Impairment of fetal hematopoietic stem cell function in the absence of Fancd2

Abstract

Fanconi Anemia (FA), results from mutations in genes necessary for DNA damage repair and often leads to progressive bone marrow failure. Although the exhaustion of the bone marrow leads to cytopenias in FA patients as they age, evidence from human FA and mouse model fetal livers suggests hematopoietic defects originate in utero which may lead to deficient seeding of the bone marrow. To address this possibility, we examined the consequences of loss of Fancd2, a central component of the FA pathway. Examination of E14.5 Fancd2 knockout (KO) fetal livers showed a decrease in total cellularity and specific declines in long-term and short-term hematopoietic stem cell (LT- and ST-HSC) numbers. Fancd2 KO fetal liver cells display similar functional defects to Fancd2 adult bone marrow cells including reduced colony forming units, increased mitomycin C sensitivity, increased LT-HSC apoptosis and heavily impaired competitive repopulation, implying these defects are intrinsic to the fetal liver and not dependent on the accumulation of DNA damage during aging. Telomere shortening, an aging-related mechanism proposed to contribute to HSC apoptosis and bone marrow failure in FA, was not observed in Fancd2 KO fetal livers. In summary, loss of Fancd2 yields significant defects to fetal liver hematopoiesis, particularly the HSC population, which mimic key phenotypes from adult Fancd2 KO bone marrow independent of aging-accrued DNA damage.

FANCI and FANCD2 have common as well as independent functions during the cellular replication stress response

“…Moreover, since I −/− and ID2 DKO cells are equally resistant to HU or APH despite harboring different residual chromatin-bound FANCD2 levels, FANCD2’s role in preventing APH/HU-induced cell death may in fact not depend on its own chromatin association. Consistent with our findings and interpretation, a role for FANCD2 in preventing apoptosis and cell death was also suggested by several groups ( 70 – 72 ). Moreover, a recent study showed that siRNA-mediated knockdown of FANCI, but not FANCD2, substantially reduced cellular apoptosis after UV treatment ( 67 ).…”

Studies in an Early Development Window Unveils a Severe HSC Defect in both Murine and Human Fanconi Anemia

“…Fancg −/− embryos were obtained at a lower frequency than expected at E11.5 and E12.5 (20% and 21.7%, respectively, instead of 25%, 958 embryos tested in total; p < 0.001). As observed in Fancc −/− and Fancd2 −/− mice ( Kamimae-Lanning et al., 2013 , Suzuki et al., 2017 ), this frequency dramatically declined at birth (11% ± 0.8, 82 postnatal day 0 [P0] to P1 pups tested, p < 0.001), suggesting a late perinatal lethality. Interestingly, at E11.5 and E12.5 Fancg −/− embryos were systematically smaller than wild-type (WT) control littermates, while at E14.5 only half of the Fancg −/− embryos remained smaller than control ( Figure 1 A).…”

“…However, a deficit in the CD34 + cell numbers in FA subjects is observed very early during childhood, even before the onset of the BMF clinical symptoms ( Ceccaldi et al., 2012 , Kelly et al., 2007 ). This, added to the early developmental defects of the hematopoietic commitment described for human embryonic stem cells after knockdown of FANCA and FANCD2 genes, led us and others to hypothesize that the FA hematopoietic defect could already begin in utero ( Ceccaldi et al., 2012 , Kamimae-Lanning et al., 2013 , Suzuki et al., 2017 , Tulpule et al., 2010 , Yoon et al., 2016 ).…”

Recent advances in understanding hematopoiesis in Fanconi Anemia

“…That functionally corrective somatic mutations in a single human HSC prenatally are capable of completely replacing the hematopoietic tissues in twins by the time of birth 62 is conclusive proof that the repopulating capacity and overall fitness of the mutant HSCs are unambiguously reduced during development and can be overtaken by even one HSC in which the mutation has been corrected. This conclusion has been confirmed in human embryonic stem cells 63 and Fancc –/– and Fancd2 –/– mice 64 , 65 . The experiment of nature 62 provides clear theoretical evidence that if a single corrected stem cell can so massively outperform all of the mutant ones during development, gene therapy for this disease early in life (or perhaps in utero 66 ) can be successful in the future.…”