Multistep phosphorelay (MSP) signaling integrates hormonal and environmental signals to control both plant development and adaptive responses. Type-A RESPONSE REGULATOR (RRA) genes, the downstream members of the MSP cascade and cytokinin primary response genes, are thought to mediate primarily the negative feedback regulation of (cytokinin-induced) MSP signaling. However, transcriptional data also suggest the involvement of RRA genes in stress-related responses. By employing evolutionary conservation with the well-characterized Arabidopsis thaliana RRA genes, we identified five and 38 novel putative RRA genes in Brassica oleracea and Brassica napus, respectively. Our phylogenetic analysis suggests the existence of gene-specific selective pressure, maintaining the homologs of ARR3, ARR6, and ARR16 as singletons during the evolution of Brassicaceae. We categorized RRA genes based on the kinetics of their cytokinin-mediated up-regulation and observed both similarities and specificities in this type of response across Brassicaceae species. Using bioinformatic analysis and experimental data demonstrating the cytokinin and abiotic stress responsiveness of the A. thaliana-derived TCSv2 reporter, we unveil the mechanistic conservation of cytokinin- and stress-mediated up-regulation of RRA genes in B. rapa and B. napus. Notably, we identify partial cytokinin dependency of cold stress-induced RRA transcription, thus further demonstrating the role of cytokinin signaling in crop adaptive responses.

CEITEC Central European Institute of Technology Masaryk University Kamenice 5 A2 625 00 Brno Czech Republic

Department of Biology University of York York UK

Faculty of Natural Sciences Novosibirsk State University Novosibirsk 630090 Russia

Institute of Cytology and Genetics Siberian Branch Russian Academy of Sciences Novosibirsk 630090 Russia

National Centre for Biomolecular Research Faculty of Science Masaryk University Kamenice 5 A2 625 00 Brno Czech Republic

PSI Ltd Drásov 66424 Drásov Czech Republic

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