Browsing by Subject "NCOA4"

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    NCOA4 is Essential for HT22 Hippocampal Neuronal Cell Survival During Iron Deficiency
    (2020-06) Bengson, Emily
    Iron is essential for proper cell function and development, serving as a cofactor for many metalloproteins. However, mismanagement of cellular iron leads to increased reactive oxygen species (ROS), which ultimately leads to ferroptosis, a newly discovered form of regulated cell death, which is iron-dependent. Neurodegeneration has been associated with brain iron accumulation and, more recently, ferroptosis. NCOA4 manages the cellular labile iron pool by controlling the release of ferritin iron via ferritinophagy. The present studies examined the capacity of hippocampal cells in handling iron fluctuation, with particular focus on NCOA4 and ferritin. HT22 mouse hippocampal cells were treated with ferric ammonium citrate (FAC) and deferoxamine (Dfo) to produce cellular iron overload and deprivation, respectively. Ferroptosis was determined by measures of ferrostatin-1 effects and Ptgs2 mRNA. For ferritinophagy studies, Ncoa4 was silenced by siRNA transfections. Functional impacts of impaired ferritinophagy were assessed via CCK-8 cell viability assays and western and qPCR analyses of iron-related genes. HT22 cells were highly susceptible to cellular iron overload. FAC-treated cells featured acute morphological changes, decreased viability, and elevated Ptgs2 mRNA abundance. Iron effects were prevented by ferrostatin-1, indicating ferroptosis by cellular iron overload. Dfo alone had minimal impact on cell morphology and viability. NCOA4 protein, but not mRNA, levels were elevated by cellular iron restriction. Ferritin turnover by iron deficiency was impaired in NCOA4-depleted cells, presumably due to impaired ferritinophagy. Moreover, HT22 cells became sensitive to iron deficiency by loss of NCOA4. Our studies demonstrate iron can induce ferroptosis in HT22 neuronal cells. We also identify NCOA4-mediated ferritinophagy as an integral process for neuronal cell survival during iron deficiency.
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    Physiological Roles and Regulation of NCOA4 in Macrophages
    (2020-09) Guggisberg, Cole
    Phagocytosing macrophages are critical to systemic iron homeostasis owing to their capability to recycle iron from senescent RBCs and store iron under systemic distress. NCOA4 has recently been identified as a key regulator of ferritin, mediating its degradation via ferritinophagy. Yet, its function in macrophages remains unclear. The present studies employed a cell culture model of J774 macrophages, to examine the role and regulation of macrophage NCOA4 by iron status, red cell iron recycling, and inflammation. Macrophage NCOA4 is responsive to iron status and inversely related to ferritin abundance. By erythrophagocytosis, ferritin peaks at 12 hours with subsequent decrease at 24 hours which is NCOA4-dependent. Hepcidin activity repressed NCOA4 preventing the turnover of ferritin between 12 and 24 hours in erythrocyte laden macrophages. Macrophages were treated with LPS, which decreased both NCOA4 transcript and protein abundance. Altogether our studies demonstrate an active role of NCOA4-mediated ferritinophagy in macrophage iron homeostasis.