Auxin determines the developmental fate of plant tissues, and local auxin concentration is precisely controlled. The role of auxin transport in modulating local auxin concentration has been widely studied but the regulation of local auxin biosynthesis is less well understood. Here, we show that TRYPTOPHAN AMINOTRANSFERASE OF ARABIDOPSIS (TAA1), a key enzyme in the auxin biosynthesis pathway in Arabidopsis thaliana is phosphorylated at Threonine 101 (T101). T101 phosphorylation status can act as an on/off switch to control TAA1-dependent auxin biosynthesis and is required for proper regulation of root meristem size and root hair development. This phosphosite is evolutionarily conserved suggesting post-translational regulation of auxin biosynthesis may be a general phenomenon. In addition, we show that auxin itself, in part via TRANS-MEMBRANE KINASE 4 (TMK4), can induce T101 phosphorylation of TAA1 suggesting a self-regulatory loop whereby local auxin signalling can suppress biosynthesis. We conclude that phosphorylation-dependent control of TAA1 enzymatic activity may contribute to regulation of auxin concentration in response to endogenous and/or external cues.

A bamboo shoot is the immature stem of the woody grass and a nutritious and popular vegetable in East Asia. However, it undergoes a rapid xylogenesis process right after harvest, even being stored in a cold chamber. To investigate the molecular regulation mechanisms of xylogenesis in Moso bamboo (Phyllostachys edulis) shoots (MBSes) during cold storage, the measurement of cell wall polymers (cellulose, hemicellulose, and lignin) and related enzyme activities (phenylalanine ammonia lyase (PAL), cinnamyl alcohol dehydrogenase (CAD), peroxidase (POD), and xylan xylosyltransferase (XylT)) and transcriptomic analysis were performed during cold storage. It was noticed that cellulose and lignin contents increased, while hemicellulose content exhibited a downward trend. PAL, CAD, and POD activity presented an upward trend generally in MBS when stored at 4 °C for 16 days. XylT activity showed a descending trend during the stages of storage, but slightly increased during the 8th to 12th days after harvest at 4 °C. Transcriptomic analysis identified 72, 28, 44, and 31 functional unigenes encoding lignin, cellulose, xylan biosynthesis enzymes, and transcription factors (TFs), respectively. Many of these secondary cell wall (SCW)-related genes showed higher expression levels in the later period of cold storage. Quantitative RT-PCR analysis of the selected genes conformed to the expression pattern. Our study provides a comprehensive analysis of MBS secondary wall biosynthesis at the molecular level during the cold storage process. The results give insight into the xylogenesis process of this economically important vegetable and shed light on solving this problem of the post-harvest industry.

Understanding plant molecular responses to flooding is crucial for strategies to increase resilience. Plants respond to submergence-induced low oxygen (hypoxia) through decreased plant cysteine oxidase (PCO) activity, which stabilizes group VII ethylene response factors (ERFVIIs), master regulators of metabolic and anatomic acclimation responses ^1–4 . Rapid reoxygenation on desubmergence induces a burst of reactive oxygen species (ROS) generation and metabolic reconfiguration ^5,6 ; however, how plants mitigate this post-hypoxic stress to facilitate submergence recovery has remained unknown. Here we report that ERFVIIs are also important in post-submergence recovery, remaining stable upon reoxygenation through ROS-mediated PCO inhibition. Stabilized ERFVIIs are retained at hypoxia-responsive promoters, becoming repressors of typical hypoxia marker genes but upregulators of genes involved in ROS homeostasis and oxidative stress protection. Our findings suggest that PCOs and ERFVIIs integrate signals from both oxygen and ROS to coordinate ERFVII stability through submergence-induced hypoxia and desubmergence stress to promote plant survival and recovery.

Plants face hypoxic conditions either chronically, as particular tissues are characterized by fluctuating or stable low oxygen levels, or acutely, when flooded. In vascular plants, transcriptional adaptive responses to hypoxia are rapidly mounted by Ethylene Response Factors VII (ERFVIIs), regulated by Plant Cysteine Oxidases (PCOs) through the cysteine branch of the N-degron pathway (Cys-NDP) for oxygen sensing. However, this relatively simple regulatory circuit, consisting of both constitutively expressed as well as hypoxia-inducible ERFVIIs and PCOs, interacts with diverse signalling cues and pathways invoked by hypoxia. To understand the share of the PCO-mediated oxygen sensing mechanism in the production of hypoxia responses, we insulated the PCO/ERFVII circuit from Arabidopsis thaliana and adapted it to Saccharomyces cerevisiae . Using a reporter gene to monitor the output of the circuit allowed us to compare the speed and amplitude of response to hypoxia in the engineered yeast and the source organism. Hypoxia triggered ERFVII stabilization both in Arabidopsis and yeast, leading to a similarly fast transcriptional response that was however larger in plants. A simple hypoxia-inducible feedback loop improved the amplitude of response in yeast, demonstrating the importance of this regulation in the endogenous PCO/ERFVII circuit. Finally, computational modelling of the yeast circuit enabled us to identify promoter competition and presence of hypoxia-inducible PCOs as key parameters that shape early hypoxia responses in plant cells. Significance Statement: We report the design, testing and optimisation of a synthetic molecular switch that activates gene expression in response to hypoxia in the yeast Saccharomyces cerevisiae . This is based on enzymes that consume molecular oxygen to regulate the stability of transcription factors in plant cells. By generating such a hybrid molecular device, we were able to demonstrate the efficacy of this hypoxia response strategy independently of the many ancillary components that affect gene regulation in plant cells. In this way, we were able to assess its activation dynamics, characterised by similarly fast induction of gene expression in both yeast and plants. Our approach also revealed the requirement of interlocked feedback loops to achieve the magnitude of gene induction measured in plants.

Plants face hypoxic conditions either chronically, as particular tissues are characterized by fluctuating or stable low oxygen levels, or acutely, when flooded. In vascular plants, transcriptional adaptive responses to hypoxia are rapidly mounted by Ethylene Response Factors VII (ERFVIIs), regulated by Plant Cysteine Oxidases (PCOs) through the cysteine branch of the N-degron pathway (Cys-NDP) for oxygen sensing. However, this relatively simple regulatory circuit, consisting of both constitutively expressed as well as hypoxia-inducible ERFVIIs and PCOs, interacts with diverse signaling cues and pathways invoked by hypoxia. To understand the share of the PCO-mediated oxygen sensing mechanism in the production of hypoxia responses, we insulated the PCO/ERFVII circuit from Arabidopsis thaliana and adapted it to Saccharomyces cerevisiae . Using a reporter gene to monitor the output of the circuit allowed us to compare the speed and amplitude of response to hypoxia in the engineered yeast and the source organism. Hypoxia triggered ERFVII stabilization both in Arabidopsis and yeast, leading to a similarly fast transcriptional response that was however larger in plants. A simple hypoxia-inducible feedback loop improved the amplitude of response in yeast, demonstrating the importance of this regulation in the endogenous PCO/ERFVII circuit. Finally, computational modeling of the yeast circuit enabled us to identify promoter competition and presence of hypoxia-inducible PCOs as key parameters that shape early hypoxia responses in plant cells.