Influences of base excision repair defects on the lethality and mutagenicity induced by Me-lex, a sequence-selective N3-adenine methylating agent.
Due to its minor groove selectivity, Me-lex preferentially generates N3-methyladenine (3-MeA) adducts in double-stranded DNA. We undertook a genetic approach in yeast to establish the influence of base excision repair (BER) defects on the processing of Me-lex lesions on plasmid DNA that harbors the p53 cDNA as target. We constructed a panel of isogenic strains containing a reporter gene to test p53 function and the following gene deletions: deltamag1, deltaapn1apn2, and deltaapn1apn2mag1. When compared with the wild-type strain, a decrease in survival was observed in deltamag1, deltaapn1apn2, and deltaapn1apn2mag1. The Me-lex-induced mutation frequency increased in the following order: wild type < deltamag1< deltaapn1apn2 = deltaapn1apn2mag1. A total of 77 mutants (23 in wild type, 31 in deltamag1, and 23 in deltaapn1apn2) were sequenced. Eighty-one independent mutations (24 in wild type, 34 in deltamag1, and 23 in deltaapn1apn2) were detected. The majority of base pair substitutions were AT-targeted in all strains (14/23, 61% in wild type; 20/34, 59%, in deltamag1; and 14/23, 61%, in deltaapn1apn2). The Mag1 deletion was associated with a significant decrease of GC > AT transitions when compared with both the wild-type and the AP endonuclease mutants. This is the first time that the impact of Mag1 and/or AP endonuclease defects on the mutational spectra caused by 3-MeA has been determined. The results suggest that 3-MeA is critical for Me-lex cytotoxicity and that its mutagenicity is slightly elevated in the absence of Mag1 glycosylase activity but significantly higher in the absence of AP endonuclease activity.
Monti
Paola
P
Mutagenesis Laboratory, National Cancer Research Institute (IST), L.go R. Benzi, 10, 16132-Genova, Italy.
Campomenosi
Paola
P
Ciribilli
Yari
Y
Iannone
Raffaella
R
Inga
Alberto
A
Shah
Dharini
D
Scott
Gina
G
Burns
Philip A
PA
Menichini
Paola
P
Abbondandolo
Angelo
A
Gold
Barry
B
Fronza
Gilberto
G
D000225:Adenine / Q000378:metabolism; D001483:Base Sequence; D004247:DNA / Q000378:metabolism; D019175:DNA Methylation; D004260:DNA Repair / Q000187:drug effects; D018076:DNA, Complementary / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D017353:Gene Deletion; D016158:Genes, p53; D005822:Genetic Vectors; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009153:Mutagens; D009429:Netropsin / Q000031:analogs & derivatives*; D010957:Plasmids / Q000378:metabolism; D011401:Promoter Regions, Genetic
D018076:DNA, Complementary; D009153:Mutagens; C110963:methyl lexitropsin; D009429:Netropsin; D004247:DNA; D000225:Adenine