Cyclobutane Pyrimidine Dimers Are Responsible for the Vast Majority of Mutations Induced by UVB Irradiation in Mammalian Cells

Abstract

The most prevalent DNA lesions induced by UVB are the cyclobutane pyrimidine dimers (CPDs) and the pyrimidine (6-4) pyrimidone photoproducts ((6-4)PPs). It has been a long standing controversy as to which of these photoproduct is responsible for mutations in mammalian cells. Here we have introduced photoproduct-specific DNA photolyases into a mouse cell line carrying the transgenic mutation reporter genes lacI and cII. Exposure of the photolyase-expressing cell lines to photoreactivating light resulted in almost complete repair of either CPDs or (6-4)PPs within less than 3 h. The mutations produced by the remaining, nonrepaired photoproducts were scored. The mutant frequency in the cII gene after photoreactivation by CPD photolyase was reduced from 127 ؋ 10 ؊5 to 34 ؋ 10 ؊5 (background, 8 -10 ؋ 10 ؊5 ). Photoreactivation with (6-4) photolyase did not lower the mutant frequency appreciably. In the lacI gene the mutant frequency after photoreactivation repair of CPDs was reduced from 148 ؋ 10 ؊5 to 28 ؋ 10 ؊5(background, 6 -10 ؋ 10 ؊5 ). Mutation spectra obtained with and without photoreactivation by CPD photolyase indicated that the remaining mutations were derived from background mutations, unrepaired CPDs, and other DNA photopoducts including perhaps a small contribution from (6-4)PPs. We conclude that CPDs are responsible for at least 80% of the UVB-induced mutations in this mammalian cell model.The UV component of sunlight is responsible for the induction of skin tumors, most notably basal cell and squamous cell carcinomas and probably also melanomas (1, 2). Mutations in the p53 gene have been found in a large percentage of human skin malignancies (3-6). The most frequent p53 mutations in skin tumors are C to T or CC to TT mutations involving dipyrimidine sequences. These are considered characteristic mutational changes that can be ascribed to solar UV irradiation (7).The most abundant UV-induced DNA photoproducts are the cis-syn cyclobutane pyrimidine dimers (CPDs) 1 and the pyrimidine (6-4) pyrimidone photoproducts ((6-4)PPs). Both lesions are produced by UVB (280 -320 nm) and UVC (200 -280 nm) irradiation in DNA (8, 9). Most of the mutagenic and carcinogenic action of sunlight has been ascribed to the UVB portion of the solar spectrum (10) with a possible role for UVA (320 -400 nm) in the induction of melanoma (11). The absorption of UVA photons by DNA is rather weak, and it is thought that indirect damaging mechanisms may involve endogenous chromophores as radiation absorbing intermediates (12). These can generate reactive oxygen species, which may then damage DNA. Of all lesions formed in DNA after UVB irradiation, the CPD is considered one of the most important ones based on its relatively high abundance, slow repair, and known mutagenicity (8, 9, 13). However, a strong case can be made that the (6-4)PPs are equally if not more important than the CPDs for inducing mutations. C to T transitions can be induced by both CPDs and (6-4)PPs in mutagenesis studies using site-specific photolesions, and (6-4)PP...

Deamination of 5-Methylcytosines within Cyclobutane Pyrimidine Dimers Is an Important Component of UVB Mutagenesis

“…Previous results using in vivo photoenzymatic removal of specific photoproducts provided evidence that CPDs are responsible for the majority of UVB-induced mutations in mammalian cells (11). The data were consistent with a higher level of formation, slower repair, and increased mutagenicity of CPDs compared with (6 -4)-photoproducts (8 -11).…”

“…We studied the mutations produced after photoproduct-specific photoreactivation and showed that the CPD is responsible for a great majority of the mutations induced by UVB irradiation (11). Earlier, we and others found that sequences that contain 5-methylcytosine within a dipyrimidine are up to 15 times more susceptible to CPD formation after exposure to natural sunlight (12) or a UVB light source (13) compared with 254-nm UVC irradiation.…”

Photorepair of RNA polymerase arrest and apoptosis after ultraviolet irradiation in normal and XPB deficient rodent cells

“…Although no photolyase is found in placental mammals, previous reports have confirmed that heterologous expression of marsupial CPD photolyase has been found to be active in human cells, either reducing mutagenesis 17,18 or preventing UV-induced apoptosis. 2 The The ESS numbers were determined in nuclei by sedimentation in alkaline sucrose gradients as described in Material and Methods.…”

UV-B Radiation Induces Epithelial Tumors in Mice Lacking DNA Polymerase η and Mesenchymal Tumors in Mice Deficient for DNA Polymerase ι

“…In general, however, CPD lesions are removed more slowly from the genome than (6-4)PPs, probably because of the distinct affinity of the global genome NER (GG-NER)-specific XPC protein complex for these UV lesions (3, 30, 57). Besides the difference in repair rates for the two photolesions in vivo, You et al reported that CPDs are responsible for the majority of UV-B-induced mutations in a mammalian mutagenesis assay system (73). Using transgenic mice carrying a specific photolyase for CPD or (6-4) PP, Jans et al also demonstrated that the CPD lesion is the principal cause of cell death, mutation, and skin cancer (24).…”