Characterization and expression analysis of the Arabidopsis mir169 family

The mir169 family is a large and conserved miRNA family in many plant species. The Arabidopsis genome contains 14 members of the mir169 family, including three tandem clusters that were transcribed as polycistrons. To investigate the regulation of mir169 expression, all the 11 promoter sequences were cloned and fused to GUS and introduced into Arabidopsis to generate pMir169:GUS transgenic plants. Histochemical GUS staining and activity assay in different tissues and developmental stages of the transgenic plants revealed unique and overlapping as well as spatio-temporal expression patterns among the family members. The endogenous transcript level of each member detected by semi-quantitative RT-PCR is largely accordant with the GUS expression profiles. Interestingly, the tandem arrayed gene pairs mir169i/j, mir169k/l and mir169m/n show greatly overlapping expression patterns. Further comparative analysis of their upstream sequences revealed significant sequence similarities, indicating that the three gene pairs may result from a duplication event. Additionally, when the transgenic plants were exposed to various abiotic stresses, a subset of family members were observed to be regulated by certain treatments, especially ABA and heat. Our results provide valuable information for functional analysis of the Arabidopsis mir169 family members as ubiquitous regulators in both developmental processes and stimuli pathways.