The physiologic role of fanconi anemia proteins in guarding against replication-associated stress during developmental HSPC expansion in the fetal liver

Fanconi anemia (FA) is a recessively inherited multisystem disorder, commonly presenting in early school age with cytopenias that result from bone marrow failure. The underlying loss of stem cells is thought to be related to defects in the coordinate activity of FA proteins in a DNA repair pathway. Additional evidence points to a potential role for innate immunity and cellular aldehyde metabolism. The challenge in dissecting the molecular events that lead to bone marrow failure is in part related to the paucity of model systems that resemble the spontaneous failure encountered in patients. We previously reported prenatal deficits in the hematopoietic stem and progenitor cell (HSPC) pool of mice with transgenic disruption of Fancc. We now provide evidence that pervasive developmental defects along with late gestational lethality occur after disruption of Fancd2, a key component of the pathway. The data demonstrate spontaneous depletion of lineage-depleted, Sca-1+, CD48-, CD150+ (SLAM) cells in the fetal liver and coincident loss of progenitor clonogenicity. Experiments indicate the rapid upregulation of genes involved in DNA repair in HSPC and the successive accrual of RAD51 foci. The accumulating DNA damage provokes a cellular stress response that upregulates Cdkn1a (P21), consistent with canonical p53 activation, and loss of miRNA 125b expression, not previously described. Distinct from models of induced HSPC loss and consistent with FA modeling in hESC, we find no evidence of apoptosis or senescence. Crucially, serial repopulation studies reveal exhaustion of the fetal HSPC pool in Fancd2-deficient offspring, but not WT littermates. Finally, our data demonstrate the rescue of progenitor formation via pharmacological inhibition of p38 MAPK. In aggregate, our studies reveal a replication-associated loss of stem cells from the developing FA HSPC pool with evidence for spontaneous DNA damage response activation. Unlike models relying on exogenous induction, we report the molecular events that lead to physiological, developmentally programmed hematopoietic deficits in FA and their origin in utero.