Chlorambucil-induced high mutation rate and suicidal gene downregulation in a base excision repair-deficient Escherichia coli strain.
Chlorambucil (CLB; N,N-bis(2-chloroethyl)-p-aminophenylbutyric acid) is a bifunctional alkylating agent widely used as an anticancer drug and also as an immunosuppressant. Its chemical structure and clinical experience indicate that CLB is mutagenic and carcinogenic. We have investigated the ability of CLB to induce mutations and gene expression changes in the wild-type (WT) Escherichia coli strain AB1157 and in the base excision repair-deficient (alkA1, tag-1) E. coli strain MV1932 using a rifampicin (rif) forward mutation system and a cDNA array method. The results showed that CLB is a potent mutagen in MV1932 cells compared with the E. coli WT strain AB1157, emphasizing the role of 3-methyladenine DNA glycosylases I and II in protecting the cells from CLB-induced DNA damage and subsequent mutations. Global gene expression profiling revealed that nine genes in WT E. coli and 100 genes in MV1932, of a total of 4290 genes, responded at least 2.5-fold to CLB. Interestingly, all of these MV1932 genes were downregulated, while 22% were upregulated in WT cells. The downregulated genes in MV1932 represented most (19/23) functional categories, and unexpectedly, many of them code for proteins responsible for genomic integrity. These include: (i) RecF (SOS-response, adaptive mutation), (ii) RecC (resistance to cross-linking agents), (iii) HepA (DNA repair, a possible substitute of RecBCD), (iv) Ssb (DNA recombination repair, controls RecBCD), and (v) SbcC (genetic recombination). Our results strongly suggest that in addition to the DNA damage itself, the downregulation of central protecting genes is responsible for the decreased cell survival (demonstrated in a previous work) and the increased mutation rate (this work) of DNA repair-deficient cells, when exposed to CLB.
Salmelin
Camilla
C
Leukemia Research Laboratory, Department of Clinical Chemistry, Tampere University Hospital and Tampere University Medical School, P.O. Box 2000, FIN-33521 Tampere, Finland.
Vilpo
Juhani
J
D000477:Alkylating Agents / Q000494:pharmacology; D001426:Bacterial Proteins / Q000235:genetics; D002699:Chlorambucil / Q000494:pharmacology*; D004260:DNA Repair / Q000235:genetics*; D004798:Enzymes / Q000235:genetics; D004926:Escherichia coli / Q000187:drug effects / Q000235:genetics*; D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects*; D005798:Genes, Bacterial / Q000187:drug effects*; D009153:Mutagens / Q000494:pharmacology*; D020411:Oligonucleotide Array Sequence Analysis; D012329:RNA, Bacterial / Q000187:drug effects
D000477:Alkylating Agents; D001426:Bacterial Proteins; D004798:Enzymes; D009153:Mutagens; D012329:RNA, Bacterial; D002699:Chlorambucil