Nonhomologous end joining (NHEJ), the direct rejoining of DNA double-strand breaks, is closely associated with illegitimate recombination and chromosomal rearrangement. This has led to the concept that NHEJ is error prone. Studies with the yeast Saccharomyces cerevisiae have revealed that this model eukaryote has a classical NHEJ pathway dependent on Ku and DNA ligase IV, as well as alternative mechanisms for break rejoining. The evolutionary conservation of the Ku-dependent process includes several genes dedicated to this pathway, indicating that classical NHEJ at least is a strong contributor to fitness in the wild. Here we review how double-strand break structure, the yeast NHEJ proteins, and alternative rejoining mechanisms influence the accuracy of break repair. We also consider how the balance between NHEJ and homologous repair is regulated by cell state to promote genome preservation. The principles discussed are instructive to NHEJ in all organisms.

In mammalian cells, repair of DNA double-strand breaks (DSBs) by nonhomologous end-joining (NHEJ) is critical for genome stability. Although the end-bridging and ligation steps of NHEJ have been reconstituted in vitro, little is known about the end-processing reactions that occur before ligation. Recently, functionally homologous end-bridging and ligation activities have been identified in prokarya. Consistent with its homology to polymerases and nucleases, we demonstrate that DNA ligase D from Mycobacterium tuberculosis (Mt-Lig) possesses a unique variety of nucleotidyl transferase activities, including gap-filling polymerase, terminal transferase, and primase, and is also a 3' to 5' exonuclease. These enzyme activities allow the Mt-Ku and Mt-Lig proteins to join incompatible DSB ends in vitro, as well as to reconstitute NHEJ in vivo in yeast. These results demonstrate that prokaryotic Ku and ligase form a bona fide NHEJ system that encodes all the recognition, processing, and ligation activities required for DSB repair.