Synthetic deconvolution of an auxin-dependent transcriptional code

How developmental signals program gene expression in space and time is still poorly understood. Here, we addressed this question for the plant master regulator, auxin. Transcriptional responses to auxin rely on a large multigenic transcription factor family, the auxin response factors (ARFs). We deconvoluted the complexity of ARF-regulated transcription using auxin-inducible synthetic promoters built from cis-element pair configurations differentially bound by ARFs. We demonstrate using cellular systems that ARF transcriptional properties are not only intrinsic but also depend on the cis-element pair configurations they bind to, thus identifying a bi-layer ARF/cis-element transcriptional code. Auxin-inducible synthetic promoters were expressed differentially in planta showing at single-cell resolution how this bi-layer code patterns transcriptional responses to auxin. Combining cis-element pair configurations in synthetic promoters created distinct patterns, demonstrating the combinatorial power of the auxin bi-layer code in generating diverse gene expression patterns that are not simply a direct translation of auxin distribution. Overall design: Single cell RNAseq approach on Arabidopsis thaliana, 7day old seedling, cultivated in vitro, MS/2, sucrose 1%, Agar 1%, and treated with IAA 1 µM for 16 h or Mock. Experiment was performed on 4 lines carrying the DR5, ER13, DR15 or IR8 synthetic promoters followed by a mTurquoise reporter.