A non-DNA-binding activity for the ATHB4 transcription factor in the control of vegetation proximity

In plants, perception of vegetation proximity by phytochrome photoreceptors activates a transcriptional network that implements a set of responses to adapt to plant competition, including elongation of stems or hypocotyls. In Arabidopsis thaliana, the homeodomain-leucine zipper (HD-Zip) transcription factor ARABIDOPSIS THALIANA HOMEOBOX 4 (ATHB4) regulates this and other responses, such as leaf polarity. To better understand the shade regulatory transcriptional network, we have carried out structure–function analyses of ATHB4 by overexpressing a series of truncated and mutated forms and analyzing three different responses: hypocotyl response to shade, transcriptional activity and leaf polarity. Our results indicated that ATHB4 has two physically separated molecular activities: that performed by HD-Zip, which is involved in binding to DNA-regulatory elements, and that performed by the ETHYLENE-RESPONSIVE ELEMENT BINDING FACTOR-associated amphiphilic repression (EAR)-containing N-terminal region, which is involved in protein−protein interaction. Whereas both activities are required to regulate leaf polarity, DNA-binding activity is not required for the regulation of the seedling responses to plant proximity, which indicates that ATHB4 works as a transcriptional cofactor in the regulation of this response. These findings suggest that transcription factors might employ alternative mechanisms of action to regulate different developmental processes.M. Gallemi and M.J.M-C. received predoctoral FPI fellowships from the Spanish Ministry of Economy and Competivity (MINECO). S.P. received a predoctoral fellowship from Catalan Agència de Gestió d'Ajuts Universitaris i de Recerca (AGAUR, FI-DGR 2015). M.S-M. received a predoctoral fellowship from the CSIC (JAEpre program). C.S. received financial support from the AGAUR (C-RED Program). Our research is supported by grants from the MINECO-FEDER (BIO2011-23489 and BIO2014-59895-P) and AGAUR (2014-SGR447 and Xarba) to J.F.M-G. We also acknowledge the support of the MINECO for the ‘Centro de Excelencia Severo Ochoa 2016–2019’ award SEV-2015-0533 and the Generalitat de Catalunya through the CERCA Program.Peer reviewe