Multiome-based identification of HSC subtype markers

Individual haematopoietic stem cells (HSCs) display heterogeneous capacities to reconstitute the major blood lineages, including platelets, erythrocytes, myeloid cells, B cells, and T cells. Single-cell transplantation assays have revealed that the HSC compartment contains lineage-restricted subtypes capable of reconstituting only specific lineages, such as platelets, alongside true multi-lineage HSCs. The proportion of these lineage-restricted HSCs increases with age, contributing to declined immunity and potentially to clonal haematopoiesis. To date, their identification and isolation have primarily relied on transgenic reporters, such as the Vwf-EGFP mouse line, which offer only moderate accuracy. In this study, we profiled highly purified adult mouse long-term HSCs (LT-HSCs; LSK CD48⁻ CD150⁺ CD34⁻) using the DOGMAseq platform, which enables simultaneous measurement of transcriptome (RNA-seq), chromatin accessibility (ATAC-seq), and surface protein expression (CITE-seq). We subsequently identified candidate clusters corresponding to platelet-biased and multi-lineage HSCs, and developed a fluorescence-activated cell sorting (FACS) strategy to enrich these populations. This strategy employs four surface markers detectable with commercially available antibodies and achieves higher enrichment specificity than the Vwf-EGFP reporter.