Aux/lAA Proteins Repress Expression of Reporter Genes Containing Natural and Highly Active Synthetic Auxin Response Elements

A highly active synthetic auxin response element (AuxRE), referred to as DR5, was created by petforming site-directed mutations in a natural composite AuxRE found in the soybean GH3 promoter. DR5 consisted of tandem direct repeats of 11 bp that included the auxin-responsive TGTCTC element. The DR5 AuxRE showed greater auxin responsiveness than a natural composite AuxRE and the GH3 promoter when assayed by transient expression in carrot protoplasts or in stably transformed Arabidopsis seedlings, and it provides a useful reporter gene for studying auxin-responsive transcription in wild-type plants and mutants. An auxin response transcription factor, ARFl, bound with specificity to the DR5 AuxRE in vitro and interacted with Aux/lAA proteins in a yeast two-hybrid system. Cotransfection experiments with natural and synthetic AuxRE reporter genes and effector genes encoding Aux/lAA proteins showed that overexpression of Aux/lAA proteins in carrot protoplasts resulted in specific repression of TGTCTC AuxRE reporter gene expression.