WRINKLED1 Is Subject to Evolutionary Conserved Negative Autoregulation.

High accumulation of storage compounds such as oil and starch are economically important traits of most agricultural crops. The genetic network determining storage compounds composition in crops has been the target of many biotechnological endeavors. Especially WRINKLED1 (WRI1), a well-known key transcription factor involved in the allocation of carbon into oil, has attracted much interest. Here we investigate the presence of an autoregulatory system involving WRI1 through transient expression in leaves. Different lengths of the Arabidopsis promotor region were coupled to a GUS reporter gene and the activity was measured when combined with constitutive expression of different WRI1 homologs from , oat ( L.), yellow nutsedge L.), and potato ( L.). We could show that increasing levels of each WRI1 homolog reduced the transcriptional activity of the Arabidopsis upstream region. Through structural analysis and domain swapping between oat and Arabidopsis WRI1, we were able to determine that the negative autoregulation was clearly dependent on the DNA-binding AP2-domains. A DNA/protein interaction assay showed that AtWRI1 is unable to bind to its corresponding upstream region indicating non-direct interaction . Taken together, our results demonstrate a negative feedback loop of expression and that it is an indirect interaction most likely caused by downstream targets of WRI1. We also show that it is possible to release expression from this autoregulation by creating semi-synthetic WRI1 homologs increasing the potential use of WRI1 in biotechnological applications.