Regulation of mRNA abundance revealed a genetic program for plant leaf acclimation to iron (Fe) limitation. The transcript for SUFB, a key component of the plastid iron-sulfur (Fe-S) assembly pathway is down-regulated early after Fe deficiency, and prior to down-regulation of mRNAs encoding abundant chloroplast Fe containing proteins, which should economize the use of Fe. What controls this system is unclear. We utilized RNA-seq. aimed to identify differentially expressed transcripts that are co-regulated with SUFB after Fe deficiency in leaves. To distinguish if lack of Fe or lack of Fe-S cofactors and associated loss of enzymatic and photosynthetic activity trigger transcriptome reprogramming, WT plants on low Fe were compared with an inducible sufb-RNAi knockdown. Fe deficiency targeted a limited set of genes and predominantly affected transcripts for chloroplast localized proteins. A set of glutaredoxin transcripts was concertedly down-regulated early after Fe deficiency, however when these same genes were down-regulated by RNAi the effect on known chloroplast Fe deficiency marker proteins was minimal. In promoters of differentially expressed genes, binding motifs for AP2/ERF transcription factors were most abundant and three AP2/ERF transcription factors were also differentially expressed early after low Fe treatment. Surprisingly, Fe deficiency in a WT on low Fe and a sufb-RNAi knockdown presented very little overlap in differentially expressed genes. sufb-RNAi produced expression patterns expected for Fe excess and up-regulation of a transcript for another Fe-S assembly component not affected by low Fe. These findings indicate that Fe scarcity, not Fe utilization, triggers reprogramming of the transcriptome in leaves.