Oxygen-Dependent Accumulation of Purine DNA Lesions in Cockayne Syndrome Cells

Cockayne Syndrome (CS) is an autosomal recessive neurodegenerative premature agingdisorder associated with defects in nucleotide excision repair (NER). Cells from CS patients, withmutations in CSA or CSB genes, present elevated levels of reactive oxygen species (ROS) andare defective in the repair of a variety of oxidatively generated DNA lesions. In this study,six purine lesions were ascertained in wild type (wt) CSA, defective CSA, wtCSB and defectiveCSB-transformed fibroblasts under different oxygen tensions (hyperoxic 21%, physioxic 5% andhypoxic 1%). In particular, the four 50,8-cyclopurine (cPu) and the two 8-oxo-purine (8-oxo-Pu)lesions were accurately quantified by LC-MS/MS analysis using isotopomeric internal standardsafter an enzymatic digestion procedure. cPu levels were found comparable to 8-oxo-Pu in all cases(3-6 lesions/106 nucleotides), slightly increasing on going from hyperoxia to physioxia to hypoxia.Moreover, higher levels of four cPu were observed under hypoxia in both CSA and CSB-defectivecells as compared to normal counterparts, along with a significant enhancement of 8-oxo-Pu. Thesefindings revealed that exposure to different oxygen tensions induced oxidative DNA damage inCS cells, repairable by NER or base excision repair (BER) pathways. In NER-defective CS patients,these results support the hypothesis that the clinical neurological features might be connected tothe accumulation of cPu. Moreover, the elimination of dysfunctional mitochondria in CS cells isassociated with a reduction in the oxidative DNA damage.