304 - Ontogenetic Regulation of NCOA4-Mediated Iron Release from Ferritin (Ferritinophagy) in the Developing Mouse Brain

The developing brain has high iron requirements that, if not met, manifest as iron deficiency (ID), which causes neurobehavioral deficits in children that persist into adolescence and adulthood. Yet, excess free iron can be cytotoxic and cause neuronal damage. Thus, brain iron levels must be tightly regulated by storing unused cellular iron in ferritin. The regulatory mechanism(s) controlling ferritin iron release during times of high brain iron demand (development) are unknown. Selective ferritin breakdown (ferritinophagy) to release iron from ferritin is essential for erythrocyte development and requires nuclear receptor coactivator 4 (NCOA4) as a cargo receptor targeting ferritin. Whether NCOA4 is expressed in the brain and regulates brain cellular iron homeostasis is unknown.


Objective:

Our goal was to test the hypothesis that NCOA4-mediated ferritinophagy is developmentally regulated with highest activity during peak iron-dependent brain development.


Design/Methods:

Cerebral cortex and hippocampus brain regions were collected from CD1 mice on embryonic day 17 (E17), postnatal day 0 (P0), P7 and P14 (n = 4-8 per age), before and during the period of rapid iron-dependent brain development (~P7-P21). qPCR and western blotting were used to determine mRNA and protein levels for ferritin and NCOA4.


Results:

Transcript abundance for Fth1 increased and Ftl1 decreased across development in the cerebral cortex (Table 1), indicating a switch from iron storage (ferritin L) to utilization (ferritin H). Ncoa4 mRNA levels were relatively stable across cortical development, with a 25% decrease between P7 and P14. Total ferritin protein abundance peaked at E17 and P0 and decreased by ~70% and ~90% at P7 and P14 (Table 2), indicating increased mobilization of stored iron between P7 and P14. NCOA4 protein levels were highest at P0/P7 just prior to the decrease in ferritin protein, suggesting the physiological importance of NCOA4-mediated ferritinophagy during this period. Similar effects were observed in the hippocampus (data not shown).


Conclusion(s):

Our findings demonstrate, for the first time, that NCOA4 is expressed in the developing brain. NCOA4 protein, but not mRNA, is regulated developmentally and is inversely correlated with ferritin protein abundance. This is consistent with the mechanism by which NCOA4 post-translationally responds to iron in other cell types. Understanding how NCOA4-mediated ferritinophagy regulates the brain labile iron pool across the lifespan will provide novel insight into common disorders of brain iron dysregulation (e.g., early-life ID and later-life neurodegenerative disorders).