DEK is required for homologous recombination repair of DNA breaks

Eric A Smith,Lisa Wiesmüller,Amom R Meetei,Ralph Scully,Linda M Starnes,Boris Gole,Abdullah M Ali,Haihong Guo,Jeremy A Daniel,Rebeca Soria,Anil G Jegga,Susanne I Wells,Lisa M Privette Vinnedge,Nicholas A Willis,Eric F Krumpelbeck,Paul R Andreassen,Ferdinand Kappes

doi:10.1038/srep44662

Abstract

DEK is a highly conserved chromatin-bound protein whose upregulation across cancer types correlates with genotoxic therapy resistance. Loss of DEK induces genome instability and sensitizes cells to DNA double strand breaks (DSBs), suggesting defects in DNA repair. While these DEK-deficiency phenotypes were thought to arise from a moderate attenuation of non-homologous end joining (NHEJ) repair, the role of DEK in DNA repair remains incompletely understood. We present new evidence demonstrating the observed decrease in NHEJ is insufficient to impact immunoglobulin class switching in DEK knockout mice. Furthermore, DEK knockout cells were sensitive to apoptosis with NHEJ inhibition. Thus, we hypothesized DEK plays additional roles in homologous recombination (HR). Using episomal and integrated reporters, we demonstrate that HR repair of conventional DSBs is severely compromised in DEK-deficient cells. To define responsible mechanisms, we tested the role of DEK in the HR repair cascade. DEK-deficient cells were impaired for γH2AX phosphorylation and attenuated for RAD51 filament formation. Additionally, DEK formed a complex with RAD51, but not BRCA1, suggesting a potential role regarding RAD51 filament formation, stability, or function. These findings define DEK as an important and multifunctional mediator of HR, and establish a synthetic lethal relationship between DEK loss and NHEJ inhibition.

Highlights

The DNA-binding and chromatin-regulating DEK oncogene is expressed across multicellular eukaryotes and is highly conserved in mammals
To determine if DEK-deficient cells require non-homologous end joining (NHEJ) for survival, we examined the need for NHEJ by inhibiting the upstream kinase, DNA-PK
We demonstrate that DEK is required for the repair of double strand breaks (DSBs) by homologous recombination (HR), and regulates multiple steps in the HR cascade by promoting γH2AX activation, enabling robust RAD51 loading, and forming a complex with RAD51 (Fig. 5d)

Summary

Introduction

The DNA-binding and chromatin-regulating DEK oncogene is expressed across multicellular eukaryotes and is highly conserved in mammals. The observed NHEJ defects in DEK-deficient cells are unlikely to fully account for the severe sensitivity to genotoxic agents, especially DNA interstrand cross linkers and topoisomerase inhibitors[14,32] This suggests additional roles for DEK in genotoxic drug tolerance and DNA repair. The γH2AX mark supports DSB repair by enhancing the recruitment of BRCA1 and key nucleases including the MRN complex and CtIP38,42–44 These factors coordinate DNA end processing into single strand DNA (ssDNA) 3′tails[38,42,43]. The resulting ssDNA is initially coated by RPA, which is efficiently replaced with a RAD51 filament through the combined activities of BRCA1, BRCA2, the RAD51 paralogs, and other factors[38,42] This RAD51 filament catalyzes strand invasion, complementary strand annealing, and the formation of a stable synaptic complex with a homologous sequence on the sister chromatid[45]

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Results

Conclusion