Increased SOD1 association with chromatin, DNA damage, p53 activation, and apoptosis in a cellular model of SOD1-linked ALS.
Mutations in the gene encoding cytosolic Cu,Zn-superoxide dismutase (SOD1) have been linked to familial amyotrophic lateral sclerosis (FALS). However the molecular mechanisms of motor neuron death are multi-factorial and remain unclear. Here we examined DNA damage, p53 activity and apoptosis in SH-SY5Y human neuroblastoma cells transfected to achieve low-level expression of either wild-type or mutant Gly(93)-->Ala (G93A) SOD1, typical of FALS. DNA damage was investigated by evaluating the levels of 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodGuo) and DNA strand breaks. Significantly higher levels of DNA damage, increased p53 activity, and a greater percentage of apoptotic cells were observed in SH-SY5Y cells transfected with G93A SOD1 when compared to cells overexpressing wild-type SOD1 and untransfected cells. Western blot, FACS, and confocal microscopy analysis demonstrated that G93A SOD1 is present in the nucleus in association with DNA. Nuclear G93A SOD1 has identical superoxide dismutase activity but displays increased peroxidase activity when compared to wild-type SOD1. These results indicate that the G93A mutant SOD1 association with DNA might induce DNA damage and trigger the apoptotic response by activating p53. This toxic activity of mutant SOD1 in the nucleus may play an important role in the complex mechanisms associated with motor neuron death observed in ALS pathogenesis.
Barbosa
Livea F
LF
Departamento de Bioquímica, Instituto de Química, Universidade de São Paulo, São Paulo, SP, Brazil.
Cerqueira
Fernanda M
FM
Macedo
Antero F A
AF
Garcia
Camila C M
CC
Angeli
José Pedro F
JP
Schumacher
Robert I
RI
Sogayar
Mari Cleide
MC
Augusto
Ohara
O
Carrì
Maria Teresa
MT
Di Mascio
Paolo
P
Medeiros
Marisa H G
MH
D000080242:8-Hydroxy-2'-Deoxyguanosine; D000690:Amyotrophic Lateral Sclerosis / Q000378:metabolism* / Q000473:pathology; D017209:Apoptosis; D002467:Cell Nucleus / Q000201:enzymology / Q000473:pathology; D002843:Chromatin / Q000378:metabolism*; D004249:DNA Damage; D003849:Deoxyguanosine / Q000031:analogs & derivatives / Q000378:metabolism; D006801:Humans; D007124:Immunoenzyme Techniques; D015227:Lipid Peroxidation; D009447:Neuroblastoma / Q000378:metabolism*; D013482:Superoxide Dismutase / Q000378:metabolism*; D000072105:Superoxide Dismutase-1; D014407:Tumor Cells, Cultured; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism*
D002843:Chromatin; C000606290:SOD1 protein, human; C495901:TP53 protein, human; D016159:Tumor Suppressor Protein p53; D000080242:8-Hydroxy-2'-Deoxyguanosine; D013482:Superoxide Dismutase; D000072105:Superoxide Dismutase-1; D003849:Deoxyguanosine