Mechanism of auxin-dependent gene regulation through composite auxin response elements

The plant signaling molecule auxin controls growth and development, largely through activating and repressing the expression of thousands of genes. Auxin-dependent transcriptional changes are mediated by DNA-binding Auxin Response Factors (ARF), whose AuxRE DNA binding sites are well-known. The identification of the first AuxRE showed this to be part of a composite element with a second motif. Indeed, systematic analysis showed other DNA motifs to be enriched in auxin-regulated promoters. Neither the basis for this enrichment nor the mechanisms for the activity of composite AuxREs is known. Here, we systematically mined Arabidopsis promoters for composite AuxRE elements enriched in auxin-responsive genes. We identified many and show that their presence is a reliable predictor of auxin response. Through mutating these elements and their higher-order modules, we demonstrate function in promoter activity. Lastly, we identified transcription factors (TFs) that bind AuxRE-associated motifs, showed their involvement in auxin response, and discovered that several of these TFs directly bind ARF proteins. We propose that ARF-TF complexes specifically bind compound motifs in promoters, and act as a source of diversification in auxin-dependent gene regulation.