Overexpression of human NOX1 complex induces genome instability in mammalian cells.
The production of reactive oxygen species (ROS) in mammalian cells is tightly regulated because of their potential to damage macromolecules, including DNA. To investigate possible links between high ROS levels, oxidative DNA damage, and genomic instability in mammalian cells, we established a novel model of chronic oxidative stress by coexpressing the NADPH oxidase human (h) NOX1 gene together with its cofactors NOXO1 and NOXA1. Transfectants of mismatch repair (MMR)-proficient HeLa cells or MMR-defective Msh2(-/-) mouse embryo fibroblasts overexpressing the hNOX1 complex displayed increased intracellular ROS levels. In one HeLa clone in which ROS were particularly elevated, reactive nitrogen species were also increased and nitrated proteins were identified with an anti-3-nitrotyrosine antibody. Overexpression of the hNOX1 complex increased the steady-state levels of DNA 8-oxo-7,8-dihydroguanine and caused a threefold increase in the HPRT mutation rate in HeLa cells. In contrast, additional oxidatively generated damage did not affect the constitutive mutator phenotype of the Msh2(-/-) fibroblasts. Because no significant changes in the expression of several DNA repair enzymes for oxidative DNA damage were identified, we suggest that chronic oxidative stress can saturate the cell's DNA repair capacity and cause significant genomic instability.
Chiera
Federica
F
Department of Environment and Primary Prevention, Istituto Superiore di Sanità, 00161 Roma, Italy.
Meccia
Ettore
E
Degan
Paolo
P
Aquilina
Gabriele
G
Pietraforte
Donatella
D
Minetti
Maurizio
M
Lambeth
David
D
Bignami
Margherita
M
D048868:Adaptor Proteins, Signal Transducing; D033942:Adaptor Proteins, Vesicular Transport / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D004249:DNA Damage; D042822:Genomic Instability; D006147:Guanine / Q000031:analogs & derivatives / Q000032:analysis / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D007041:Hypoxanthine Phosphoribosyltransferase / Q000235:genetics; D051379:Mice; D051718:MutS Homolog 2 Protein / Q000235:genetics; D016296:Mutagenesis; D000074624:NADPH Oxidase 1; D019255:NADPH Oxidases / Q000235:genetics / Q000378:metabolism*; D018384:Oxidative Stress
D048868:Adaptor Proteins, Signal Transducing; D033942:Adaptor Proteins, Vesicular Transport; C476420:NOXA1 protein, human; C476419:NOXO1 protein, human; C024829:8-hydroxyguanine; D006147:Guanine; D000074624:NADPH Oxidase 1; D019255:NADPH Oxidases; C402799:NOX1 protein, human; D007041:Hypoxanthine Phosphoribosyltransferase; C497173:Msh2 protein, mouse; D051718:MutS Homolog 2 Protein