Separate Roles of FAN1 and Fanconi Anemia Proteins in DNA Interstrand Crosslink Repair and Human Disease

DNA interstrand crosslink (ICL) repair is vital for cellular proliferation and survival. Defects in the repair of DNA crosslinks have been associated with congenital abnormalities, bone marrow and kidney failure, liver dysfunction, and cancer. DNA nucleases play a significant role during the repair of ICLs and they function at multiple repair steps. Here, we assessed the contributions of FANCD2/FANCI-Associated Nuclease 1 (FAN1) to the repair of ICL lesions and studied the consequences of its deficiency, which results in rare chronic kidney disease - Karyomegalic Interstitial Nephritis (KIN). FAN1 is a highly conserved nuclease from yeast to humans. It was first identified in an RNAi screen for proteins necessary for ICL repair and also as a direct interacting partner of FANCI/FANCD2 (ID2) complex of the Fanconi anemia pathway. Using cells isolated from human KIN patients and Fan1-deficient mice, we showed that ICL repair function of FAN1 is non-overlapping with Fanconi anemia proteins, yet redundant with SNM1A, an exonuclease that was previously implicated in ICL repair. Furthermore, the Fan1 knockout mouse model recapitulates many of the phenotypes observed in human KIN patients, including renal karyomegaly and liver dysfunction. Fan1-deficient mice displayed hypersensitivity to ICL-inducing chemotherapeutic drugs and developed bone marrow failure following the treatment. We also demonstrated that formaldehyde and acetaldehyde are likely not the endogenous damaging agents that lead to KIN in the absence of FAN1. In parallel studies, we explored the possibility of rescuing ICL sensitivity of Fanconi anemia patient cells through an inhibition of non-homologous end joining repair (NHEJ) factors. Using both genetic and chemical approaches, we showed that NHEJ suppression could not compensate for the lack of FA pathway in the repair of ICLs