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Mutations in a shuttle vector exposed to activated mitomycin C
Author(s) -
Srikanth Nadadur S.,
Mudipalli Anuradha,
Maccubbin Alexander E.,
Gurtoo Hira L
Publication year - 1994
Publication title -
molecular carcinogenesis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.254
H-Index - 97
eISSN - 1098-2744
pISSN - 0899-1987
DOI - 10.1002/mc.2940100105
Subject(s) - biology , shuttle vector , guanine , transversion , mitomycin c , microbiology and biotechnology , dna , base pair , mutation , mutagenesis , genetics , plasmid , dna damage , gene , vector (molecular biology) , nucleotide , recombinant dna
The cytotoxicity of the potent antibiotic and antitumor agent mitomycin C (MMC) is due to its irreversible binding to DNA. Alkylating species generated by bioreductive activation of MMC are known to cause monoadducts and cross‐links in DNA by specifically binding to guanine residues. To gain insight into how these lesions lead to base‐ and sequence‐specific mutations, shuttle vector pSP189 was treated with MMC chemically reduced by treatment with sodium borohydride, replicated in human Ad293 cells, rescued in bacteria, and analyzed for mutations in the supF tRNA gene sequence. The MMC‐induced mutations were predominantly base substitutions. Eighty‐four percent of the base substitutions were transversions, with G:C←T:A the major transversion. Single base deletions were the other major mutational event, and 77% of these were G:C deletions. Base positions 115, 123, and 163 were mutational hot spots based on the frequency of independent mutations. Identification of a single MMC adduct (presumed to be a modified G on the basis of its R f value) and clustering of MMC‐induced mutations at three GC‐rich areas (nt 100–123, 152–163, and 168–176) suggested that the mutational spectrum we found was due to binding of MMC to guanine on either strand of the plasmid DNA. © 1994 Wiley‐Liss, Inc.