Iron (Fe) is a key element of many proteins involved in primary cellular processes such as energy production, DNA synthesis and repair (Fuss et al. 2015). Unbalanced iron levels are poorly tolerated thus a tight control over iron trafficking and storage is necessary in order to ensure cell growth and genome stability (Zhang et al. 2014). Indeed, under low iron condition, cells arrest in G1 phase and restrain DNA synthesis till normal iron levels are restored (Lui et al 2015). The Nuclear Receptor CoActivator 4 (NCOA4) has been recently involved in cellular and systemic iron homeostasis as a selective cargo receptor of ferritin, the iron storage macromolecule (Mancias et al. 2014). Under iron deficient condition, NCOA4 promotes the autophagic degradation of ferritin (ferritinophagy) restoring appropriate iron levels. Conversely, excess iron induces the proteosomal degradation of NCOA4 by HERC2 ubiquitin ligase allowing iron storage into ferritin (Mancias et al. 2015). Lack of ferritinophagy in NCOA4-deficient mice leads to tissue iron overload as well as anaemia because of inability to mobilize iron from deposits (Bellelli et al. 2016). In addition to this iron-related function, we uncovered that nuclear NCOA4 binds to MCM7, a member of the core engine of the replicative DNA Helicase and blocks DNA replication origin activation (Bellelli et al. 2014). Since low iron levels impair cell proliferation and block DNA replication, we hypothesized that cytosolic and nuclear NCOA4 functions (eg ferritinophagy and DNA replication origin control, respectively) were linked in order to couple iron availability to DNA metabolism. We first evaluated the effects of iron chelation on NCOA4 levels and found that Deferoxamine (DFO) treatment of HeLa cell lines increased both cytosolic and nuclear NCOA4 fraction. In particular, under low iron conditions, we found that NCOA4 not only induced ferritin degradation but also increased its interaction with MCM7 at canonical DNA origins in order to block DNA replication. Consistently, upon iron chelation, NCOA4-/- HeLa cells failed to block DNA synthesis, and underwent replication stress with a robust activation of the DNA damage response (DDR) and a permanent cell cycle arrest, as confirmed by low colony-forming ability of NCOA4 deficient cells. Then, we asked whether the dual function of NCOA4 was relevant during DNA Damage, when replication is blocked and many iron-requiring enzymes work to repair DNA. Interestingly, although DNA damage did not substantially reduce labile iron pool, as suggested by no change of IRP2 stability, treatment with genotoxic agents, such as Hydroxyurea (HU), decreased NCOA4 binding to HERC2. The HU-dependent NCOA4 stabilization promoted block of DNA replication origin activation and, as iron depletion, increased ferritin turnover. Collectively, our data identify NCOA4 functions as two faces of a common response, which aims at coordinating cellular iron availability to DNA metabolism. In this light, NCOA4 by controlling activation of DNA origins reduces replication stress and by regulating cellular iron levels sustains DNA iron-requiring enzymatic activities in order to maintain genome stability.
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