N-2-aminofluorene and N-2 acetylaminofluorene adducts: the local sequence context of an adduct and its chemical structure determine its replication properties.
The strong rat liver carcinogen, N-2-acetylaminofluorene, forms mainly two types of guanine adducts at the C-8 position, the acetylaminofluorene adduct (dGuo-C8-AAF) and the aminofluorene adduct (dGuo-C8-AF). We have constructed different oligonucleotides bearing a single AF lesion at each of the guanine residues of the NarI mutagenesis hot spot (G1G2CG3CC) and analysed the structural distortion induced by this DNA adduct according to the sequence context. At position G1 and G2, the deformation induced by the AF adduct is smaller than the deformation induced by the corresponding acetylated form of this adduct (i.e. the AAF adduct at the G1 and G2), whereas both AF and AAF adducts induce a similar structural change when bound to G3. Single-stranded oligonucleotides modified with AF adducts were used in primer extension replication assays using purified DNA polymerases (PolIII holoenzyme, Klenow fragment (exo+ and exo-), Sequenase 2.0) and the data compared to the AAF containing substrates. Translesion synthesis (complete bypass) is found with all tested polymerases when AF adducts are bound to G1 or G2 while little or no bypass is seen when the AF adduct is bound to G3. On the other hand, irrespective of its position within the NarI sequence, AAF adducts completely block DNA synthesis. The results described in this paper show that the sole knowledge of the chemical structure of an adduct neither determines uniquely the conformational change it induces at the DNA level nor its replication properties. Indeed, although AF adducts are in most cases non-distorting adducts and as a consequence non-replication blocking lesions (as exemplified by adducts at G1 or G2), some AF adducts (as at position G3) behave almost as AAF adducts in terms of the structural distortion induced and its replication blocking property. These findings stress the strong modulation by the local sequence context of the structural and biological consequences of a given adduct.
Belguise-Valladier
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UPR 9003: Cancérogenèse et Mutägenèse Moléculaire et Structurale CNRS, Ecole Supérieure de Biotechnologie de Strasbourg, France.
Fuchs
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D015073:2-Acetylaminofluorene / Q000737:chemistry*; D001483:Base Sequence; D018736:DNA Adducts / Q000737:chemistry*; D004256:DNA Polymerase I / Q000378:metabolism; D004258:DNA Polymerase III / Q000378:metabolism; D004261:DNA Replication; D005449:Fluorenes / Q000737:chemistry*; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D013329:Structure-Activity Relationship
D018736:DNA Adducts; D005449:Fluorenes; D015073:2-Acetylaminofluorene; D004256:DNA Polymerase I; D004258:DNA Polymerase III