BackgroundThe phenylpropanoid pathway is responsible for the synthesis of numerous compounds important for plant growth and responses to the environment. In the first committed step of phenylpropanoid biosynthesis, the enzyme phenylalanine ammonia-lyase (PAL) deaminates L-phenylalanine into trans-cinnamic acid that is then converted into p-coumaric acid by cinnamate-4-hydroxylase (C4H). Recent studies showed that the Kelch repeat F-box (KFB) protein family of ubiquitin ligases control phenylpropanoid biosynthesis by promoting the proteolysis of PAL. However, this ubiquitin ligase family, alternatively named Kiss Me Deadly (KMD), was also implicated in cytokinin signaling as it was shown to promote the degradation of type-B ARRs, including the key response activator ARR1. Considering that ubiquitin ligases typically have narrow target specificity, this dual targeting of structurally and functionally unrelated proteins appeared unusual.ResultsHere we show that the KFBs indeed target PAL but not ARR1. Moreover, we show that changes in early phenylpropanoid biosynthesis alter cytokinin sensitivity – as reported earlier - but that the previously documented cytokinin growth response changes are primarily the result of altered auxin signaling. We found that reduced PAL accumulation decreased, whereas the loss of C4H function increased the strength of the auxin response. The combined loss of function of both enzymes led to a decrease in auxin sensitivity, indicating that metabolic events upstream of C4H control auxin sensitivity. This auxin/phenylpropanoid interaction impacts both shoot and root development and revealed an auxin-dependent stimulatory effect of trans-cinnamic acid feeding on leaf expansion and thus biomass accumulation.ConclusionsCollectively, our results show that auxin-regulated plant growth is fine-tuned by early steps in phenylpropanoid biosynthesis and suggest that metabolites accumulating upstream of the C4H step impact the auxin response mechanism.Electronic supplementary materialThe online version of this article (10.1186/s12870-018-1477-0) contains supplementary material, which is available to authorized users.

The recent discovery of SPINDLY (SPY)-catalyzed protein O-fucosylation revealed a novel mechanism for regulating nucleocytoplasmic protein functions in plants. Genetic evidence indicates important roles of SPY in diverse developmental and physiological processes. However, the upstream signal controlling SPY activity and the downstream substrate proteins O-fucosylated by SPY remain largely unknown. Here, we demonstrated that SPY mediates sugar-dependent growth in Arabidopsis (Arabidopsis thaliana). We further identified hundreds of O-fucosylated proteins using lectin affinity chromatography followed by mass spectrometry. All the O-fucosylation events quantified in our proteomic analyses were undetectable or dramatically decreased in the spy mutants, and thus likely catalyzed by SPY. The O-fucosylome includes mostly nuclear and cytosolic proteins. Many O-fucosylated proteins function in essential cellular processes, phytohormone signaling, and developmental programs, consistent with the genetic functions of SPY. The O-fucosylome also includes many proteins modified by O-linked-N-acetylglucosamine (O-GlcNAc) and by phosphorylation downstream of the target of rapamycin (TOR) kinase, revealing the convergence of these nutrient signaling pathways on key regulatory functions such as post-transcriptional/translational regulation and phytohormone responses. Our study identified numerous targets of SPY/O-fucosylation and potential nodes of crosstalk among sugar/nutrient signaling pathways, enabling future dissection of the signaling network that mediates sugar regulation of plant growth and development.

Cytokinins control critical aspects of plant development and environmental responses. Perception of cytokinin ultimately leads to the activation of proteins belonging to the type-B Response Regulator family of cytokinin response activators. In Arabidopsis thaliana, ARR1 is one of the most abundantly expressed type-B Response Regulators.We investigated the link between cytokinin signaling, protein synthesis, plant growth and osmotic stress tolerance.We show that the increased cytokinin signaling in ARR1 gain-of-function transgenic lines is associated with increased rates of protein synthesis, which lead to growth inhibition and hypersensitivity to osmotic stress. Cytokinin-induced growth inhibition and osmotic stress hypersensitivity were rescued by treatments with ABA, a hormone known to inhibit protein synthesis. We also demonstrate that cytokinin-induced protein synthesis requires isoforms of the ribosomal protein L4 encoded by the cytokinin-inducible genes RPL4A and RPL4D, and that RPL4 loss-of-function increases osmotic stress tolerance and decreases sensitivity to cytokinin-induced growth inhibition.These findings reveal that an increase in protein synthesis negatively impacts growth and osmotic stress tolerance and explain some of the adverse effects of elevated cytokinin action on plant development and stress physiology.

Proteins are workhorses in the cell; they form stable and more often dynamic transient protein–protein interactions, assemblies, and networks and have an intimate interplay with DNA and RNA. These network interactions underlie fundamental biological processes and play essential roles in cellular function. The proximity-dependent biotinylation labeling approach combined with mass spectrometry (PL-MS) has recently emerged as a powerful technique to dissect the complex cellular network at the molecular level. In PL-MS, by fusing a genetically encoded proximity-labeling (PL) enzyme to a protein or a localization signal peptide, the enzyme is targeted to a protein complex of interest or to an organelle, allowing labeling of proximity proteins within a zoom radius. These biotinylated proteins can then be captured by streptavidin beads and identified and quantified by mass spectrometry. Recently engineered PL enzymes such as TurboID have a much-improved enzymatic activity, enabling spatiotemporal mapping with a dramatically increased signal-to-noise ratio. PL-MS has revolutionized the way we perform proteomics by overcoming several hurdles imposed by traditional technology, such as biochemical fractionation and affinity purification mass spectrometry. In this review, we focus on biotin ligase–based PL-MS applications that have been or are likely to be adopted by the plant field. We discuss the experimental designs and review the different choices for engineered biotin ligases, enrichment, and quantification strategies. Lastly, we review the validation and discuss future perspectives. Expected final online publication date for the Annual Review of Plant Biology, Volume 74 is May 2023. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.