Ceruloplasmin is a ferroxidase that interacts with ferroportin to export cellular iron, but is not expressed in neurons. We recently reported that the amyloid precursor protein (APP) is the analogous iron-exporting chaperone for neurons and other cells. The ferroxidase activity of APP has since been called into question. Using a triplex Fe²⁺ oxidation assay, we analyzed the activity of a soluble form of APP (sAPPα) within a buffer of physiological pH and anionic charge, and determined that iron oxidation originated from phosphate. Using various techniques such as flow-cytometry to measure surface presented proteins, we confirmed that endogenous APP is essential for ferroportin persistence on the neuronal surface. Therefore, despite lacking ferroxidase activity, APP still supports iron export from neurons.

Proteolytic cleavage of the amyloid precursor protein (APP) into the Aβ peptide has been an extensively researched mechanism for Alzheimer’s disease, but the normal function of the protein is less understood. APP functions to regulate neuronal iron content by stabilizing the surface presentation of ferroportin—the only iron exporter channel of cells. The present study aims to quantify the contribution of APP to brain and peripheral iron by examining the lifetime impact on brain and liver iron levels in APP knockout mice. Consistent with previous reports, we found that wild-type mice exhibited an age-dependent increase in iron and ferritin in the brain, while no age-dependent changes were observed in the liver. APP ablation resulted in an exaggeration of age-dependent iron accumulation in the brain and liver in mice that was assessed at 8, 12, 18, and 22 months of age. Brain ferroportin levels were decreased in APP knockout mice, consistent with a mechanistic role for APP in stabilizing this iron export protein in the brain. Iron elevation in the brain and liver of APP knockout mice correlated with decreased transferrin receptor 1 and increased ferritin protein levels. However, no age-dependent increase in brain ferritin iron saturation was observed in APP-KO mice despite similar protein expression levels potentially explaining the vulnerability of APP-KO mice to parkinsonism and traumatic brain sequelae. Our results support a crucial role of APP in regulating brain and peripheral iron, and show that APP may act to oppose brain iron elevation during aging.

Pyroglutamate-modified amyloid-β (pE-Aβ) is a highly neurotoxic amyloid-β (Aβ) isoform and is enriched in the brains of individuals with Alzheimer disease compared with healthy aged controls. Pyroglutamate formation increases the rate of Aβ oligomerization and alters the interactions of Aβ with Cu²⁺ and lipids; however, a link between these properties and the toxicity of pE-Aβ peptides has not been established. We report here that Aβ3pE-42 has an enhanced capacity to cause lipid peroxidation in primary cortical mouse neurons compared with the full-length isoform (Aβ(1–42)). In contrast, Aβ(1–42) caused a significant elevation in cytosolic reactive oxygen species, whereas Aβ3pE-42 did not. We also report that Aβ3pE-42 preferentially associates with neuronal membranes and triggers Ca²⁺ influx that can be partially blocked by the N-methyl-d-aspartate receptor antagonist MK-801. Aβ3pE-42 further caused a loss of plasma membrane integrity and remained bound to neurons at significantly higher levels than Aβ(1–42) over extended incubations. Pyroglutamate formation was additionally found to increase the relative efficiency of Aβ-dityrosine oligomer formation mediated by copper-redox cycling.