Advanced microscopy techniques used for comparison of UVA- and γ-irradiation-induced DNA damage in the cell nucleus and nucleolus
Jazyk angličtina Země Česko Médium print
Typ dokumentu srovnávací studie, časopisecké články, práce podpořená grantem
PubMed
25369346
PII: FB2014A0037
Knihovny.cz E-zdroje
- MeSH
- 53BP1 MeSH
- antigeny jaderné MeSH
- buněčné jadérko účinky záření MeSH
- červený fluorescenční protein MeSH
- chromozomální proteiny, nehistonové metabolismus MeSH
- DNA vazebné proteiny metabolismus MeSH
- histony metabolismus MeSH
- kinetika MeSH
- luminescentní proteiny metabolismus MeSH
- mikroskopie metody MeSH
- myši MeSH
- poškození DNA * MeSH
- pyrimidinové dimery metabolismus MeSH
- ultrafialové záření * MeSH
- záření gama * MeSH
- zvířata MeSH
- Check Tag
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- srovnávací studie MeSH
- Názvy látek
- 53BP1 MeSH
- antigeny jaderné MeSH
- chromozomální proteiny, nehistonové MeSH
- DNA vazebné proteiny MeSH
- H2AX protein, mouse MeSH Prohlížeč
- histony MeSH
- luminescentní proteiny MeSH
- nucleolar organizer region associated proteins MeSH Prohlížeč
- pyrimidinové dimery MeSH
- Trp53bp1 protein, mouse MeSH Prohlížeč
Every day, genomes are affected by genotoxic factors that create multiple DNA lesions. Several DNA repair systems have evolved to counteract the deleterious effects of DNA damage. These systems include a set of DNA repair mechanisms, damage tolerance processes, and activation of cell-cycle checkpoints. This study describes selected confocal microscopy techniques that investigate DNA damage-related nuclear events after UVA- and γ-irradiation and compare the DNA damage response (DDR) induced by the two experimental approaches. In both cases, we observed induction of the nucleotide excision repair (NER) pathway and formation of localized double-strand breaks (DSBs). This was confirmed by analysis of cyclobutane pyrimidine dimers (CPDs) in the DNA lesions and by increased levels of γH2AX and 53BP1 proteins in the irradiated genome. DNA damage by UVA-lasers was potentiated by either BrdU or Hoechst 33342 pre-sensitization and compared to non-photosensitized cells. DSBs were also induced without BrdU or Hoechst 33342 pre-treatment. Interestingly, no cyclobutane pyrimidine dimers (CPDs) were detected after 405 nm UVA laser micro-irradiation in non-photosensitized cells. The effects of UVA and γ-irradiation were also studied by silver staining of nucleolar organizer regions (AgNORs). This experimental approach revealed changes in the morphology of nucleoli after genome injury. Additionally, to precisely characterize DDR in locally induced DNA lesions, we analysed the kinetics of the 53BP1 protein involved in DDR by fluorescence recovery after photobleaching (FRAP).
Function of heterochromatin protein 1 during DNA repair
HP1β-dependent recruitment of UBF1 to irradiated chromatin occurs simultaneously with CPDs