Cellular iron depletion and the mechanisms involved in the iron-dependent regulation of the growth arrest and DNA damage family of genes

Iron plays a crucial part in proliferation while iron deficiency results in G1/S arrest, DNA damage, and apoptosis. However, the precise role of iron in cell cycle control remains unclear. We showed that iron depletion using the iron chelators, desferrioxamine (DFO), or 2-hydroxy-1-napthylaldehyde isonicotinoyl hydrazone (311), increased the mRNA levels of the growth arrest and DNA damage 45α gene, GADD45α (Darnell, G. and Richardson, D. R. (1999) Blood 94, 781-792). In this study, we examined the effect of iron depletion on up-regulating GADD family members involved in growth control, including cell cycle arrest, apoptosis, andDNArepair, making them therapeutic targets for tumor suppression.Weshowed the GADD family members were up-regulated by cellular iron depletion. Further, upregulation of GADD45α after iron deprivation was independent of hypoxia-inducible factor-1α (HIF-1α), octamer-1 (Oct-1), p53 and early growth response 1 (Egr1). We then analyzed the regulatory elements responsible for iron depletion-mediated regulation of GADD45α and identified the specific transcription factor/s involved. This region was within -117 bp and -81 bp relative to the start codon where the consensus sequences of three transcription factors are located: the CCAAT-binding factor/nuclear factor-Y (NF-Y), the stabilizing molecule v-MYB and the enhancer, CCAAT enhancer-binding protein (CEBPα). Mutation analysis, shRNA studies, Western blotting, and electrophoretic mobility shift assays led to the identification of NF-Y in the transcriptional up-regulation of GADD45α after iron depletion. Furthermore, like GADD45α, NF-YA was upregulated after iron chelation and down-regulated by iron supplementation. These results are important for understanding the mechanisms of iron depletion-mediated cell cycle arrest, DNA damage repair, and apoptosis.