Influence of nucleotide excision repair of Escherichia coli on radiation-induced mutagenesis of double-stranded M13 DNA.
To investigate a possible role of nucleotide excision repair (NER) of E. coli in the removal of gamma-radiation-induced DNA lesions, double-stranded M13mp10 DNA, which contains a part of the lac operon, including the promoter/operator region, the lacZ alpha gene and a 144 basepair (bp) inframe insert in the lacZ alpha gene, as mutational target was gamma-irradiated in a phosphate buffer under N2. Subsequently, the radiation-exposed DNA was transfected to wild-type or NER-deficient (uvrA-) E. coli, mutants in the mutational target selected, followed by characterization of the mutants by sequence analysis. Both the mutations obtained from wild-type and uvrA- E. coli appeared to consist mainly of bp substitutions. However, in contrast to wild-type cells, a relatively large proportion of the mutations obtained from the NER-deficient cells (about 25%) is represented by -1 bp deletions, indicating that NER may be responsible for the removal of lesions which cause this particular type of frameshift. Comparison of the bp substitutions between both E. coli strains showed considerable differences. Thirty per cent of all bp substitutions in the NER-deficient host are T/A-->C/G transitions which are virtually absent in wild-type E. coli. This indicates that NER is involved in the elimination of lesions responsible for these transitions. This may also be true for a part of the lesions which cause C/G-->T/A transitions, which make up 52% of the bp substitutions in uvrA- cells versus 17% in wild-type cells. Strikingly, C/G-->G/C transversions appeared to be only formed in wild-type, where they make up 22% of all bp substitutions, and not in the NER-deficient E. coli. This result suggests, that due to the action of NER, a particular type of mutation may be introduced. A similar indication holds for C/G-->A/T transversions, which are predominant in wild-type (58%) and in the minority in uvrA- cells (15%).
Braun
J E
JE
Department of Medical Oncology, Faculty of Medicine, Vrije Universiteit, Amsterdam, Netherlands.
Wanamarta
A H
AH
Westmijze
E J
EJ
Wientjes
N M
NM
Wijker
C A
CA
Lafluer
M V
MV
Retèl
J
J
D001483:Base Sequence; D003090:Coliphages / Q000528:radiation effects*; D004260:DNA Repair; D004279:DNA, Viral / Q000528:radiation effects; D004926:Escherichia coli / Q000235:genetics*; D005720:Gamma Rays; D008969:Molecular Sequence Data; D016296:Mutagenesis / Q000528:radiation effects; D009584:Nitrogen; D017384:Sequence Deletion
D004279:DNA, Viral; D009584:Nitrogen