The Ferroxidase Hephaestin But Not Amyloid Precursor Protein is Required for Ferroportin-Supported Iron Efflux in Primary Hippocampal Neurons
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Iron efflux in mammalian cells is mediated by the ferrous iron exporter ferroportin (Fpn); Fpn plasma membrane localization and function are supported by a multicopper ferroxidase and/or the soluble amyloid precursor protein (sAPP). Fpn and APP are ubiquitously expressed in all cell types in the central nervous system including neurons. In contrast, neuronal ferroxidase(s) expression has not been well characterized. Using primary cultures of hippocampal neurons, we examined the molecular mechanism of neuronal Fe efflux in detail. Developmental increases of Fpn, APP, and the ferroxidase hephaestin (Hp) were observed in hippocampal neurons. Iron efflux in these neurons depended on the level of Fpn localized at the cell surface; as noted, Fpn stability is supported by ferroxidase activity, an enzymatic activity that is required for Fe efflux. Iron accumulation increases and iron efflux decreases in Hp knockout neurons. In contrast, suppression of endogenous APP by RNAi knockdown does not affect surface Fpn stability or Fe efflux. These data support the model that the neuronal ferroxidase Hp plays a unique role in support of Fpn-mediated Fe efflux in primary hippocampal neurons. Our data also demonstrate that Hp ferroxidase activity relies on copper bioavailability, which suggests neuronal iron homeostasis will be modulated by cellular copper status.
KeywordsPrimary hippocampal neurons Iron efflux Ferroportin (Fpn) Hephaestin (Hp) Amyloid precursor protein (APP) Ferroxidase
This work was supported by Grants DK053820 and NS095063 from the National Institutes of Health to DJK.
Changyi Ji and Daniel J. Kosman designed the study, analyzed the data, and wrote the paper. Changyi Ji, Brittany Steimle, and Danielle Bailey performed the experiments.
Compliance with Ethical Standards
Conflict of interest
The authors declare that they have no competing interests.
All contributors to this manuscript have participated in the Ethical Standards and Scientific Integrity program conducted by the Vice President of Research at the University at Buffalo.
Research Involved in Animal Rights
All applicable international, national, and institutional guidelines for the care and use of animals were followed. All procedures performed in studies involving animals were in accordance with the ethical standards of the State University of New York. The use of animals in this research was approved and supervised by the Animal Care and Use Committee and the Division of Comparative Medicine and Laboratory Animal Facilities in the Jacobs School of Medicine and Biomedical Sciences, The University at Buffalo.
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