Flavonols regulate root hair development by modulating accumulation of reactive oxygen species in the root epidermis.

Reactive oxygen species (ROS) are signaling molecules produced by tissue-specific respiratory burst oxidase homolog (RBOH) enzymes to drive development. In Arabidopsis thaliana, ROS produced by RBOHC was previously reported to drive root hair elongation. We identified a specific role for one ROS, H2O2, in driving root hair initiation and demonstrated that localized synthesis of flavonol antioxidants control the level of H2O2 and root hair formation. Root hairs form from trichoblast cells that express RBOHC and have elevated H2O2 compared with adjacent atrichoblast cells that do not form root hairs. The flavonol-deficient tt4 mutant has elevated ROS in trichoblasts and elevated frequency of root hair formation compared with the wild type. The increases in ROS and root hairs in tt4 are reversed by genetic or chemical complementation. Auxin-induced root hair initiation and ROS accumulation were reduced in an rbohc mutant and increased in tt4, consistent with flavonols modulating ROS and auxin transport. These results support a model in which localized synthesis of RBOHC and flavonol antioxidants establish patterns of ROS accumulation that drive root hair formation.

RBOH-Dependent ROS Synthesis and ROS Scavenging by Plant Specialized Metabolites To Modulate Plant Development and Stress Responses.

Reactive oxygen species (ROS) regulate plant growth and development. ROS are kept at low levels in cells to prevent oxidative damage, allowing them to be effective signaling molecules upon increased synthesis. In plants and animals, NADPH oxidase/respiratory burst oxidase homolog (RBOH) proteins provide localized ROS bursts to regulate growth, developmental processes, and stress responses. This review details ROS production via RBOH enzymes in the context of plant development and stress responses and defines the locations and tissues in which members of this family function in the model plant Arabidopsis thaliana. To ensure that these ROS signals do not reach damaging levels, plants use an array of antioxidant strategies. In addition to antioxidant machineries similar to those found in animals, plants also have a variety of specialized metabolites that scavenge ROS. These plant specialized metabolites exhibit immense structural diversity and have highly localized accumulation. This makes them important players in plant developmental processes and stress responses that use ROS-dependent signaling mechanisms. This review summarizes the unique properties of plant specialized metabolites, including carotenoids, ascorbate, tocochromanols (vitamin E), and flavonoids, in modulating ROS homeostasis. Flavonols, a subclass of flavonoids with potent antioxidant activity, are induced during stress and development, suggesting that they have a role in maintaining ROS homeostasis. Recent results using genetic approaches have shown how flavonols regulate development and stress responses through their action as antioxidants.

Local and systemic signaling of iron status and its interactions with homeostasis of other essential elements.

Iron (Fe) is essential for plant growth and development. However, alkaline soils, which occupy approximately 30% of the world's arable lands, are considered Fe-limiting for plant growth because insoluble Fe (III) chelates prevail under these conditions. In contrast, high bioavailability of Fe in acidic soils can be toxic to plants due to the ability of Fe ions to promote oxidative stress. Therefore, plants have evolved sophisticated mechanisms to sense and respond to the fluctuation of Fe availability in the immediate environment and to the needs of developing shoot tissues to preclude deficiency while avoiding toxicity. In this review, we focus on recent advances in our understanding of local and systemic signaling of Fe status with emphasis on the contribution of Fe, its interaction with other metals and metal ligands in triggering molecular responses that regulate Fe uptake and partitioning in the plant body.

Brachypodium distachyon as a model system for studies of copper transport in cereal crops.

Copper (Cu) is an essential micronutrient that performs a remarkable array of functions in plants including photosynthesis, cell wall remodeling, flowering, and seed set. Of the world's major cereal crops, wheat, barley, and oat are the most sensitive to Cu deficiency. Cu deficient soils include alkaline soils, which occupy approximately 30% of the world's arable lands, and organic soils that occupy an estimated 19% of arable land in Europe. We used Brachypodium distachyon (brachypodium) as a proxy for wheat and other grain cereals to initiate analyses of the molecular mechanisms underlying their increased susceptibility to Cu deficiency. In this report, we focus on members of the CTR/COPT family of Cu transporters because their homologs in A. thaliana are transcriptionally upregulated in Cu-limited conditions and are involved either in Cu uptake from soils into epidermal cells in the root, or long-distance transport and distribution of Cu in photosynthetic tissues. We found that of five COPT proteins in brachypodium, BdCOPT3, and BdCOPT4 localize to the plasma membrane and are transcriptionally upregulated in roots and leaves by Cu deficiency. We also found that BdCOPT3, BdCOPT4, and BdCOPT5 confer low affinity Cu transport, in contrast to their counterparts in A. thaliana that confer high affinity Cu transport. These data suggest that increased sensitivity to Cu deficiency in some grass species may arise from lower efficiency and, possibly, other properties of components of Cu uptake and tissue partitioning systems and reinforce the importance of using brachypodium as a model for the comprehensive analyses of Cu homeostasis in cereal crops.

OPT3 Is a Phloem-Specific Iron Transporter That Is Essential for Systemic Iron Signaling and Redistribution of Iron and Cadmium in Arabidopsis.

Iron is essential for both plant growth and human health and nutrition. Knowledge of the signaling mechanisms that communicate iron demand from shoots to roots to regulate iron uptake as well as the transport systems mediating iron partitioning into edible plant tissues is critical for the development of crop biofortification strategies. Here, we report that OPT3, previously classified as an oligopeptide transporter, is a plasma membrane transporter capable of transporting transition ions in vitro. Studies in Arabidopsis thaliana show that OPT3 loads iron into the phloem, facilitates iron recirculation from the xylem to the phloem, and regulates both shoot-to-root iron signaling and iron redistribution from mature to developing tissues. We also uncovered an aspect of crosstalk between iron homeostasis and cadmium partitioning that is mediated by OPT3. Together, these discoveries provide promising avenues for targeted strategies directed at increasing iron while decreasing cadmium density in the edible portions of crops and improving agricultural productivity in iron deficient soils.

The CTR/COPT-dependent copper uptake and SPL7-dependent copper deficiency responses are required for basal cadmium tolerance in A. thaliana.

Copper (Cu) homeostasis in plants is maintained by at least two mechanisms: (1) the miRNA-dependent reallocation of intracellular Cu among major Cu-enzymes and important energy-related functions; (2) the regulation of the expression of Cu transporters including members of the CTR/COPT family. These events are controlled by the transcription factor SPL7 in Arabidopsis thaliana. Cadmium (Cd), on the other hand, is a non-essential and a highly toxic metal that interferes with homeostasis of essential elements by competing for cellular binding sites. Whether Cd affects Cu homeostasis in plants is unknown. We found that Cd stimulates Cu accumulation in roots of A. thaliana and increases mRNA expression of three plasma membrane-localized Cu uptake transporters, COPT1, COPT2 and COPT6. Further analysis of Cd sensitivity of single and triple copt1copt2copt6 mutants, and transgenic plants ectopically expressing COPT6 suggested that Cu uptake is an essential component of Cd resistance in A. thaliana. Analysis of the contribution of the SPL7-dependent pathway to Cd-induced expression of COPT1, COPT2 and COPT6 showed that it occurs, in part, through mimicking the SPL7-dependent transcriptional Cu deficiency response. This response also involves components of the Cu reallocation system, miRNA398, FSD1, CSD1 and CSD2. Furthermore, seedlings of the spl7-1 mutant accumulate up to 2-fold less Cu in roots than the wild-type, are hypersensitive to Cd, and are more sensitive to Cd than the triple copt1copt2copt6 mutant. Together these data show that exposure to excess Cd triggers SPL7-dependent Cu deficiency responses that include Cu uptake and reallocation that are required for basal Cd tolerance in A. thaliana.

COPT6 is a plasma membrane transporter that functions in copper homeostasis in Arabidopsis and is a novel target of SQUAMOSA promoter-binding protein-like 7.

Among the mechanisms controlling copper homeostasis in plants is the regulation of its uptake and tissue partitioning. Here we characterized a newly identified member of the conserved CTR/COPT family of copper transporters in Arabidopsis thaliana, COPT6. We showed that COPT6 resides at the plasma membrane and mediates copper accumulation when expressed in the Saccharomyces cerevisiae copper uptake mutant. Although the primary sequence of COPT6 contains the family conserved domains, including methionine-rich motifs in the extracellular N-terminal domain and a second transmembrane helix (TM2), it is different from the founding family member, S. cerevisiae Ctr1p. This conclusion was based on the finding that although the positionally conserved Met(106) residue in the TM2 of COPT6 is functionally essential, the conserved Met(27) in the N-terminal domain is not. Structure-function studies revealed that the N-terminal domain is dispensable for COPT6 function in copper-replete conditions but is important under copper-limiting conditions. In addition, COPT6 interacts with itself and with its homolog, COPT1, unlike Ctr1p, which interacts only with itself. Analyses of the expression pattern showed that although COPT6 is expressed in different cell types of different plant organs, the bulk of its expression is located in the vasculature. We also show that COPT6 expression is regulated by copper availability that, in part, is controlled by a master regulator of copper homeostasis, SPL7. Finally, studies using the A. thaliana copt6-1 mutant and plants overexpressing COPT6 revealed its essential role during copper limitation and excess.