Nonhomologous End-Joining (NHEJ)

Stable Identifier
Homo sapiens
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The nonhomologous end joining (NHEJ) pathway is initiated in response to the formation of DNA double-strand breaks (DSBs) induced by DNA-damaging agents, such as ionizing radiation. DNA DSBs are recognized by the MRN complex (MRE11A:RAD50:NBN), leading to ATM activation and ATM-dependent recruitment of a number of DNA damage checkpoint and repair proteins to DNA DSB sites (Lee and Paull 2005). The ATM phosphorylated MRN complex, MDC1 and H2AFX-containing nucleosomes (gamma-H2AX) serve as scaffolds for the formation of nuclear foci known as ionizing radiation induced foci (IRIF) (Gatei et al. 2000, Paull et al. 2000, Stewart et al. 2003, Stucki et al. 2005). Ultimately, both BRCA1:BARD1 heterodimers and TP53BP1 (53BP1) are recruited to IRIF (Wang et al. 2007, Pei et al. 2011, Mallette et al. 2012), which is necessary for ATM-mediated CHEK2 activation (Wang et al. 2002, Wilson et al. 2008). In G1 cells, TP53BP1 promotes NHEJ by recruiting RIF1 and PAX1IP, which displaces BRCA1:BARD1 and associated proteins from the DNA DSB site and prevents resection of DNA DSBs needed for homologous recombination repair (HRR) (Escribano-Diaz et al. 2013, Zimmermann et al. 2013, Callen et al. 2013). TP53BP1 also plays an important role in ATM-mediated phosphorylation of DCLRE1C (ARTEMIS) (Riballo et al. 2004, Wang et al. 2014). Ku70:Ku80 heterodimer (also known as the Ku complex or XRCC5:XRCC6) binds DNA DSB ends, competing away the MRN complex and preventing MRN-mediated resection of DNA DSB ends (Walker et al. 2001, Sun et al. 2012). The catalytic subunit of the DNA-dependent protein kinase (DNA-PKcs, PRKDC) is then recruited to DNA-bound Ku to form the DNA-PK holoenzyme. Two DNA-PK complexes, one at each side of the break, bring DNA DSB ends together, joining them in a synaptic complex (Gottlieb 1993, Yoo and Dynan 2000). DNA-PK complex recruits DCLRE1C (ARTEMIS) to DNA DSB ends (Ma et al. 2002). PRKDC-mediated phosphorylation of DCLRE1C, as well as PRKDC autophosphorylation, enables DCLRE1C to trim 3'- and 5'-overhangs at DNA DSBs, preparing them for ligation (Ma et al. 2002, Ma et al. 2005, Niewolik et al. 2006). The binding of inositol phosphate may additionally stimulate the catalytic activity of PRKDC (Hanakahi et al. 2000). Other factors, such as polynucleotide kinase (PNK), TDP1 or TDP2 may remove unligatable damaged nucleotides from 5'- and 3'-ends of the DSB, converting them to ligatable substrates (Inamdar et al. 2002, Gomez-Herreros et al. 2013). DNA ligase 4 (LIG4) in complex with XRCC4 (XRCC4:LIG4) is recruited to ligatable DNA DSB ends together with the XLF (NHEJ1) homodimer and DNA polymerases mu (POLM) and/or lambda (POLL) (McElhinny et al. 2000, Hsu et al. 2002, Malu et al. 2002, Ahnesorg et al. 2006, Mahajan et al. 2002, Lee et al. 2004, Fan and Wu 2004). After POLL and/or POLM fill 1- or 2-nucleotide long single strand gaps at aligned DNA DSB ends, XRCC4:LIG4 performs the ligation of broken DNA strands, thus completing NHEJ. The presence of NHEJ1 homodimer facilitates the ligation step, especially at mismatched DSB ends (Tsai et al. 2007). Depending on other types of DNA damage present at DNA DSBs, NHEJ can result in error-free products, produce dsDNA with microdeletions and/or mismatched bases, or result in translocations (reviewed by Povrik et al. 2012).

Literature References
PubMed ID Title Journal Year
22179609 Human Ku70/80 protein blocks exonuclease 1-mediated DNA resection in the presence of human Mre11 or Mre11/Rad50 protein complex

Sun, J, Chen, DJ, Davis, AJ, Lee, KJ

J. Biol. Chem. 2012
22373579 RNF8- and RNF168-dependent degradation of KDM4A/JMJD2A triggers 53BP1 recruitment to DNA damage sites

Sixma, TK, Cui, G, Mer, G, Richard, S, Young, LC, Mallette, FA, Hendzel, MJ, Mattiroli, F

EMBO J. 2012
10802669 ATM-dependent phosphorylation of nibrin in response to radiation exposure

Spring, K, Lavin, MF, Concannon, P, Gatti, RA, Desai-Mehta, A, Khanna, K, Cerosaletti, KM, Young, D, Gatei, M, Kozlov, S

Nat. Genet. 2000
12509254 Defining interactions between DNA-PK and ligase IV/XRCC4

Chen, DJ, Yannone, SM, Hsu, HL

DNA Repair (Amst.) 2002
23306437 53BP1 regulates DSB repair using Rif1 to control 5' end resection

Zimmermann, M, Buonomo, SB, de Lange, T, Lottersberger, F, Sfeir, A

Science 2013
21293379 MMSET regulates histone H4K20 methylation and 53BP1 accumulation at DNA damage sites

You, Z, Lou, Z, Chesi, M, Pei, H, Bergsagel, PL, Luo, K, Wang, L, Qin, Y, Zhang, L, Fei, F

Nature 2011
11955432 Hairpin opening and overhang processing by an Artemis/DNA-dependent protein kinase complex in nonhomologous end joining and V(D)J recombination.

Schwarz, K, Pannicke, U, Lieber, MR, Ma, Y

Cell 2002
23505375 TDP2-dependent non-homologous end-joining protects against topoisomerase II-induced DNA breaks and genome instability in cells and in vivo

Huylebroeck, D, Quintero, C, Zeng, Z, Romero-Granados, R, Gómez-Herreros, F, Alvarez-Quilón, A, Caldecott, KW, Vermeire, L, Umans, L, Cortés-Ledesma, F, Ju, L

PLoS Genet. 2013
23727112 53BP1 mediates productive and mutagenic DNA repair through distinct phosphoprotein interactions

Ge, K, Callen, E, Daniel, JA, Polato, F, Alt, FW, Wesemann, DR, Wong, N, Fernandez-Capetillo, O, Chen, HT, Kruhlak, MJ, Sleckman, BP, Nussenzweig, MC, Starnes, LM, Di Virgilio, M, Lee, JE, Nieto-Soler, M, Santos, M, Faryabi, RB, Tubbs, A, Nussenzweig, A

Cell 2013
12607005 MDC1 is a mediator of the mammalian DNA damage checkpoint.

Elledge, SJ, Bignell, CR, Taylor, AM, Stewart, GS, Wang, B

Nature 2003
10572166 Geometry of a complex formed by double strand break repair proteins at a single DNA end: recruitment of DNA-PKcs induces inward translocation of Ku protein.

Dynan, WS, Yoo, S

Nucleic Acids Res 2000
22529269 Artemis C-terminal region facilitates V(D)J recombination through its interactions with DNA Ligase IV and DNA-PKcs

Hanna, M, Kozlov, M, Greene, M, Malu, S, Tempst, P, Erdjument-Bromage, H, Kurosawa, A, Escalante, CR, De Ioannes, P, Cortes, P, Aggarwal, AK, Pena, J, Francis, D, Adachi, N, Villa, A, Vezzoni, P

J. Exp. Med. 2012
8422676 The DNA-dependent protein kinase: requirement for DNA ends and association with Ku antigen.

Gottlieb, TM

Cell 1993
12023295 Conversion of phosphoglycolate to phosphate termini on 3' overhangs of DNA double strand breaks by the human tyrosyl-DNA phosphodiesterase hTdp1

Rasouli-Nia, A, Lees-Miller, SP, Inamdar, KV, Povirk, LF, Zhou, T, Pouliot, JJ

J Biol Chem 2002
10757784 Ku recruits the XRCC4-ligase IV complex to DNA ends

Snowden, CM, Ramsden, DA, McCarville, J, Nick McElhinny, SA

Mol. Cell. Biol. 2000
19001859 NFBD1/MDC1, 53BP1 and BRCA1 have both redundant and unique roles in the ATM pathway

Stern, DF, Wilson, KA

Cell Cycle 2008
12364621 53BP1, a mediator of the DNA damage checkpoint.

Elledge, SJ, Carpenter, PB, Wang, B, Matsuoka, S

Science 2002
16093244 The DNA-dependent protein kinase catalytic subunit phosphorylation sites in human Artemis

Lieber, MR, Schwarz, K, Lu, H, Pannicke, U, Niewolik, D, Ma, Y

J. Biol. Chem. 2005
23333306 A cell cycle-dependent regulatory circuit composed of 53BP1-RIF1 and BRCA1-CtIP controls DNA repair pathway choice

Durocher, D, Canny, MD, Fradet-Turcotte, A, Tká?, J, Xing, M, Rosebrock, AP, Cook, MA, Orthwein, A, Munro, M, Young, JT, Escribano-Díaz, C, Xu, D

Mol. Cell 2013
11030616 Binding of inositol phosphate to DNA-PK and stimulation of double-strand break repair.

West, SC, Bartlet-Jones, M, Chappell, C, Hanakahi, LA, Pappin, D

Cell 2000
10959836 A critical role for histone H2AX in recruitment of repair factors to nuclear foci after DNA damage.

Paull, TT, Bonner, WM, Kirchgessner, CU, Gellert, M, Rogakou, EP, Yamazaki, V

Curr Biol 2000
17470781 Cernunnos/XLF promotes the ligation of mismatched and noncohesive DNA ends

Kim, SA, Chu, G, Tsai, CJ

Proc. Natl. Acad. Sci. U.S.A. 2007
16377563 MDC1 directly binds phosphorylated histone H2AX to regulate cellular responses to DNA double-strand breaks

Smerdon, SJ, Clapperton, JA, Jackson, SP, Yaffe, MB, Stucki, M, Mohammad, D

Cell 2005
15790808 ATM activation by DNA double-strand breaks through the Mre11-Rad50-Nbs1 complex

Paull, TT, Lee, JH

Science 2005
16439205 XLF interacts with the XRCC4-DNA ligase IV complex to promote DNA nonhomologous end-joining

Smith, P, Jackson, SP, Ahnesorg, P

Cell 2006
17525340 Abraxas and RAP80 form a BRCA1 protein complex required for the DNA damage response

Elledge, SJ, Matsuoka, S, Gygi, SP, Wang, B, Smogorzewska, A, Zhang, D, Ballif, BA

Science 2007
25512557 PTIP associates with Artemis to dictate DNA repair pathway choice

Lobrich, M, Aroumougame, A, Gong, Z, Chen, J, Chen, D, Li, Y, Wang, J

Genes Dev. 2014
14561766 Implication of DNA polymerase lambda in alignment-based gap filling for nonhomologous DNA end joining in human nuclear extracts

Lee, JW, Bebenek, K, Garcia-Diaz, M, Blanco, L, Wang, Z, Kunkel, TA, Povirk, LF, Zhou, T

J. Biol. Chem. 2004
15451442 DNA polymerase lambda can elongate on DNA substrates mimicking non-homologous end joining and interact with XRCC4-ligase IV complex

Fan, W, Wu, X

Biochem. Biophys. Res. Commun. 2004
12077346 Association of DNA polymerase mu (pol mu) with Ku and ligase IV: role for pol mu in end-joining double-strand break repair

Mahajan, KN, Ramsden, DA, Nick McElhinny, SA, Mitchell, BS

Mol. Cell. Biol. 2002
15574327 A pathway of double-strand break rejoining dependent upon ATM, Artemis, and proteins locating to gamma-H2AX foci

Recio, MJ, Löbrich, M, Kühne, M, Jackson, SP, Parker, AR, Doherty, A, Dahm, K, Rief, N, Riballo, E, Smith, GC, Fricke, A, Gennery, A, Jeggo, PA, Reis, C, Krempler, A

Mol. Cell 2004
11493912 Structure of the Ku heterodimer bound to DNA and its implications for double-strand break repair.

Goldberg, J, Corpina, RA, Walker, JR

Nature 2001
16914548 DNA-PKcs dependence of Artemis endonucleolytic activity, differences between hairpins and 5' or 3' overhangs

Kulesza, P, Schwarz, K, Lieber, MR, Lu, H, Wang, LC, Pannicke, U, Zandi, E, Niewolik, D, Ma, Y

J. Biol. Chem. 2006
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