The recently described doppel protein (Dpl) is a homologue of the prion protein (PrPc). This protein, expressed in the brains of mice that lack the expression of PrPc, causes neuronal death as the mice age. Previous studies have suggested this neuronal damage is caused by oxidative assault and changes in the activity of NOS proteins. We investigated the toxicity of Dpl in cell culture models and showed that Dpl was toxic to neurons. This toxicity was inhibited by the expression of PrPc and possibly involved direct interaction between the two proteins. The mechanism of toxicity involved stimulation of nitric oxide production via activation of the nitric oxide synthases, nNOS and iNOS. This mechanism of toxicity is quite different from that of PrPSc and does not require the protein to change conformation. These results provide the first evidence for the mechanism of Dpl toxicity.