Recent epidemiological and clinical data suggest that persons with low folic acid levels and elevated homocysteine levels are at increased risk of Alzheimer's disease (AD), but the underlying mechanism is unknown. We tested the hypothesis that impaired one-carbon metabolism resulting from folic acid deficiency and high homocysteine levels promotes accumulation of DNA damage and sensitizes neurons to amyloid beta-peptide (Abeta) toxicity. Incubation of hippocampal cultures in folic acid-deficient medium or in the presence of methotrexate (an inhibitor of folic acid metabolism) or homocysteine induced cell death and rendered neurons vulnerable to death induced by Abeta. Methyl donor deficiency caused uracil misincorporation and DNA damage and greatly potentiated Abeta toxicity as the result of reduced repair of Abeta-induced oxidative modification of DNA bases. When maintained on a folic acid-deficient diet, amyloid precursor protein (APP) mutant transgenic mice, but not wild-type mice, exhibited increased cellular DNA damage and hippocampal neurodegeneration. Levels of Abeta were unchanged in the brains of folate-deficient APP mutant mice. Our data suggest that folic acid deficiency and homocysteine impair DNA repair in neurons, which sensitizes them to oxidative damage induced by Abeta.