Mutation spectrum resulting in M13mp2 phage DNA exposed to N-nitrosoproline with UVA irradiation.

N-nitrosoproline (NPRO) is endogenously formed from proline and nitrite. In an effort to delineate the mechanism of NPRO-induced photomutagenicity, we investigated the mutagenic spectrum of NPRO on M13mp2 DNA with UVA irradiation. Following exposure to NPRO and UVA, the mutation frequency increased significantly in an NPRO and UVA dose-dependent manner. The sequence data derived from seventy of the mutants indicated that mutagenesis resulted mainly from an increase in single-base substitutions, the most frequent being GC to CG transversions. Non-clustering of the GC to CG mutations suggests that NPRO+UVA damage to DNA is random. These transversions may be caused by guanine adducts in DNA or in part by oxidatively modified guanine in DNA exposed to NPRO and UVA.

Mutation, DNA strand cleavage and nitric oxide formation caused by N-nitrosoproline with sunlight: a possible mechanism of UVA carcinogenicity.

N-Nitrosoproline (NPRO) is endogenously formed from proline and nitrite. NPRO has been reported to be nonmutagenic and noncarcinogenic. In this study, we have detected the direct mutagenicity of NPRO plus natural sunlight towards Salmonella typhimurium. Furthermore, formation of 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG), a mutagenic lesion, was observed in calf thymus DNA treated with NPRO plus simulated sunlight. The treatment with NPRO and sunlight induced single strand breaks in the superhelical replicative form of phage M13mp2 DNA. Single-strand DNA breaks also occurred in the human fibroblast cells on treatment with NPRO plus UVA, as detected by the comet assay. An analysis using scavengers suggested that both reactive oxygen species and NO radical mediate the strand breaks. The formation of nitric oxide was observed in NPRO solution irradiated with UVA. We analyzed the photodynamic spectrum of mutation induction and DNA breakage using monochromatic radiation at a series of wavelengths between 300 and 400 nm. Both mutation frequencies and DNA breakage were highest at the absorption maximum of NPRO, 340 nm. The co-mutagenic and co-toxic actions of NPRO and sunlight merit attention as possible mechanisms increasing the carcinogenic risk from UVA irradiation.