APE1 is dispensable for S-region cleavage but required for its repair in class switch recombination

Abstract

Activation-induced cytidine deaminase (AID) is essential for antibody diversification, namely somatic hypermutation (SHM) and class switch recombination (CSR). The deficiency of apurinic/ apyrimidinic endonuclease 1 (Ape1) in CH12F3-2A B cells reduces CSR to ∼20% of wild-type cells, whereas the effect of APE1 loss on SHM has not been examined. Here we show that, although APE1's endonuclease activity is important for CSR, it is dispensable for SHM as well as IgH/c-myc translocation. Importantly, APE1 deficiency did not show any defect in AID-induced S-region break formation, but blocked both the recruitment of repair protein Ku80 to the S region and the synapse formation between Sμ and Sα. Knockdown of end-processing factors such as meiotic recombination 11 homolog (MRE11) and carboxy-terminal binding protein (CtBP)-interacting protein (CtIP) further reduced the remaining CSR in Ape1-null CH12F3-2A cells. Together, our results show that APE1 is dispensable for SHM and AID-induced DNA breaks and may function as a DNA end-processing enzyme to facilitate the joining of broken ends during CSR.class switch recombination | somatic hypermutation | DNA cleavage | DNA synapse formation | end processing

Regulation of the DNA Repair Complex during Somatic Hypermutation and Class-Switch Recombination

“…The number of proteins participating in this repair is large. Although the detailed mechanisms are still controversial (7), it is well established that base excision repair (BER) enzymes, including the uracil DNA glucosylase (UNG), apurinic/ apyrimidinic endonuclease 1 (APE1), x-ray repair cross-complementing 1 (XRCC1; a scaffold protein involved in BER), mismatch repair (MMR) enzymes, including MSH2, MSH6, and MLH1, and errorprone translesion DNA polymerases, especially polymerase h, play important roles in SH and CSR (1,(7)(8)(9). However, knowledge of interactions among these proteins during SH/CSR has been almost completely lacking.…”

Antibody repertoire and gene expression dynamics of diverse human B cell states during affinity maturation

“…One surprising result from our reconstruction of human B cell activation and GC entry was the discovery that several genes associated with class switch recombination were most highly expressed prior to GC entry, with a specific enrichment of these genes in the preGC B cell population (Figure 3H-I). This included APEX1 expression, of which its translated product APE1 is required for class switch recombination to occur in a dose-dependent manner (Masani et al, 2013, Xu et al, 2014a) and is expressed almost exclusively by non-GC B cells (Figure S2B; Roco et al, 2019). We also found that preGC B cells had the highest expression of IgH germline transcripts (GLTs) (Figure 3J), which preceded switching from IgM/IgD to other isotypes (Figure 3K), in fitting with a recent study describing GLT transcription prior to GC formation in mouse models (Roco et al, 2019).…”

Chromatin remodeller SMARCA4 recruits topoisomerase 1 and suppresses transcription-associated genomic instability

“…Genomic DNAs were prepared from CH12F3-2A cells stimulated with CIT for 24 h. DSBs were analysed by ligation-mediated PCR as previously described 65 66 . Briefly, living CH12F3-2A cells were isolated by Percoll density gradient centrifugation and embedded in low-melting-point agarose followed by DNA extraction within low-melting agarose plugs.…”