NR4A2 Promotes DNA Double-strand Break Repair Upon Exposure to UVR

Abstract

Exposure of melanocytes to ultraviolet radiation (UVR) induces the formation of UV lesions that can produce deleterious effects in genomic DNA. Encounters of replication forks with unrepaired UV lesions can lead to several complex phenomena, such as the formation of DNA double-strand breaks (DSBs). The NR4A family of nuclear receptors are transcription factors that have been associated with mediating DNA repair functions downstream of the MC1R signaling pathway in melanocytes. In particular, emerging evidence shows that upon DNA damage, the NR4A2 receptor can translocate to sites of UV lesion by mechanisms requiring post-translational modifications within the N-terminal domain and at a serine residue in the DNA-binding domain at position 337. Following this, NR4A2 aids in DNA repair by facilitating chromatin relaxation, allowing accessibility for DNA repair machinery. Using A2058 and HT144 melanoma cells engineered to stably express wild-type or mutant forms of the NR4A2 proteins, we reveal that the expression of functional NR4A2 is associated with elevated cytoprotection against UVR. Conversely, knockdown of NR4A2 expression by siRNA results in a significant loss of cell viability after UV insult. By analyzing the kinetics of the ensuing 53BP1 and RAD51 foci following UV irradiation, we also reveal that the expression of mutant NR4A2 isoforms, lacking the ability to translocate, transactivate, or undergo phosphorylation, display compromised repair capacity. These data expand the understanding of the mechanism by which the NR4A2 nuclear receptor can facilitate DNA DSB repair. .

Identification of activity-inducedEgr3-dependent genes reveals genes associated with DNA damage response and schizophrenia

“…We found genes involved in regulation of behavior and DNA damage response pathways to be significantly overrepresented in our DEG list. Our results suggest that Egr3 is necessary for ECS-dependent stimulation of a subset of genes involved in regulation of nervous system function including regulation of memory ( Npas4 51 , Nr4a2 52 ), neurogenesis and synaptic plasticity ( Gadd45b 40 , Bdnf 64 ), behavior ( Fos 65 , Fosb 66 ) and DNA damage response, particularly, DNA repair ( Cenpa 43 , GADD45 family proteins Gadd45b and Gadd45g 37, 38 , Fos and Fosb that are part of the AP-1 transcription factor 48 , and Nr4a2 54 ). We also report that two genes that show elevated expression in schizophrenia ( Calb2 58 , Mef2c 57 ) are upregulated in mice lacking Egr3 .…”

NR4A Nuclear Receptors Target Poly-ADP-Ribosylated DNA-PKcs Protein to Promote DNA Repair

“…We find that DBD of NR4A is bi-functional and can bind DNA to regulate transcription (in a sequence-specific fashion) or PAR to facilitate DSB repair. Given that NR4A2 overexpression can accelerate DSB repair ( Malewicz et al., 2011 , Yin et al., 2017 ), it appears that the DNA-PK assembly is a rate-limiting step in this process. Thus, physiological and pathological conditions in which the expression of NR4A is altered, such as stress responses and various cancers, might be associated with variable NHEJ activity, with important consequences for cell physiology and drug responses.…”

“…Typically, the Zn1 contacts DNA ( Meinke and Sigler, 1999 ), while Zn2 is not involved in direct DNA binding. A direct role for NR4As has been discovered in DNA DSB repair, but the mechanism remains elusive ( Jagirdar et al., 2013 , Malewicz et al., 2011 , Ramirez-Herrick et al., 2011 , Smith et al., 2008 , Yin et al., 2017 ). Here we show that NR4As’ DBD is functionally unique, because it is able to bind both DNA and PAR.…”

Genetic variation in IRF4 expression modulates growth characteristics, tyrosinase expression and interferon‐gamma response in melanocytic cells

“…IRF4, BRN2 and empty vector lentivirus were generated using the Lenti‐X HT packaging system (Clontech) according to the manufacturer's protocol. A2058 and HT144 melanoma cells were transduced with lentivirus as previously described (Yin et al., ), and cells were selected in media containing 0.75 μg/ml puromycin and expression confirmed by Western blotting. MB cell strains and established melanoma lines were transfected with 20 nmol/l of non‐targeting siRNA (scramble/control, Dharmacon) and siRNAs targeting IRF4 (GE Dharmacon), BRN2 (Shanghai GenePharma) and MITF (Shanghai GenePharma) using Lipofectamine 2000 (Invitrogen) according to manufacturer's guidelines.…”

“…IRF4, BRN2 and empty vector lentivirus were generated using the Lenti-X HT packaging system (Clontech) according to the manufacturer's protocol. A2058 and HT144 melanoma cells were transduced with lentivirus as previously described (Yin et al, 2017), and cells were selected in media containing 0.75 μg/ml…”