Indole-3-acetic acid (IAA), the natural auxin in plants, regulates many aspects of plant growth and development. Extensive analyses have elucidated the components of auxin biosynthesis, transport, and signaling, but the physiological roles and molecular mechanisms of auxin degradation remain elusive. Here, we demonstrate that the dioxygenase for auxin oxidation (DAO) gene, encoding a putative 2-oxoglutarate-dependent-Fe (II) dioxygenase, is essential for anther dehiscence, pollen fertility, and seed initiation in rice. Rice mutant lines lacking a functional DAO display increased levels of free IAA in anthers and ovaries. Furthermore, exogenous application of IAA or overexpression of the auxin biosynthesis gene OsYUCCA1 phenocopies the dao mutants. We show that recombinant DAO converts the active IAA into biologically inactive 2-oxoindole-3-acetic acid (OxIAA) in vitro. Collectively, these data support a key role of DAO in auxin catabolism and maintenance of auxin homeostasis central to plant reproductive development.

4-Coumaric acid:CoA ligase (4CL) is the central enzyme of the plant-specific phenylpropanoid pathway. It catalyzes the synthesis of hydroxycinnamate-CoA thioesters, the precursors of lignin and other important phenylpropanoids, in two-step reactions involving the formation of hydroxycinnamate-AMP anhydride and then the nucleophilic substitution of AMP by CoA. In this study, we determined the crystal structures of Populus tomentosa 4CL1 in the unmodified (apo) form and in forms complexed with AMP and adenosine 59-(3-(4-hydroxyphenyl)propyl)phosphate (APP), an intermediate analog, at 2.4, 2.5, and 1.9 Å resolution, respectively. 4CL1 consists of two globular domains connected by a flexible linker region. The larger N-domain contains a substrate binding pocket, while the C-domain contains catalytic residues. Upon binding of APP, the C-domain rotates 818 relative to the N-domain. The crystal structure of 4CL1-APP reveals its substrate binding pocket. We identified residues essential for catalytic activities (Lys-438, Gln-443, and Lys-523) and substrate binding (Tyr-236, Gly-306, Gly-331, Pro-337, and Val-338) based on their crystal structures and by means of mutagenesis and enzymatic activity studies. We also demonstrated that the size of the binding pocket is the most important factor in determining the substrate specificities of 4CL1. These findings shed light on the enzymatic mechanisms of 4CLs and provide a solid foundation for the bioengineering of these enzymes.

Bark tissue of Populus 3 canescens can hyperaccumulate cadmium, but microstructural, transcriptomic, and physiological response mechanisms are poorly understood. Histochemical assays, transmission electron microscopic observations, energydispersive x-ray microanalysis, and transcriptomic and physiological analyses have been performed to enhance our understanding of cadmium accumulation and detoxification in P. 3 canescens. Cadmium was allocated to the phloem of the bark, and subcellular cadmium compartmentalization occurred mainly in vacuoles of phloem cells. Transcripts involved in microstructural alteration, changes in nutrition and primary metabolism, and stimulation of stress responses showed significantly differential expression in the bark of P. 3 canescens exposed to cadmium. About 48% of the differentially regulated transcripts formed a coregulation network in which 43 hub genes played a central role both in cross talk among distinct biological processes and in coordinating the transcriptomic regulation in the bark of P. 3 canescens in response to cadmium. The cadmium transcriptome in the bark of P. 3 canescens was mirrored by physiological readouts. Cadmium accumulation led to decreased total nitrogen, phosphorus, and calcium and increased sulfur in the bark. Cadmium inhibited photosynthesis, resulting in decreased carbohydrate levels. Cadmium induced oxidative stress and antioxidants, including free proline, soluble phenolics, ascorbate, and thiol compounds. These results suggest that orchestrated microstructural, transcriptomic, and physiological regulation may sustain cadmium hyperaccumulation in P. 3 canescens bark and provide new insights into engineering woody plants for phytoremediation.

To investigate how N-fertilization affects the growth, carbon and nitrogen (N) physiology, and wood properties of poplars with contrasting growth characteristics, slow-growing (Populus popularis, Pp) and fast-growing (P. alba×P. glandulosa, Pg) poplar saplings were exposed to different N levels. Above-ground biomass, leaf area, photosynthetic rates (A), instantaneous photosynthetic nitrogen use efficiency (PNUE i), chlorophyll and foliar sugar concentrations were higher in Pg than in Pp. Foliar nitrate reductase (NR) activities and root glutamate synthase (GOGAT) activities were higher in Pg than in Pp as were the N amount and NUE of new shoots. Lignin contents and calorific values of Pg wood were less than that of Pp wood. N-fertilization reduced root biomass of Pg more than of Pp, but increased leaf biomass, leaf area, A, and PNUEi of Pg more than of Pp. Among 13 genes involved in the transport of ammonium or nitrate or in N assimilation, transcripts showed more pronounced changes to N-fertilization in Pg than in Pp. Increases in NR activities and N contents due to N-fertilization were larger in Pg than in Pp. In both species, N-fertilization resulted in lower calorific values as well as shorter and wider vessel elements/fibres. These results suggest that growth, carbon and N physiology, and wood properties are more sensitive to increasing N availability in fast-growing poplars than in slow-growing ones, which is probably due to prioritized resource allocation to the leaves and accelerated N physiological processes in fast-growing poplars under higher N levels.

Bicyclic analogues of the plant hormone abscisic acid (ABA) were designed to incorporate the structural elements and functional groups of the parent molecule that are required for biological activity. The resulting tetralone analogues were predicted to have enhanced biological activity in plants, in part because oxidized products would not cyclize to forms corresponding to the inactive catabolite phaseic acid. The tetralone analogues were synthesized in seven steps from 1-tetralone and a range of analogues were accessible through a second route starting with 2-methyl-1-naphthol. Tetralone ABA 8 was found to have greater activity than ABA in two bioassays. The absolute configuration of (+)-8 was established by X-ray crystallography of a RAMP hydrazone derivative. The hydroxymethyl compounds 10 and 11, analogues for studying the roles of 8- and 9-hydroxy ABA 3 and 6, were also synthesized and found to be active.

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Summary Here, we studied the poplar C19 gibberellin 2‐oxidase (GA2ox) gene subfamily. We show that a set of paralogous gene pairs differentially regulate shoot and root development. PtGA2ox4 and its paralogous gene PtGA2ox5 are primarily expressed in aerial organs, and overexpression of PtGA2ox5 produced a strong dwarfing phenotype characteristic of GA deficiency. Suppression of PtGA2ox4 and PtGA2ox5 led to increased biomass growth, but had no effect on root development. By contrast, the PtGA2ox2 and PtGA2ox7 paralogous pair was predominantly expressed in roots, and when these two genes were RNAi‐suppressed it led to a decrease of root biomass. The morphological changes in the transgenic plants were underpinned by tissue‐specific increases in bioactive GAs that corresponded to the predominant native expression of the targeted paralogous gene pair. Although RNAi suppression of both paralogous pairs led to changes in wood development, they were much greater in the transgenics with suppressed PtGA2ox4 and PtGA2ox5. The degree of gene suppression in independent events was strongly associated with phenotypes, demonstrating dose‐dependent control of growth by GA2ox RNA concentrations. The expression and transgenic modifications reported here show that shoot‐ and leaf‐expressed PtGA2ox4 and PtGA2ox5 specifically restrain aerial shoot growth, while root‐expressed PtGA2ox2 and PtGA2ox7 promote root development.

Phytohormones act at relatively low concentrations as major regulatory factors of plant growth and development, and cross talk of phytohormones is currently of great interest throughout the plant science community. To meet this demand, a method that is capable of simultaneously analyzing diverse plant hormones is essential. This paper introduces a high-performance liquid chromatographic separation technique coupled with sensitive and selective ion trap mass spectrometry to simultaneously determine 24 or more acidic and alkaline phytohormones, including auxin, cis- and trans-abscisic acid, 11 cytokinins, and 10 gibberellins, in a single injection of sample. A binary solid-phase extraction using Oasis MCX cartridges for cations and Oasis MAX cartridges for anions was used to prepurify more than 24 acidic and alkaline phytohormones from a single plant extract. The method showed good linearity for all 24 phytohormones with R(2) values ranging from 0.9903 to 0.9997. Limits of detection for most of the phytohormones were in the femtomole range with some extending into the sub-femtomole range. This method was applied to hundreds of plant samples comprising different tissues from various plants, including herbaceous, woody climbing, and woody plants to demonstrate feasibility and to validate the methodology.