Auxin represents a key signal in plants, regulating almost every aspect of their growth and development. Major breakthroughs have been made dissecting the molecular basis of auxin transport, perception, and response. In contrast, how plants control the metabolism and homeostasis of the major form of auxin in plants, indole-3-acetic acid (IAA), remains unclear. In this paper, we initially describe the function of the Arabidopsis thaliana gene DIOXYGENASE FOR AUXIN OXIDATION 1 (AtDAO1). Transcriptional and translational reporter lines revealed that AtDAO1 encodes a highly root-expressed, cytoplasmically localized IAA oxidase. Stable isotope-labeled IAA feeding studies of loss and gain of function AtDAO1 lines showed that this oxidase represents the major regulator of auxin degradation to 2-oxoindole-3-acetic acid (oxIAA) in Arabidopsis Surprisingly, AtDAO1 loss and gain of function lines exhibited relatively subtle auxin-related phenotypes, such as altered root hair length. Metabolite profiling of mutant lines revealed that disrupting AtDAO1 regulation resulted in major changes in steady-state levels of oxIAA and IAA conjugates but not IAA. Hence, IAA conjugation and catabolism seem to regulate auxin levels in Arabidopsis in a highly redundant manner. We observed that transcripts of AtDOA1 IAA oxidase and GH3 IAA-conjugating enzymes are auxin-inducible, providing a molecular basis for their observed functional redundancy. We conclude that the AtDAO1 gene plays a key role regulating auxin homeostasis in Arabidopsis, acting in concert with GH3 genes, to maintain auxin concentration at optimal levels for plant growth and development.

Centre for Plant Integrative Biology Plant and Crop Science Division School of Biosciences University of Nottingham Loughborough LE12 5RD United Kingdom

Department of Agronomy and Horticulture University of Nebraska Lincoln NE 68583 0915

Department of Plant Biology and Crop Science Rothamsted Research Hertfordshire AL5 2JQ United Kingdom

Faculty of Science Palacký University CZ 78371 Olomouc Czech Republic

Integrated Molecular Plant Physiology Research Biology Department Antwerp University 2020 Antwerp Belgium

Laboratory of Growth Regulators Centre of the Region Haná for Biotechnological and Agricultural Research Institute of Experimental Botany Academy of Sciences of the Czech Republic CZ 78371 Olomouc Czech Republic

Umeå Plant Science Centre Department of Forest Genetics and Plant Physiology Swedish University of Agricultural Sciences SE 901 83 Umea Sweden

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