Site-specific mutagenesis in Escherichia coli by N2-deoxyguanosine adducts derived from the highly carcinogenic fjord-region benzo[c]phenanthrene 3,4-diol 1,2-epoxides.
Although there have been numerous studies of site-specific mutagenesis by dGuo adducts of benzo[a]pyrene diol epoxides (B[a]P DEs), the present study represents the first example of site-specific mutagenesis by dGuo adducts of the highly carcinogenic benzo[c]phenanthrene 3,4-diol 1,2-epoxides (B[c]Ph DEs). The eight adducts that would result from cis- and trans-opening at C-1 of four optically active isomers of B[c]Ph DEs by the N(2)-amino group of dGuo were incorporated into 5'-TTCGAATCCTTCCCCC (context III) and 5'-GGGGTTCCCGAGCGGC (context IV) at the underlined site. These modified oligonucleotides along with unmodified controls were ligated into single-stranded M13mp7L2, which were then used to transfect SOS-induced Escherichia coli. Upon replication of the lesions in each of the two sequence contexts, mutational analysis of the progeny was performed by differential hybridization. For the 16 adducts, the mutation frequencies varied over 2 orders of magnitude with a reasonably even distribution (0.4-1% for three adducts, 1-2% for six adducts, 3-7.4% for five adducts, and one adduct each at 11 and 39%). For all but this last adduct, the mutation frequency for a given B[c]Ph DE adduct was less than for its B[a]P analogue with the same stereochemistry in the same sequence. For the vectors containing adducts with S configuration at the site of attachment of the hydrocarbon to the dGuo base, the main base substitution was G --> T followed by G --> A. In contrast, for the vectors containing adducts with R configuration, the main base substitution was G --> A. The most notable observation in the present study is the low frequency of mutations induced by the B[c]Ph DE-dGuo adducts relative to their B[a]P counterparts. A possible structural basis for this difference is proposed.
Ramos
Leilani A
LA
Laboratory of Comparative Carcinogenesis (Formerly Chemistry of Carcinogenesis Laboratory), National Cancer Institute-Frederick, Frederick, Maryland 21702, USA.
Pontén
Ingrid
I
Dipple
Anthony
A
Kumar
Subodh
S
Yagi
Haruhiko
H
Sayer
Jane M
JM
Kroth
Heiko
H
Kalena
Govind
G
Jerina
Donald M
DM
D001483:Base Sequence; D002273:Carcinogens / Q000737:chemistry / Q000633:toxicity*; D004247:DNA / Q000187:drug effects* / Q000235:genetics; D018736:DNA Adducts / Q000737:chemistry / Q000235:genetics*; D003849:Deoxyguanosine / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics*; D005822:Genetic Vectors; D016297:Mutagenesis, Site-Directed; D009152:Mutagenicity Tests; D009841:Oligonucleotides / Q000138:chemical synthesis / Q000235:genetics; D010616:Phenanthrenes / Q000737:chemistry / Q000633:toxicity*; D017354:Point Mutation; D013014:SOS Response, Genetics; D013237:Stereoisomerism; D014162:Transfection
D002273:Carcinogens; D018736:DNA Adducts; D009841:Oligonucleotides; D010616:Phenanthrenes; C090203:1,2-epoxy-3,4-dihydroxy-1,2,3,4-tetrahydrobenzo(c)phenanthrene; D004247:DNA; D003849:Deoxyguanosine