Multifaceted regulation and functions of 53BP1 in NHEJ‑mediated DSB repair (Review)

Abstract

The repair of DNA double‑strand breaks (DSBs) is crucial for the preservation of genomic integrity and the maintenance of cellular homeostasis. Non‑homologous DNA end joining (NHEJ) is the predominant repair mechanism for any type of DNA DSB during the majority of the cell cycle. NHEJ defects regulate tumor sensitivity to ionizing radiation and anti‑neoplastic agents, resulting in immunodeficiencies and developmental abnormalities in malignant cells. p53‑binding protein 1 (53BP1) is a key mediator involved in DSB repair, which functions to maintain a balance in the repair pathway choices and in preserving genomic stability. 53BP1 promotes DSB repair via NHEJ and antagonizes DNA end overhang resection. At present, novel lines of evidence have revealed the molecular mechanisms underlying the recruitment of 53BP1 and DNA break‑responsive effectors to DSB sites, and the promotion of NHEJ‑mediated DSB repair via 53BP1, while preventing homologous recombination. In the present review article, recent advances made in the elucidation of the structural and functional characteristics of 53BP1, the mechanisms of 53BP1 recruitment and interaction with the reshaping of the chromatin architecture around DSB sites, the post‑transcriptional modifications of 53BP1, and the up‑ and downstream pathways of 53BP1 are discussed. The present review article also focuses on the application perspectives, current challenges and future directions of 53BP1 research.

Keywords: DNA double strand break; Pax transactivation domain‑interacting protein; RAP1‑interacting factor 1; non‑homologous end joining; p53‑binding protein 1.

Figure 1

Domain structures and functions of 53BP1. Recruitment of 53BP1 to DSB sites requires the minimal focus forming region, comprising the OD, the GAR motif, the tandem Tudor domain, the UDR motif, and the LC8 binding domain. The N-terminal S-T/Q phosphorylation sites mediate interactions with PTIP and RIF1/Shieldin/CST/Polα/Primase axis, which control DNA end resection. The C-terminal includes two BRCT domains connected in series, which are 53BP1 interacting with other proteins in a phosphorylation-independent pathway, such as p53 and MUM1 (or EXPAND1). 53BP1; p53-binding protein 1; DSB, double-strand break; OD, oligomerization domain; GAR, glycine-arginine-rich; UDR, ubiquitin-dependent recognition; LC8, dynein light chain; PTIP, Pax transactivation domain-interacting protein; BRCT, breast cancer type 1 susceptibility protein carboxyl-terminal; RIF1, RAP1-interacting factor 1; MUM1, mutated melanoma-associated antigen 1; CST, CTC1-STN1-TEN1; Polα, polymerase-α; DYNLL1, dynein light chain 1; PRMT1, protein arginine N-methyltransferase 1; H2AX, H2A histone family member X.

Figure 2

Graphic representation of 53BP1 recruitment and its nano-foci formation around DSB sites (DSBs). (A) DSB formation triggers a range of protein modifications that orchestrate the cellular response and DNA repair. DNA-PKcs, Ku70/Ku80, XRCC4 and etc. bind to DSBs, followed ligate end by one after another recruiting or activating MRN complex, ATM, γH2AX and MDC1. This provides a positive feedback loop for DSB signal amplification. MDC1 recruits RNF8, which cooperates with RNF168 to catalyze histone H2A ubiquitylation at DSBs. H2AK15ub, together with H4K20me2, mediates 53BP1 recruitment at DSBs. In its ATM-phosphorylated form, 53BP1 interacts with RIF1 and PTIP, which promote NHEJ repair. (B) 3D reorganization of 53BP1 foci and chromatin architecture. 53BP1 binds to histone modifications on damaged chromatin at the vicinity of the DSB and recruits RIF1, which elicits the assembly of the Shieldin complex. Shieldin complex protects broken DNA ends from nucleolytic degradation by resection factors. The spreading of 53BP1 on chromatin occurs over megabases around the DSB and is shaped by chromatin topology with the formation of distinct 53BP1 nanodomains (close to 100 nm) corresponding to chromatin TADs. RIF1 and Cohesin complex lead to the 'loop extrusion' and promote the circularization of 53BP1 nanodomains into one ring-like micro-domain. This ring-like structure can limit the recruitment of BRCA1/CtIP and prevent excessive cleavage of DNA breaks. DSB, double-strand break; DNA-PKcs, DNA-dependent protein kinase catalytic subunit; XRCC4, X-ray repair cross complementing protein 4; RIF1, replication timing regulatory factor 1; TAD, topologically associated domain; XLF, XRCC4-like factor; LIG4, DNA ligase IV; H2AX, H2A histone family member X; ATM, ataxia-telangiectasia mutated; MRN, MRE11-RAD50-NBS1; MDC1, mediator of DNA damage checkpoint protein 1; RNF, ring finger protein; 53BP1; p53-binding protein 1; BRCA1, breast cancer type 1 susceptibility protein; CtIP, C-terminal binding protein (CtBP)-interacting protein; PTIP, Pax transactivation domain-interacting protein; NHEJ, non-homologous DNA end joining.

Figure 3

Upstream regulators of 53BP1 in NHEJ repair. (A) In a stress-free environment, TIRR inhibits the histone binding function of 53BP1 by binding to its Tudor domain, which is known as the 'off switch'. However, upon DNA damage, 53BP1 is recruited to chromatin and promotes DSB NHEJ repair, which is known as the 'on switch'. (B) At DSB ends, the assembly of phosphorylated DNA-PKcs serves as a platform to recruit Artemis, 53BP1 and other NHEJ factors. Post-transcriptional modification of DNA-PKcs affects its ability to promote NHEJ repair. The autophosphorylation or MEK5-dependent phosphorylation of DNA-PKcs contributes to 53BP1 recruitment, and induces DSB-induced microtubule dynamics stress response. The CRL4ADTL-induced ubiquitination degradation of DNA-PKcs inhibits the NHEJ repair. (C) The cell cycle phase is an important determinant of the repair pathway selection at DSB sites. In the G1 phase, the phosphorylation of Chk1 (S317, S345), regulated by ATM and ATR, induces the formation of 53BP1 foci following DNA damage. In the S/G2 phase, the recruitment of 53BP1 is inhibited by the phosphorylation of Chk1 (S59), FOXK1, BRCA1 and acetylated LMNB1 (K134). DSB, double-strand break; DNA-PKcs, DNA-dependent protein kinase catalytic subunit; 53BP1; p53-binding protein 1; NHEJ, non-homologous DNA end joining; Chk1, checkpoint kinase 1; ATM, ataxia-telangiectasia mutated; 53BP1; p53-binding protein 1; BRCA1, breast cancer type 1 susceptibility protein; LMNB1, lamin B1; TIRR, Tudor-interacting repair regulator; CUL4A, cullin 4A; USP14, ubiquitin-specific protease 14; XRCC4, X-ray repair cross complementing protein 4; XLF, XRCC4-like factor; LIG4, DNA ligase IV; DMSR, DNA induced DSB-induced microtubule dynamics stress response; PTIP, Pax transactivation domain-interacting protein; TopBP1, topoisomerase IIβ binding protein 1; ASF1A, anti-silencing function 1A histone chaperone; MDC1, mediator of DNA damage checkpoint protein 1; HR, homologous recombination; FOXK1, forkhead box K1.

Figure 4

Two main downstream pathways of 53BP1. (A) DSB end protection by the ssDNA-binding Shieldin complex (REV7, SHLD3, SHLD2 and SHLD1) limits resection by EXO1 and DNA2-BLM. (B) Shieldin recruits CST and Polα/Primase, promoting the fill in reaction to counteract the DSBs end resection and leave a considerable 3′ overhang (50-60 nt). The initial 5′ end resection also occurs to allow ssDNA binding by Shieldin and CST. The CST/Polα/Primase-mediated fill in reaction helps to control the DSB repair by 53BP1, RIF1 and Shieldin. (C) Sequential phosphorylation events on multiple Ku/DNA-PKcs amino acid clusters favors the initial processing of DNA ends by Artemis. Artemis binds to 53BP1 to promote NHEJ, and consequently to prevent end resection and RAD51-dependent HR repair. DSB, double-strand break; ssDNA, 3′ single-stranded DNA; CST, CTC1-STN1-TEN1; Polα, polymerase-α; 53BP1; p53-binding protein 1; NHEJ, non-homologous DNA end joining; RIF1, replication timing regulatory factor 1; DNA-PKcs, DNA-dependent protein kinase catalytic subunit; PTIP, Pax transactivation domain-interacting protein; XRCC4, X-ray repair cross complementing protein 4; XLF, XRCC4-like factor; LIG4, DNA ligase IV; BRCT, breast cancer type 1 susceptibility protein carboxyl-terminal; CtIP, C-terminal binding protein (CtBP)-interacting protein.