Lethality induced by a single site-specific double-strand break in a dispensable yeast plasmid.
Cells of the yeast Saccharomyces cerevisiae are delayed in the G2 phase of the cell cycle following chromosomal DNA damage. This arrest is RAD9-dependent and suggests a signaling mechanism(s) between chromosomal lesions and cell cycling. We examined the global nature of growth inhibition caused by an HO endonuclease-induced double-strand break (DSB) at a 45-bp YZ sequence (from MAT YZ) in a non-yeast region of a dispensable single-copy plasmid. The presence of an unrepaired DSB results in cellular death even though the plasmid is dispensable. Loss of cell viability is partially dependent on the RAD9 gene product. Following induction of the DSB, 40% of RAD+ and 49% of rad9 delta cells [including both unbudded (G1) and budded (S plus G2) cells] did not progress further in the cell cycle. The remaining RAD+ cells progressed to form microcolonies (< 30 cells) containing aberrantly shaped inviable cells. For the rad9 delta mutant, the majority of the remaining cells produced viable colonies accounting for the greater survival of the rad9 delta strain. Based on the profound effects of a single nonchromosomal DNA lesion, this system provides a convenient means for studying the signaling effects of a DNA lesion, as well as for designing strategies for modulating cell proliferation.
Laboratory of Molecular Genetics, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, NC 27709.
C B
CB
Bennett
A L
AL
Lewis
K K
KK
Baldwin
M A
MA
Resnick
D002453:Cell Cycle / Q000235:genetics; D018797:Cell Cycle Proteins; D002455:Cell Division; D015825:Chromosomes, Fungal / Q000502:physiology*; D004249:DNA Damage; D015252:Deoxyribonucleases, Type II Site-Specific / Q000096:biosynthesis; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D005800:Genes, Fungal; D005804:Genes, Lethal; D010957:Plasmids; D015183:Restriction Mapping; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins; D013379:Substrate Specificity
D018797:Cell Cycle Proteins; D005656:Fungal Proteins; D029701:Saccharomyces cerevisiae Proteins; C069930:rad9 protein; C530845:HO protein, S cerevisiae; C037725:SCEI protein, S cerevisiae; D015252:Deoxyribonucleases, Type II Site-Specific