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| Author(s): |
Barbara Helen Cortat Santos
Total Authors: 1
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| Document type: | Master's Dissertation |
| Press: | São Paulo. |
| Institution: | Universidade de São Paulo (USP). Instituto de Biociências (IBIOC/SB) |
| Defense date: | 2010-10-06 |
| Examining board members: |
Carlos Frederico Martins Menck;
Eduardo Gorab;
Nadja Cristhina de Souza Pinto Lardner
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| Advisor: | Carlos Frederico Martins Menck |
| Abstract | |
Ultraviolet radiation (UV) is absorbed by different cellular molecules, including DNA in which induces structural distortions. The most common lesions induced by UV radiation are the cyclobutane pyrimidine (CPD) and the photoproduct (6-4)-pyrimidine-pyrimidone [(6-4)PP]. These lesions can be repaired by the photoreactivation, characterized by a single protein (photolyase) that removes lesions using visible light (320-500 nm) as energy source. Two types of photolyases had been identified that differ by their substrate specificity: CPD-photolyase and (6-4)-photolyase. Another repair mechanism is the nucleotide excision repair (NER), a mechanism that involves multiple steps and proteins. While the genotoxic effects of UVB and UVC are already relatively well-understood and accepted, there is still controversy about the genotoxicity of UVA radiation, due to its low absorption by DNA. Some authors believe that the major effects are generated indirectly by the production of reactive oxygen species, while others believe that UVA can cause damage to DNA directly, inducing the formation of pyrimidine dimers. The aim of this study was to assess the genotoxic effects of UVA radiation in human fibroblasts deficient and proficient in NER, using recombinant adenovirus expressing the photolyases to verify if CPDs and (6-4)PPs are generated by UVA and whether they had any importance in the responses observed after irradiation. It was found that cells deficient in the XPA gene are more sensitive to UV radiation compared to wild type cells. By immunological detection, we confirm the generation of CPD, (6-4)PP and Dewar, photoisomer of the (6-4)PP lesion, in the genome of human cells after irradiation with UVA. Using adenoviral vectors for the transduction of photolyases specific for CPD or (6-4)PP lesions, we confirm that in fact these lesions are generated in human cells deficient in DNA repair after irradiation with UVA. Moreover, these viruses allowed us to verify the biological relevance of these lesions in the induction of cell death in irradiated XP-A cells. In fact, our data indicates that for low doses of UVA radiation, these lesions play important roles in the induction of death. We cannot rule out, however, that indirect lesions (probably caused by oxidative stress) could also have a role in the induction of death by UVA radiation, which seems to be more important in intermediate and high doses of this radiation. (AU) | |