Formation of Cyclobutane Pyrimidine Dimers after UVA Exposure (Dark-CPDs) Is Inhibited by an Hydrophilic Extract of Polypodium leucotomos

Exposure to sun and especially to ultraviolet radiation (UVR) exerts well known detrimental effects on skin which are implicated in malignancy. UVR induces production of cyclobutane pyrimidine dimers (CPDs), immediately during exposure and even hours after the exposure, these latter being called dark-CPDs, as consequence of the effects of different reactive species that are formed. Fernblock® (FB), an aqueous extract of Polypodium leucotomos, has proven to have photoprotective and antioxidant effects on skin. The aim of our work was to investigate the potential photoprotective effect of FB against dark-CPD formation. Murine melanocytes (B16-F10) were exposed to UVA radiation and the production of dark-CPDs and different reactive oxygen and nitrogen species (ROS and RNS) was measured. Significant dark-CPD formation could be seen at 3h after UVA irradiation, which was inhibited by the pre-treatment of cells with FB. Formation of nitric oxide, superoxide and peroxynitrite was increased after irradiation, consistent with the increased CPD formation. FB successfully reduced the production of these reactive species. Hence, these results show how dark-CPDs are formed in UVA irradiated melanocytes, and that FB acts as a potential antioxidant and ROS scavenger, preventing the DNA damage induced by sun exposure.

Xeroderma Pigmentosum: Gene Variants and Splice Variants

The nucleotide excision repair (NER) is essential for the repair of ultraviolet (UV)-induced DNA damage, such as cyclobutane pyrimidine dimers (CPDs) and 6,4-pyrimidine-pyrimidone dimers (6,4-PPs). Alterations in genes of the NER can lead to DNA damage repair disorders such as Xeroderma pigmentosum (XP). XP is a rare autosomal recessive genetic disorder associated with UV-sensitivity and early onset of skin cancer. Recently, extensive research has been conducted on the functional relevance of splice variants and their relation to cancer. Here, we focus on the functional relevance of alternative splice variants of XP genes.

The major mechanism of melanoma mutations is based on deamination of cytosine in pyrimidine dimers as determined by circle damage sequencing

Sunlight-associated melanomas carry a unique C-to-T mutation signature. UVB radiation induces cyclobutane pyrimidine dimers (CPDs) as the major form of DNA damage, but the mechanism of how CPDs cause mutations is unclear. To map CPDs at single-base resolution genome wide, we developed the circle damage sequencing (circle-damage-seq) method. In human cells, CPDs form preferentially in a tetranucleotide sequence context (5′-Py-T<>Py-T/A), but this alone does not explain the tumor mutation patterns. To test whether mutations arise at CPDs by cytosine deamination, we specifically mapped UVB-induced cytosine-deaminated CPDs. Transcription start sites (TSSs) were protected from CPDs and deaminated CPDs, but both lesions were enriched immediately upstream of the TSS, suggesting a mutation-promoting role of bound transcription factors. Most importantly, the genomic dinucleotide and trinucleotide sequence specificity of deaminated CPDs matched the prominent mutation signature of melanomas. Our data identify the cytosine-deaminated CPD as the leading premutagenic lesion responsible for mutations in melanomas.

2,6-diaminopurine promotes repair of DNA lesions under prebiotic conditions

High-yielding and selective prebiotic syntheses of RNA and DNA nucleotides involve UV irradiation to promote the key reaction steps and eradicate biologically irrelevant isomers. While these syntheses were likely enabled by UV-rich prebiotic environment, UV-induced formation of photodamages in polymeric nucleic acids, such as cyclobutane pyrimidine dimers (CPDs), remains the key unresolved issue for the origins of RNA and DNA on Earth. Here, we demonstrate that substitution of adenine with 2,6-diaminopurine enables repair of CPDs with yields reaching 92%. This substantial self-repairing activity originates from excellent electron donating properties of 2,6-diaminopurine in nucleic acid strands. We also show that the deoxyribonucleosides of 2,6-diaminopurine and adenine can be formed under the same prebiotic conditions. Considering that 2,6-diaminopurine was previously shown to increase the rate of nonenzymatic RNA replication, this nucleobase could have played critical roles in the formation of functional and photostable RNA/DNA oligomers in UV-rich prebiotic environments.

The effect of flanking bases on direct and triplet sensitized cyclobutane pyrimidine dimer formation in DNA depends on the dipyrimidine, wavelength and the photosensitizer

Cyclobutane pyrimidine dimers (CPDs) are the major products of DNA produced by direct absorption of UV light, and result in C to T mutations linked to human skin cancers. Most recently a new pathway to CPDs in melanocytes has been discovered that has been proposed to arise from a chemisensitized pathway involving a triplet sensitizer that increases mutagenesis by increasing the percentage of C-containing CPDs. To investigate how triplet sensitization may differ from direct UV irradiation, CPD formation was quantified in a 129-mer DNA designed to contain all 64 possible NYYN sequences. CPD formation with UVB light varied about 2-fold between dipyrimidines and 12-fold with flanking sequence and was most frequent at YYYR and least frequent for GYYN sites in accord with a charge transfer quenching mechanism. In contrast, photosensitized CPD formation greatly favored TT over C-containing sites, more so for norfloxacin (NFX) than acetone, in accord with their differing triplet energies. While the sequence dependence for photosensitized TT CPD formation was similar to UVB light, there were significant differences, especially between NFX and acetone that could be largely explained by the ability of NFX to intercalate into DNA.