title: The S. cerevisiae Mag1 3-methyladenine DNA glycosylase modulates susceptibility to homologous recombination.
abstract: DNA glycosylases, such as the Mag1 3-methyladenine (3MeA) DNA glycosylase, initiate the base excision repair (BER) pathway by removing damaged bases to create abasic apurinic/apyrimidinic (AP) sites that are subsequently repaired by downstream BER enzymes. Although unrepaired base damage may be mutagenic or recombinogenic, BER intermediates (e.g. AP sites and strand breaks) may also be problematic. To investigate the molecular basis for methylation-induced homologous recombination events in Saccharomyces cerevisiae, spontaneous and methylation-induced recombination were studied in strains with varied MAG1 expression levels. We show that cells lacking Mag1 have increased susceptibility to methylation-induced recombination, and that disruption of nucleotide excision repair (NER; rad4) in mag1 cells increases cellular susceptibility to these events. Furthermore, expression of Escherichia coli Tag 3MeA DNA glycosylase suppresses recombination events, providing strong evidence that unrepaired 3MeA lesions induce recombination. Disruption of REV3 (required for polymerase zeta (Pol zeta)) in mag1 rad4 cells causes increased susceptibility to methylation-induced toxicity and recombination, suggesting that Pol zeta can replicate past 3MeAs. However, at subtoxic levels of methylation damage, disruption of REV3 suppresses methylation-induced recombination, indicating that the effects of Pol zeta on recombination are highly dose-dependent. We also show that overproduction of Mag1 can increase the levels of spontaneous recombination, presumably due to increased levels of BER intermediates. However, additional APN1 endonuclease expression or disruption of REV3 does not affect MAG1-induced recombination, suggesting that downstream BER intermediates (e.g. single strand breaks) are responsible for MAG1-induced recombination, rather than uncleaved AP sites. Thus, too little Mag1 sensitizes cells to methylation-induced recombination, while too much Mag1 can put cells at risk of recombination induced by single strand breaks formed during BER.
authors.affiliation: Division of Bioengineering and Environmental Health, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
authors.forename: C A
authors.initials: CA
authors.lastname: Hendricks
authors.forename: M
authors.initials: M
authors.lastname: Razlog
authors.forename: T
authors.initials: T
authors.lastname: Matsuguchi
authors.forename: A
authors.initials: A
authors.lastname: Goyal
authors.forename: A L
authors.initials: AL
authors.lastname: Brock
authors.forename: B P
authors.initials: BP
authors.lastname: Engelward
mesh_terms: D001080:Apurinic Acid / Q000737:chemistry / Q000378:metabolism; D002470:Cell Survival / Q000235:genetics; D004249:DNA Damage; D045647:DNA Glycosylases; D019175:DNA Methylation; D004260:DNA Repair / Q000235:genetics*; D004271:DNA, Fungal / Q000235:genetics; D004259:DNA-Directed DNA Polymerase / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*; D004926:Escherichia coli / Q000201:enzymology; D009699:N-Glycosyl Hydrolases / Q000502:physiology*; D011743:Pyrimidines / Q000737:chemistry / Q000378:metabolism; D011995:Recombination, Genetic; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics
chemical_list: D004271:DNA, Fungal; D011743:Pyrimidines; D029701:Saccharomyces cerevisiae Proteins; D001080:Apurinic Acid; C425929:DNA polymerase zeta; D004259:DNA-Directed DNA Polymerase; C092602:REV3 protein, S cerevisiae; D045647:DNA Glycosylases; D009699:N-Glycosyl Hydrolases; C445668:MAG1 protein, S cerevisiae