heterogeneity in the HSC compartment

Hematopoietic stem cells (HSCs) are considered a heterogeneous cell population. In an attempt to further resolve the HSC compartment, we characterized a retinoic acid (RA) reporter mouse line. Sub-fractionation of the HSC compartment in RA-CFP reporter mice demonstrated that RA-CFP-dim HSCs were largely non-proliferative and displayed superior engraftment potential in comparison to RA-CFP-bright HSCs. Gene expression analysis demonstrated higher expression of RA-target genes in RA-CFP-dim HSCs, contrasting the RA-CFP reporter expression, but both RA-CFP-dim and –bright HSCs responded efficiently to ATRA in vitro. Single-cell RNA-sequencing of >1,200 HSCs showed that differences in cell cycle activity constitutedthe main driver of transcriptional heterogeneity in HSCs. Moreover, further analysis of the single-cell RNA-seq data revealed that stochastic low-level expression of distinct lineage-affiliated transcriptional programs is a common feature of HSCs. Collectively, this work demonstrates the utility of the RA-CFP reporter line as a tool for the isolation of superior HSCs. - Whole genome bisulphite sequencing of approximately 1000 CFP dim HSCs versus CFP bright HSCs were generated in duplicates, using Illumina Next Seq 500. - RNA-seq of HSCs. - Single cell RNA-seq of HSCs.