During plant development, a precise balance of cytokinin is crucial for correct growth and patterning, but it remains unclear how this is achieved across different cell types and in the context of a growing organ. Here we show that in the root apical meristem, the TMO5/LHW complex increases active cytokinin levels via two cooperatively acting enzymes. By profiling the transcriptomic changes of increased cytokinin at single-cell level, we further show that this effect is counteracted by a tissue-specific increase in CYTOKININ OXIDASE 3 expression via direct activation of the mobile transcription factor SHORTROOT. In summary, we show that within the root meristem, xylem cells act as a local organizer of vascular development by non-autonomously regulating cytokinin levels in neighbouring procambium cells via sequential induction and repression modules.

Department of Applied Mathematics Computer Science and Statistics Ghent University Ghent Belgium

Department of Biomedical Molecular Biology Ghent University Ghent Belgium

Department of Plant Biotechnology and Bioinformatics Ghent University Ghent Belgium

Laboratory of Growth Regulators Institute of Experimental Botany of the Czech Academy of Sciences and Faculty of Science of Palacký University Olomouc Czech Republic

Laboratory of Hormonal Regulations in Plants Institute of Experimental Botany Czech Academy of Sciences Prague Czech Republic

School of Biosciences University of Nottingham Loughborough UK

School of Mathematical Sciences University of Nottingham Nottingham UK

VIB Center for Inflammation Research Data Mining and Modelling for Biomedicine Ghent Belgium

VIB Center for Plant Systems Biology Ghent Belgium

VIB Flow Core VIB Center for Inflammation Research Ghent Belgium

VIB Protein Service Facility VIB Center for Inflammation Research Ghent Belgium

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