The 3'-flap endonuclease XPF-ERCC1 promotes alternative end joining and chromosomal translocation during B cell class switching.

Wanyu Bai,Pingyue Chen,Chun Chen,Jiejie Xu,Guangchao Zhu,Feilong Meng,Hongman Xue,Junchao Dong

doi:10.1016/j.celrep.2021.109756

Abstract

Robust alternative end joining (A-EJ) in classical non-homologous end joining (c-NHEJ)-deficient murine cells features double-strand break (DSB) end resection and microhomology (MH) usage and promotes chromosomal translocation. The activities responsible for removing 3' single-strand overhangs following resection and MH annealing in A-EJ remain unclear. We show that, during class switch recombination (CSR) in mature mouse B cells, the structure-specific endonuclease complex XPF-ERCC1SLX4, although not required for normal CSR, represents a nucleotide-excision-repair-independent 3' flap removal activity for A-EJ-mediated CSR. B cells deficient in DNA ligase 4 and XPF-ERCC1 exhibit further impaired class switching, reducing joining to the resected S region DSBs without altering the MH pattern in S-S junctions. In ERCC1-deficient A-EJ cells, 3' single-stranded DNA (ssDNA) flaps that are generated predominantly in S/G2 phase of the cell cycle are susceptible to nuclease resolution. Moreover, ERCC1 promotes c-myc-IgH translocation in Lig4-/- cells. Our study reveals an important role of the flap endonuclease XPF-ERCC1 in A-EJ and oncogenic translocation in mouse B cells.

Highlights

DNA double-strand breaks (DSBs) arise from constant assault by environmental factors and cellular metabolites and from programmed processes during antibody receptor gene diversification in developing lymphocytes (Alt et al, 2013)
Classical non-homologous end joining (NHEJ) (c-NHEJ) in mammalian cells is catalyzed by Ku70/80 and XRCC4/DNA ligase 4 (Lig4) complexes, and DNA-PKcs and Artemis in certain circumstances (Boboila et al, 2012; Chang et al, 2017). c-NHEJ dominates repair of programmed DSBs that occur in V(D)J recombination and class switch recombination (CSR) in developing T and B lymphocytes, and much insight into the molecular mechanisms of c-NHEJ has been gained by studying these processes
ERCC1 is required for alternative end joining (A-EJ)- but not c-NHEJ-mediated CSR in mouse B cells To elucidate the functions of XPF-ERCC1 in mouse CSR, we first generated an Ercc1 deletion mutation in the CH12F3 cell line that can be stimulated to switch to IgA in culture sgRNA chr7:19,345,071-19,356,524 ll WT #1 #2 Lig4-/- #1 #2 53bp1-/- #1 #2

Summary

SUMMARY

Robust alternative end joining (A-EJ) in classical non-homologous end joining (c-NHEJ)-deficient murine cells features double-strand break (DSB) end resection and microhomology (MH) usage and promotes chromosomal translocation. We show that, during class switch recombination (CSR) in mature mouse B cells, the structure-specific endonuclease complex XPF-ERCC1SLX4, not required for normal CSR, represents a nucleotide-excision-repair-independent 30 flap removal activity for A-EJ-mediated CSR. B cells deficient in DNA ligase 4 and XPF-ERCC1 exhibit further impaired class switching, reducing joining to the resected S region DSBs without altering the MH pattern in S-S junctions. In ERCC1-deficient A-EJ cells, 30 single-stranded DNA (ssDNA) flaps that are generated predominantly in S/G2 phase of the cell cycle are susceptible to nuclease resolution. Our study reveals an important role of the flap endonuclease XPF-ERCC1 in A-EJ and oncogenic translocation in mouse B cells

INTRODUCTION

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DISCUSSION

METHOD DETAILS