Specificity in auxin responses is not explained by the promoter preferences of activator ARFs

Auxin is essential for almost every developmental process within plants. How a single small molecule can lead to a plethora of downstream responses has puzzled researchers for decades. It has been hypothesized that one source for such diversity is distinct promoter-binding and activation preferences for different members of the AUXIN RESPONSE FACTOR (ARF) family of transcription factors. We systematically tested this hypothesis by engineering varied promoter sequences in a heterologous yeast system and quantifying transcriptional activation by ARFs from two species, Arabidopsis thaliana and Zea mays. By harnessing the user-defined and scalable nature of our synthetic system, we elucidated promoter design rules for optimal ARF function, discovered novel ARF-responsive promoters, and characterized the impact of ARF dimerization on their activation potential. We found no evidence for specificity in ARF-promoter interactions, suggesting that the diverse auxin responses observed in plants may be driven by factors outside the core auxin response machinery.