#NHEJ

Repair Pathway

Final ligation in NHEJ is done by

Final sealing of processed DNA ends in classical non-homologous end joining performed by DNA ligase IV, dedicated ligase distinct from replication ligase I and base excision ligase III. Ligase IV forms stable obligate complex with XRCC4 scaffold and accessory factor XLF/Cernunnos plus PAXX, enhancing adenylation and end alignment. After Ku and DNA-PKcs orchestrate end processing, XRCC4-Ligase IV complex catalyzes ATP-dependent phosphodiester bond formation joining broken termini. Patient mutations in LIG4 cause immunodeficiency and radiosensitivity, highlighting essential role in lymphocyte V(D)J recombination and general double-strand break repair pathway completion.

Ref: Lodish Chapter 11: NHEJ ligation by Ligase IV XRCC4 complex; J Biol Chem 2018 Mechanism of Ligase IV end joining

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Repair Pathway

Broken DNA ends in NHEJ are recognized by

Ku heterodimer composed of 70 kDa and 80 kDa subunits forms preformed ring with central aperture approximately 30 angstroms accommodating double-stranded DNA ends without sequence specificity. Abundant nuclear protein diffuses, threading onto broken terminus within seconds, protecting end from nucleolytic degradation and acting as scaffold for non-homologous end joining machinery. Ku recruits DNA-dependent protein kinase catalytic subunit, Artemis nuclease, polymerases mu and lambda, and XRCC4-Ligase IV complex. By synapsing two Ku-bound ends, it holds broken chromosome together. Rapid binding also antagonizes resection, steering repair away from homologous recombination.

Ref: Alberts Molecular Biology of Cell Chapter 5: Ku70/Ku80 end recognition in NHEJ; Nature Struct Mol Biol 2017 Ku-DNA complex

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Repair Pathway

NHEJ is predominant in which cell cycle phase?

Pathway choice for double-strand break repair depends strongly on cell cycle stage and resection control. In G0 quiescence and G1 gap where sister chromatids absent, extensive 5' resection blocked by 53BP1-RIF1-Shieldin complex and Ku end-binding factor prevents homology search. Therefore non-homologous end joining dominates, initiated by Ku70/Ku80 rapidly capping ends and recruiting DNA-PKcs and ligase IV. In S/G2, CDK phosphorylation activates CtIP enabling resection favoring homologous recombination. Thus NHEJ predominance in G0/G1 reflects availability of template and protective mechanisms limiting homologous recombination when homolog unavailable.

Ref: Berg Biochemistry Chapter 28: DSB repair cell cycle choice; NCBI Bookshelf NHEJ in G1 predominance regulation

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Recombination

Ku70/Ku80 proteins function in

After double-strand break, Ku heterodimer comprising Ku70 and Ku80 forms ring-shaped structure that threads onto each DNA end with high affinity irrespective of sequence, protecting ends from excessive resection. Ku serves as scaffold recruiting DNA-dependent protein kinase catalytic subunit, end-processing factors, and Ligase IV complex. By blocking homologous recombination machinery, Ku channels repair toward NHEJ, especially in mammalian G1 cells or immune lymphocytes generating immunoglobulin diversity. Absence of Ku shifts repair toward microhomology-mediated alternative end joining or homologous recombination pathways, increasing resection length and genome instability.

Ref: Lodish et al., Molecular Cell Biology, 9th ed., Chapter 12: Ku70 Ku80 Function in NHEJ Pathway

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Recombination

Non-homologous end joining (NHEJ) is

Non-homologous end joining ligates broken chromosome ends without requiring homologous donor template, acting throughout cell cycle especially G1. Core factors Ku70/Ku80 anchor DNA ends, DNA-PKcs protects, Artemis trims, polymerases mu/lambda add nucleotides, and Lig4-XRCC4 seals ligation. Since processing removes or adds bases before joining, small insertions or deletions often arise at junction, causing frameshifts and mutations. This error-prone nature distinguishes it from homologous recombination which copies intact template accurately, but affords rapid survival when sister chromatid unavailable, critical for V(D)J recombination and class switching.

Ref: Alberts et al., Molecular Biology of the Cell, 7th ed., Chapter 5: NHEJ as Error-Prone End Joining Mechanism

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