A study has uncovered a unique pattern of DNMT3A mutations in frequent blood donors that respond specifically to erythropoietin, revealing a previously unknown mechanism of clonal selection in hematopoietic stem cells.
In the study, published in Blood, researchers from an international collaboration led by scientists at the German Cancer Research Center, The Francis Crick Institute, and German Red Cross Blood Donation Centre examined clonal hematopoiesis (CH) in 217 older male volunteer donors with extensive donation histories (> 100 lifetime donations) compared with 212 age-matched controls with fewer than 10 lifetime donations.
While the overall incidence of CH was similar between frequent donors and controls, the researchers identified a distinct mutational pattern in the DNMT3A gene among frequent donors. These mutations exhibited different environmental response patterns compared with the well-known preleukemic R882 mutations.
"Our data demonstrate a nuanced ongoing Darwinian evolution at the somatic stem cell level, with erythropoietin (EPO) identified as a novel environmental factor that favors [hematopoietic stem cells] (HSC) carrying certain DNMT3A mutations," the study authors stated.
The researchers found that DNMT3A variants from frequent donors had significantly lower stability scores compared with variants in the control cohort. These mutations typically caused quantitatively reduced but qualitatively normal enzyme activity, unlike the aberrant functionality seen with R882 mutations.
When the researchers introduced these mutations into human HSCs using CRISPR-Cas9 editing, they observed that the frequent donor DNMT3A mutations expanded in EPO-rich environments but not under inflammatory stimuli. Conversely, cells carrying the preleukemic R882H mutation responded to interferon-gamma but showed no expansion with EPO.
Single-cell analysis revealed that HSCs with frequent donor DNMT3A variants maintained balanced lineage output during homeostasis but exhibited preferential erythroid differentiation under erythropoietic stress. This contrasted sharply with R882-mutated HSCs, which demonstrated a pronounced myeloid bias regardless of environmental conditions.
The molecular mechanism appeared to involve altered DNMT3A transcript abundance, with the W305* variant showing reduced levels of protein-coding transcripts and increased nonsense-mediated decay transcripts. Downregulation of DNMT3A in cell models resulted in upregulation of hemoglobin genes and the erythropoietin receptor.
This research established a reference CH data set from healthy individuals subject to a specific type of environmental stress and suggested that EPO-responsive DNMT3A mutations with moderate fitness advantages can be selected over time in individuals undergoing repeated blood donation.
These findings had significant implications for understanding the dynamics of clonal selection in HSCs and may inform future research into donor safety and the long-term effects of frequent blood donation.
The authors declared having no competing interests.
