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.
DEK is an oncoprotein that is overexpressed in many forms of cancer and participates in numerous cellular pathways. Of these different pathways, relevant interacting partners and functions of DEK are well described in regard to the regulation of chromatin structure, epigenetic marks, and transcription. Most of this understanding was derived by investigating DNA-binding and chromatin processing capabilities of the oncoprotein. To facilitate the generation of mechanism-driven hypotheses regarding DEK activities in underexplored areas, we have developed the first DEK interactome model using tandem-affinity purification and mass spectrometry. With this approach, we identify IMPDH2, DDX21, and RPL7a as novel DEK binding partners, hinting at new roles for the oncogene in de novo nucleotide biosynthesis and ribosome formation. Additionally, a hydroxyurea-specific interaction with replication protein A (RPA) was observed, suggesting that a DEK-RPA complex may form in response to DNA replication fork stalling. Taken together, these findings highlight diverse activities for DEK across cellular pathways and support a model wherein this molecule performs a plethora of functions.DDX21, DEK, IMPDH2, interactome, mass spectrometry, ribosome, RPA | I N TR ODU C TI ONDEK is a DNA-binding and predominantly nuclear protein that was first identified as a DEK-NUP214 fusion protein in AML. 1 Since this discovery, DEK has been classified as an oncoprotein and shown to be overexpressed in many diverse tumor types, 2 wherein the degree of overexpression was linked to worse prognosis, advanced stage tumors, and chemotherapy resistance. 3-9 Cellular functions of the oncoprotein include activities in modifying chromatin structure, 2,10-13 histone chaperoning, 14,15 epigenetic modification and transcription regulation, [16][17][18][19]20,21 DNA repair,22,23 DNA replication fork restart, 24 mitotic non-disjunction events, 25 evasion of senescence and apoptosis, 19,26,27 proliferation 28-30 cancer stem cell fitness and invasion, 28,31 inflammation, 32,33 and metabolic reprogramming. 34 Precise molecular mechanisms whereby DEK regulates these cellular processes remain unclear in many cases. For instance, while DEK is necessary for optimal non-homologous end joining (NHEJ) DNA repair, the mechanism of action or interacting partners remain elusive. 23 For an oncogene that is widely implicated in human carcinogenesis and outcome, determining how DEK operates is imperative. However, this has proven challenging as the oncoprotein DEK has no known enzymatic activity or paralogs. 2 While significant progress has been made to understand molecular DEK activities in transcription via chromatin remodeling, 2,[10][11][12][13][16][17][18][19] there is a dearth of information regarding 88 | V C 2017 Wiley Periodicals, Inc.wileyonlinelibrary.com/journal/prot Proteins. 2018;86:88-97.
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