Simultaneous lineage and transcriptome analysis of hematopoietic stem cell fates

Bone marrow transplantation therapy relies on the life-long regenerative capacity of hematopoietic stem cells (HSCs). HSCs present a wide complexity of regenerating behaviours at the clonal level, but the mechanisms underlying this diversity are still unclear. Recent advances in single cell sequencing have described transcriptional differences amongst HSCs, providing a possible explanation for their functional heterogeneity. However, the destructive nature of sequencing assays prevents simultaneous observation of stem cell functions. Thus, it is unclear to what extent transcriptome states carry information about stem cell outcomes. Here, we applied expressible barcodes to simultaneously recover lineages and transcriptomes from single adult HSCs, and investigated their clonal trajectories during bone marrow reconstitution in vivo. Using clonal readouts, we unveiled a previously underappreciated subset of quiescent HSCs that cell-autonomously retains native-like behaviours. Transcriptome enrichment using clonal information identified a distinct molecular signature that characterizes functional long-term repopulating HSCs. Probing this signature through in vivo CRISPR screening, we described the transcription factor Tcf15 to be required, and sufficient, to drive HSC quiescence and long-term self-renewal. In situ, Tcf15 labels the subset of native multipotent HSCs and its expression is inherited in a hierarchical fashion. In conclusion, our work elucidates molecular programs associated with stem cell function, and identifies a mechanism for the maintenance of the native-like quiescent state. Single-cell mRNA sequencing of barcoded hematopoietic stem cells (HSC libraries) and progenitor cells (KIT libraries) using the inDrop (v3) pipeline. Sample 1: single cell mRNA sequencing libraries from 2 primary recipients (recipient 1 and 2). Sample 2: single cell mRNA sequencing libraries from 2 primary recipients (recipient 3 and 4). Sample 3:single cell mRNA sequencing libraries from 1 primary recipient (recipient 5). Sample 4: single cell mRNA sequencing libraries from secondary recipients. Sample 5: single cell mRNA sequencing libraries from CROPseq experiment replicate A and replicate B. Sample 6: single cell mRNA sequencing library from TetO-Tcf15 experiment (wt vs. TetO-Tcf15 KIT cells).