Modulation of cellular iron metabolism by hydrogen peroxide -: Effects of H2O2 on the expression and function of iron-responsive element-containing mRNAs in B6 fibroblasts

Cellular iron uptake and storage are coordinately controlled by binding of iron-regulatory proteins (IRP), IRP1 and IRP2, to iron-responsive elements (IREs) within the mRNAs encoding transferrin receptor (TfR) and ferritin. Under conditions of iron starvation, both IRP1 and IRP2 bind with high affinity to cognate IREs, thus stabilizing TfR and inhibiting translation of ferritin mRNAs, The IRE/IRP regulatory system receives additional input by oxidative stress in the form of H2O2 that leads to rapid activation of IRP1. Here we show that treating murine B6 fibroblasts with a pulse of 100 muM H2O2 for 1 h is sufficient to alter critical parameters of iron homeostasis in a time-dependent manner. First, this stimulus inhibits ferritin synthesis for at least 8 h, leading to a significant (50%) reduction of cellular ferritin content. Second, treatment with H2O2 induces a similar to4-fold increase in TfR mRNA levels within 2-6 h, and subsequent accumulation of newly synthesized protein after 4 h, This is associated with a profound increase in the cell surface expression of TfR, enhanced binding to fluorescein-tagged transferrin, and stimulation of transferrin-mediated iron uptake into cells, Under these conditions, no significant alterations are observed in the levels of mitochondrial aconitase and the (D) under bar ivalent (M) under bar etal (T) under bar ransporter DMT1, although both are encoded by two as yet lesser characterized IRE-containing mRNAs, Finally. H2O2-treated cells display an increased capacity to sequester Fe-59 in ferritin, despite a reduction in the ferritin pool, which results in a rearrangement of Fe-59 intracellular distribution. Our data suggest that H2O2 regulates cellular iron acquisition and intracellular iron distribution by both IRP1-dependent and -independent mechanisms.