Clonal analysis of lineage fate in native hematopoiesis

Hematopoiesis, the process of mature blood and immune cell production, is functionally organized as a hierarchy, with self-renewing hematopoietic stem cells (HSCs) and multipotent progenitor (MPP) cells sitting at the very top(1,2). Multiple models have been proposed as to what the earliest lineage choices are in these primitive hematopoietic compartments, the cellular intermediates, and the resulting lineage trees that emerge from them(3–10). Given that the bulk of studies addressing lineage outcomes have been performed in the context of hematopoietic transplantation, current lineage branching models are more likely to represent roadmaps of lineage potential rather than native fate. Here, we utilize transposon (Tn) tagging to clonally trace the fates of progenitors and stem cells in unperturbed hematopoiesis. Our results describe a distinct clonal roadmap in which the megakaryocyte (Mk) lineage arises largely independently of other hematopoietic fates. Our data, combined with single cell RNAseq, identify a functional hierarchy of uni- and oligolineage producing clones within the MPP population. Finally, our results demonstrate that traditionally defined long-term HSCs (LT-HSCs) are a significant source of Mk-restricted progenitors, suggesting that the Mk-lineage is the predominant native fate of LT-HSCs. Our study provides evidence for a substantially revised roadmap for unperturbed hematopoiesis, and highlights unique properties of MPPs and HSCs in situ.