The D380 residue in MCO1 regulates neural iron homeostasis and fitness in Drosophila

Iron dyshomeostasis has been implicated in the aging and pathogenesis of numerous neurodegenerative diseases, however, the underlying mechanisms are largely unknown. Previously, we determined that Drosophila melanogaster multicopper oxidase-1 (MCO1) has dual-functional catalytic activity of both ferroxidase and ascorbate oxidase (FAO) in vitro, but how MCO1 functions in vivo and what roles it plays in the nervous system are unclear. By using a pan-neuronal elav-Gal4 driver and the CRISPR-Cas9 system, we characterized MCO1 roles in the nervous iron metabolism and dissected the contributions of its ferroxidase and ascorbate oxidase activity in the development, survival and locomotion of D. melanogaster in vivo. We found that MCO1 responds to the dietary iron levels and regulates the iron contents in Drosophila head. Altering the nervous MCO1 expression significantly affects the expression of the key iron metabolic genes, including Malvolio and the Transferrins . MCO1 expression in yeast can partially complement the △Fet3 and △CCC2 mutant cell defects, facilitating iron accumulation in the cell. Abolishing both FAO activities in MCO1 ΔExon IV flies or specifically disrupting ferroxidase activity in MCO1 D380A flies resulted in significant accumulation of ferrous iron and reactive oxygen species (ROS) in the aging Drosophila brain severely impaired development, reduced locomotion, and shortened lifespan. In contrast, elimination of ascorbate oxidase activity alone in MCO1 H374S flies suppressed ferrous iron accumulation and substantially extended lifespan. Our findings underscore MCO1’s role as a vital ferroxidase in the Drosophila brain, providing insights for modulating aging as well as the progress of neurodegenerative diseases.