RESUMEN
BACKGROUND: Cis-acting elements are essential genomic sequences that control gene expression. In higher eukaryotes, a series of cis-elements function cooperatively. However, further studies are required to examine the co-regulation of multiple cis-elements on a promoter. The aim of this study was to propose a model of cis-element networks that cooperatively regulate gene expression in rice under iron (Fe) deficiency. RESULTS: We developed a novel clustering-free method, microarray-associated motif analyzer (MAMA), to predict novel cis-acting elements based on weighted sequence similarities and gene expression profiles in microarray analyses. Simulation of gene expression was performed using a support vector machine and based on the presence of predicted motifs and motif pairs. The accuracy of simulated gene expression was used to evaluate the quality of prediction and to optimize the parameters used in this method. Based on sequences of Oryza sativa genes upregulated by Fe deficiency, MAMA returned experimentally identified cis-elements responsible for Fe deficiency in O. sativa. When this method was applied to O. sativa subjected to zinc deficiency and Arabidopsis thaliana subjected to salt stress, several novel candidate cis-acting elements that overlap with known cis-acting elements, such as ZDRE, ABRE, and DRE, were identified. After optimization, MAMA accurately simulated more than 87% of gene expression. Predicted motifs strongly co-localized in the upstream regions of regulated genes and sequences around transcription start sites. Furthermore, in many cases, the separation (in bp) between co-localized motifs was conserved, suggesting that predicted motifs and the separation between them were important in the co-regulation of gene expression. CONCLUSIONS: Our results are suggestive of a typical sequence model for Fe deficiency-responsive promoters and some strong candidate cis-elements that function cooperatively with known cis-elements.
RESUMEN
Iron (Fe) deficiency, a worldwide agricultural problem on calcareous soil with low Fe availability, is also a major human nutritional deficit. Plants induce Fe acquisition systems under conditions of low Fe availability. Previously, we reported that an Fe-deficiency-inducible basic helix-loop-helix (bHLH) transcription factor, OsIRO2, is responsible for regulation of the genes involved in Fe homeostasis in rice. Using promoter-GUS transformants, we showed that OsIRO2 is expressed throughout a plant's lifetime in a spatially and temporally similar manner to the genes OsNAS1, OsNAS2 and TOM1, which is involved in Fe absorption and translocation. During germination, OsIRO2 expression was detected in embryos. OsIRO2 expression in vegetative tissues was restricted almost exclusively to vascular bundles of roots and leaves, and to the root exodermis under Fe-sufficient conditions, and expanded to all tissues of roots and leaves in response to Fe deficiency. OsIRO2 expression was also detected in flowers and developing seeds. Plants overexpressing OsIRO2 grew better, and OsIRO2-repressed plants showed poor growth compared to non-transformant rice after germination. OsIRO2 overexpression also resulted in improved tolerance to low Fe availability in calcareous soil. In addition to increased Fe content in shoots, the overexpression plants accumulated higher amounts of Fe in seeds than non-transformants when grown on calcareous soil. These results suggest that OsIRO2 is synchronously expressed with genes involved in Fe homeostasis, and performs a crucial function in regulation not only of Fe uptake from soil but also Fe transport during germination and Fe translocation to grain during seed maturation.
