Identification of iron-deficiency responsive microRNA genes and cis-elements in Arabidopsis

MicroRNAs (miRNAs) are a class of endogenous non-coding small RNAs that bind to their target mRNAs to repress their translation or induce their degradation. Recent studies have shown that several miRNAs regulate plant adaptation to sulfate and phosphate deficiency. However, whether miRNAs are involved in regulation of stress response to iron (Fe) deficiency is unknown. In this study, we carried out a survey of Arabidopsis miRNA genes in response to Fe deficiency and identified IDE1/IDE2 (Iron-deficiency responsive cis-Element 1 and 2) in their promoter regions. We constructed a small RNA library from Arabidopsis seedlings under Fe deficiency. Sequence analysis revealed 8 conserved miRNA genes in 5 families, all of which were up-regulated during Fe deficiency. Further, we analyzed cis-regulatory elements upstream of all miRNA genes in Arabidopsis and found 24 miRNA genes containing IDE1/IDE2 motifs in their promoter regions. Transcriptional analysis using RT-PCR showed that 70.8% (17/24) of the IDE-containing miRNA genes were expressed in response to Fe deficiency. We presented a putative interaction model between protein-coding genes and miRNA genes under Fe deficiency. Our analytic approach is useful and efficient because it is applicable to cis-element finding for miRNAs responding to other abiotic stresses. Also, the data obtained in this study may aid our understanding of the role of Fe deficiency responsive specific sequences upstream of miRNA genes and the functional implications of miRNA genes in response to Fe stress in plants.