The non-homologous end-joining pathway of S. cerevisiae works effectively in G1-phase cells, and religates cognate ends correctly and non-randomly

Abstract

DNA double-strand breaks (DSBs) are potentially lethal lesions repaired by two major pathways: homologous recombination (HR) and non-homologous end-joining (NHEJ). Homologous recombination preferentially reunites cognate broken ends. In contrast, non-homologous end-joining could ligate together any two ends, possibly generating dicentric or acentric fragments, leading to inviability. Here, we characterize the yeast NHEJ pathway in populations of pure G1 phase cells, where there is no possibility of repair using a homolog. We show that in G1 yeast cells, NHEJ is a highly effective repair pathway for gamma-ray induced breaks, even when many breaks are present. Pulsed-field gel analysis showed chromosome karyotypes following NHEJ repair of cells from populations with multiple breaks. The number of reciprocal translocations was surprisingly low, perhaps zero, suggesting that NHEJ preferentially re-ligates the “correct” broken ends instead of randomly-chosen ends. Although we do not know the mechanism, the preferential correct ligation is consistent with the idea that broken ends are continuously held together by protein-protein interactions or by larger scale chromatin structure.

Saccharomyces cerevisiae Mhr1 can bind Xho I-induced mitochondrial DNA double-strand breaks in vivo

“…Using an in vivo plasmid rejoining assay, it was shown that yku80 null mutants were as defective as yku70, yku80 double mutants in repairing restriction endonuclease (EcoRI, Xho I, or PstI) induced DSBs [47]. Following association of Yku80 with Yku70, the Yku heterodimer binds DNA ends [48] to mediate nuclear DNA DSB repair via C-NHEJ, particularly in G1-phase haploid yeast cells when a homologous DNA template is unavailable for recombinational repair [49]. With regard to its role in mtDNA DSB repair, indirect evidence exists for the involvement of the Yku complex in suppressing mtDNA deletions [18].…”

A New Assay Capturing Chromosome Fusions Shows a Protection Trade-off at Telomeres and NHEJ Vulnerability to Low Density Ionising Radiation

“…Chromosome stability requires that DNA double-strand breaks (DSBs) are repaired by canonical NHEJ or homologous recombination. NHEJ is an efficient pathway directly resealing DSB ends, most often accurately (1)(2)(3)(4). In line with other DNA repair pathways, NHEJ repair sometimes comes with errors.…”

“…In line with other DNA repair pathways, NHEJ repair sometimes comes with errors. Its contribution to mutagenesis is twofold: sequence changes at the break sites and chromosomal rearrangements (1,2,(5)(6)(7)(8)(9)(10). NHEJdependent chromosomal rearrangements include dicentric chromosomes that lead to catastrophic mutational processes, namely chromosome breakage-fusion-bridge cycles and chromothripsis (11)(12)(13).…”

Functional Applications of Systems Biology Tools: Identification of Novel DNA Repair Factors and Peptide Design

“…Yeast has long served as an effective model for both pathways and provided numerous insights into the molecular mechanisms underlying DSB repair 124,131,190,191 . In haploid yeast, HR dominates from S-phase until division when a homologous chromosome is present while NHEJ is the preferred mechanism during G1 when a template sequence is generally not available 192 . Each pathway employs a unique set of proteins to facilitate repair and the highly amenable yeast model facilitates studies of both.…”

“…No other DNA lesion is as severe as a DSB. When a breaks occur, cells halt cell cycle progression until this structural lesion is resolved 192…”