| Summary: | <p>Iron is an essential micronutrient as it contributes to the oxygen-carrying capacity of blood and is a cofactor for many enzymes. However, excess iron is toxic as it promotes the formation of reactive oxygen species (ROS), which react with and alter the functions of biomolecules. Moreover, there is no regulated iron excretion mechanism from our body. Hence, iron homeostasis is tightly regulated by the hormone hepcidin that restricts iron absorption and controls iron distribution in the body. Accumulation of iron in the liver increases the expression of bone morphogenetic protein-6 (<em>Bmp6</em>), which induces hepcidin synthesis via Bmp responsive elements in the hepcidin promoter. The mechanism by which excess iron is ‘sensed’, leading to increased <em>Bmp6</em>, is unknown. Since excess iron induces oxidative stress, we hypothesized that <em>Bmp6</em> expression is upregulated during iron-overload via the Nrf2-driven antioxidant response pathway. Nrf2 is a transcriptional activator that is activated by oxidative stimuli and binds to antioxidant responsive elements (AREs) to induce the expression of a battery of ARE-regulated antioxidant genes. Bach1 is a transcriptional repressor that binds AREs and suppresses gene expression. Haem can induce the degradation of Bach1 and derepress the expression of ARE-regulated genes. In this thesis, we demonstrated the simultaneous induction of <em>Bmp6</em> and classical Nrf2/ARE-driven genes in mice and cell lines by iron and hemin (haem-chloride). ChIP-sequencing analyses showed binding of Nrf2 to a conserved ARE within intron 1 of <em>Bmp6</em>. The antioxidant mitoTEMPO blunted the activation of Nrf2 and upregulation of <em>Bmp6</em> by iron. Furthermore, siRNA-mediated knockdown of <em>Nrf2</em> decreased basal <em>Bmp6</em> expression and inhibited the upregulation of <em>Bmp6</em> by iron <em>in vitro</em>, whereas knockdown of Bach1 increased <em>Bmp6</em> expression. Similarly, the upregulation of <em>Bmp6</em> and hepcidin expression was blunted or completely abrogated in several models of iron overloading in <em>Nrf2</em>-knockout mice. <em>Nrf2</em>-knockout mice were more prone to iron accumulation and susceptible to oxidative stress-induced liver damage. Deletion of <em>Nrf2</em> in <em>Hfe</em>-knockout haemochromatosis mice, and a SNP associated with reduced NRF2 expression in HFE-hereditary haemochromatosis patients, worsened the iron accumulation phenotype; conversely, pharmacological activation of Nrf2 upregulated the <em>Bmp6</em>/hepcidin axis and alleviated iron accumulation and oxidative stress in <em>Hfe</em>-knockout mice. In summary, Nrf2 links cellular and systemic iron homeostasis, is required for the upregulation of <em>Bmp6</em> by iron, and is an important modifier and therapeutic target for iron overload disorders.</p>
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