WRN regulates pathway choice between classical and alternative non-homologous end joining.

Raghavendra A Shamanna,Vilhelm A Bohr,Jane Tian,Jessica K De Freitas,Deborah L Croteau,Huiming Lu

doi:10.1038/ncomms13785

Abstract

Werner syndrome (WS) is an accelerated ageing disorder with genomic instability caused by WRN protein deficiency. Many features seen in WS can be explained by the diverse functions of WRN in DNA metabolism. However, the origin of the large genomic deletions and telomere fusions are not yet understood. Here, we report that WRN regulates the pathway choice between classical (c)- and alternative (alt)-nonhomologous end joining (NHEJ) during DNA double-strand break (DSB) repair. It promotes c-NHEJ via helicase and exonuclease activities and inhibits alt-NHEJ using non-enzymatic functions. When WRN is recruited to the DSBs it suppresses the recruitment of MRE11 and CtIP, and protects the DSBs from 5′ end resection. Moreover, knockdown of Wrn, alone or in combination with Trf2 in mouse embryonic fibroblasts results in increased telomere fusions, which were ablated by Ctip knockdown. We show that WRN regulates alt-NHEJ and shields DSBs from MRE11/CtIP-mediated resection to prevent large deletions and telomere fusions.

Highlights

Werner syndrome (WS) is an accelerated ageing disorder with genomic instability caused by WRN protein deficiency
In vitro nonhomologous end joining (NHEJ) was performed with DNA substrates containing cohesive and non-cohesive ends and cellular extracts prepared from HeLa or U2OS cells treated with control and WRN short hairpin RNA or with small interfering RNA (Supplementary Fig. 1A)
In U2OS cells, WRN knockdown inhibited total NHEJ by B61% on non-cohesive substrates (Fig. 1b). These results suggest that WRN promotes NHEJ in vitro

Summary

Introduction

Werner syndrome (WS) is an accelerated ageing disorder with genomic instability caused by WRN protein deficiency. We report that WRN regulates the pathway choice between classical (c)- and alternative (alt)-nonhomologous end joining (NHEJ) during DNA double-strand break (DSB) repair. It promotes c-NHEJ via helicase and exonuclease activities and inhibits alt-NHEJ using non-enzymatic functions. We show that WRN regulates alt-NHEJ and shields DSBs from MRE11/CtIP-mediated resection to prevent large deletions and telomere fusions. WS cells and Wrn knockout mouse cells show genome instability, often with large deletions and telomere fusions[3,5,6,7,8]. The resected ends are ligated by DNA ligase I or ligase III (refs 19,20–23)

Methods

Results

Conclusion