Fam72acontrols the balance between error-prone and error-free DNA repair during antibody diversification

Abstract

Efficient humoral responses rely on DNA damage, mutagenesis and error-prone DNA repair. B cell receptor diversification through somatic hypermutation (SHM) and class switch recombination (CSR) are initiated by cytidine deamination in DNA mediated by activation induced cytidine deaminase (AID)1 and by the subsequent excision of the resulting uracils by Uracil DNA glycosylase (UNG) and by mismatch repair (MMR) proteins2–4. Although uracils arising in DNA are faithfully repaired2–7, it is not known how these pathways are co-opted to generate mutations and double stranded DNA breaks (DSBs) in the context of SHM and CSR2,4,8. Here we have performed a genome-wide CRISPR/Cas9 knockout screen for genes involved in CSR. The screen identified FAM72A, a protein that interacts with the nuclear isoform of UNG (UNG2)9 and that is overexpressed in several cancers9. We show that the FAM72A-UNG2 interaction controls the protein levels of UNG2 and that CSR is defective in Fam72a−/− B cells due to the specific upregulation of UNG2. Moreover, we show that in Fam72a−/− B cells SHM is reduced by 5-fold and that upregulation of UNG2 results in a skewed mutation pattern. Our results are consistent with a model in which FAM72A interacts with UNG2 to control its physiological level by triggering its degradation. Consequently, deficiency in Fam72a leads to supraphysiological levels of UNG2 and enhanced uracil excision, shifting the balance from error-prone to error-free DNA repair. Our findings have potential implications for tumorigenesis, as Fam72a overexpression would lead to reduced UNG2 levels, shifting the balance toward mutagenic DNA repair and rendering cells more prone to acquire mutations.

Identification of a Chemotherapeutic Lead Molecule for the Potential Disruption of the FAM72A-UNG2 Interaction to Interfere with Genome Stability, Centromere Formation, and Genome Editing

“…Accumulating evidence indicates the involvement of FAM72A in tumorigenesis [ 24 , 25 , 26 , 29 , 87 , 88 ]. Elevated FAM72A causes reduced UNG2 levels, eventually leading to new mutations [ 24 , 25 , 27 , 28 , 29 ]. Our data pave the way for new investigative experimental approaches to validate the prevention of cancer by interfering with the FAM72A-UNG2 signaling pathways using withaferin B. Withaferin B is a potential candidate for future investigations in the interference with genome stability, centromere formation, and genome editing, and on potential therapeutic strategies for the treatment of cancer.…”

“…Recent data show that decreased levels of FAM72A lead to hyperphysiological UNG2 levels, an increased uracil correction, and, thus, error-free DNA repair. In contrast, the binding of FAM72A with UNG2 antagonizes UNG2 activity and causes UNG2 degradation in B cells, leading to increased levels of genome-wide deoxyuracils and, therefore, mediating increased levels of U•G mispairs that engage in mutagenic mismatch repair, promoting the error-prone processing of activation-induced cytidine deaminase (AID)-induced deoxyuracils [ 27 , 28 ]. Thus, FAM72A bridges BER and mismatch repair in order to modulate antibody diversification during B cell and antibody maturation [ 27 , 28 ].…”

“…In contrast, the binding of FAM72A with UNG2 antagonizes UNG2 activity and causes UNG2 degradation in B cells, leading to increased levels of genome-wide deoxyuracils and, therefore, mediating increased levels of U•G mispairs that engage in mutagenic mismatch repair, promoting the error-prone processing of activation-induced cytidine deaminase (AID)-induced deoxyuracils [ 27 , 28 ]. Thus, FAM72A bridges BER and mismatch repair in order to modulate antibody diversification during B cell and antibody maturation [ 27 , 28 ]. Overall, an increased FAM72A level could lead to reduced UNG2 levels and could thus shift the balance of appropriate mutagenic DNA repair, therefore making the cells more susceptible to mutations, with possible effects on tumor development [ 24 , 25 , 27 , 28 , 29 ].…”

“…Thus, FAM72A bridges BER and mismatch repair in order to modulate antibody diversification during B cell and antibody maturation [ 27 , 28 ]. Overall, an increased FAM72A level could lead to reduced UNG2 levels and could thus shift the balance of appropriate mutagenic DNA repair, therefore making the cells more susceptible to mutations, with possible effects on tumor development [ 24 , 25 , 27 , 28 , 29 ].…”