Higher plants maintain iron homeostasis by regulating the expression of iron (Fe)-related genes in accordance with Fe availability. The transcription factor IDEF1 regulates the response to Fe deficiency in Oryza sativa (rice) by recognizing CATGC sequences within the Fe deficiency-responsive cis-acting element IDE1. To investigate the function of IDEF1 in detail, we analyzed the response to Fe deficiency in transgenic rice plants exhibiting induced or repressed IDEF1 expression. Fe-deficiency treatment in hydroponic culture revealed that IDEF1 knock-down plants are susceptible to early-stage Fe deficiency, in contrast to IDEF1-induced plants. Time-course expression analyses using quantitative reverse-transcriptase PCR revealed that the IDEF1 expression level was positively correlated with the level of induction of the Fe utilization-related genes OsIRO2, OsYSL15, OsIRT1, OsYSL2, OsNAS1, OsNAS2, OsNAS3 and OsDMAS1, just after the onset of Fe starvation. However, this overall transactivation mediated by IDEF1 became less evident in subsequent stages. Microarray and in-silico analyses revealed that genes positively regulated by IDEF1, especially at the early stage, exhibit over-representation of CATGC and IDE1-like elements within the proximal promoter regions. These results indicate the existence of early and subsequent responses to Fe deficiency, with the former requiring IDEF1 more specifically. Proximal regions of IDEF1-regulated gene promoters also showed enrichment of RY elements (CATGCA), which regulate gene expression during seed maturation. The expression of several genes encoding late embryogenesis abundant proteins, including Osem, was induced in Fe-deficient roots and/or leaves in an IDEF1-dependent manner, suggesting a possible function of seed maturation-related genes in Fe-deficient vegetative organs.