Abstract

Transitions between subsets of differentiating hematopoietic cells are widely regarded as unidirectional . Here, we introduce clonal phylogenetic tracer (CP-tracer) that sequentially introduces genetic barcodes, enabling high-resolution analysis of ~100,000 subclones derived from ~500 individual hematopoietic stem cells (HSC). This revealed previously uncharacterized HSC functional subsets and identified bidirectional fate transitions between myeloid-biased and lineage-balanced HSC. Contrary to the prevailing view that the more self-renewing My-HSCs unidirectionally transition to balanced-HSCs, phylogenetic tracing revealed durable lineage bidirectionality with the transition favoring My-HSC accumulation over time. Further, balanced-HSCs mature through distinct intermediates-My-HSCs and lymphoid-biased-HSCs-with lymphoid competence here shown by CRISPR/Cas9 screening to be dependent on the homeobox gene, . Hhex enables Ly-HSC differentiation, but its expression declines with age. These findings establish HSC plasticity and as a determinant of myeloid-lymphoid balance with each changing over time to favor the age-related myeloid bias of the elderly.