The non-homologous end-joining activity is required for Fanconi anemia fetal HSC maintenance

Recent studies have shown that deficiency in the Fanconi anemia (FA) DNA repair pathway enhances the error-prone non-homologous end-joining (NHEJ) repair, leading to increased genomic instability, and that genetic or pharmacological inhibition of the NHEJ pathway could rescue the FA phenotype.

First, we exposed LSK cells from WT and Fanca^-/- mice to DNA-PKcs inhibitor NU7026 or Ku70 knockdown to examine whether inhibition of NHEJ sensitizes Fanca^-/- HSPCs to PARP inhibitor (PARPi)- or interstrand crosslinking (ICL)-induced cell death and genomic instability. We then generated DNA-PKcs^3A/3AFanca^-/- mice to investigate the effect of specific inactivation of NHEJ on fetal HSCs. Lastly, we used two p53 mutant models to test whether specific inactivation of the p53 function in apoptosis is sufficient to rescue embryonic lethality and fetal HSC depletion in Fanca^-/- DNA-PKcs^3A/3A mice.

Inhibition of NHEJ sensitizes HSPCs from Fanca^-/- mice to PARP inhibition- and ICL-induced cell death and genomic instability and further decreases Fanca^-/- HSPC proliferation and hematopoietic repopulation in irradiated transplant recipients. Specific inactivation of NHEJ activity by the knockin DNA-PKcs^3A/3A mutation in two FA mouse models, Fanca^-/- and Fancc^-/-, leads to embryonic lethality. DNA-PKcs^3A/3A causes fetal HSC depletion in developing Fanca^-/- embryos due to increased HSC apoptosis and cycling. Both p53^-/- and a knockin p53^515C mutation, which selectively impairs the p53 function in apoptosis, can rescue embryonic lethality and fetal HSC depletion in Fanca^-/- DNA-PKcs^3A/3A mice.

These results demonstrate that the NHEJ pathway functions to maintain Fanconi anemia fetal HSCs.