OPT3 is a plasma membrane transporter preferentially expressed in phloem cells during iron starvation . Cell-specific microarrays and OPT3pro:GUS analysis under Fe-limiting conditions show preferential expression of OPT3 in phloem cells, suggesting a role of OPT3 in long-distance transport processes. Notably, shoot specific expression of OPT3 in the opt3-2 background rescued the constitutively high expression of IRT1 in roots, the seed Cd over-accumulation phenotype, and the seedling sensitivity to Cd . These results suggest that the impaired metal homeostasis in opt3-2 roots is caused by a disruption of the shoot-to-root signaling of the leaf metal status. Thus, OPT3 is the first shoot-expressed gene required for proper communication from leaves to roots to maintain metal homeostasis at the whole-plant level. Several Arabidopsis and tomato mutants displaying an Fe-deficiency response in roots can be rescued by foliar application of Fe ; these experiments suggest that shoot-to-root Fe signaling plays an important role in Fe homeostasis which in turn could also impact the uptake and accumulation of other transition metals such as Zn, Mn, and Cd, as seen in opt3-2 . Foliar application of Fe does not repress the Fe-deficiency response in opt3-2 roots to wild-type levels , suggesting that source-to-sink transport of Fe, or a molecule mediating Fe signaling, is impaired in opt3-2. Radiotracer experiments using 59Fe demonstrate that the movement of Fe between leaves is impaired in opt3-2 ; whether this leaf-to-leaf transport occurs as Fe2+ or as an Fe–ligand complex remains to be determined. OPT3 is a member of the oligopeptide transporter family and members of this family have been shown to mediate the transport of a broad range of peptides . Arabidopsis OPT3 has also been reported to rescue the ability of yeast mutants defective in Cu and Mn transport to grow on low concentrations of these transition metals . However, so far there is no direct evidence to suggest that OPT3 mediates the transport of transition metals, in the ionic form or complexed with a ligand,raspberry grow in pots or whether OPT3 mediates the transport of a ligand that facilitates the uptake and accumulation of transition metals into the cell.
In fact, our OPT3 localization experiments in yeast show that OPT3–YFP fusions are unable to transit out of the ER to the plasma membrane . This intracellular localization of OPT3 makes it difficult to interpret the ability of yeast strains defective in transition metal transport to grow on minimal media when expressing OPT3. Glutathione is a small peptide that has also gained recent attention in metal-status signaling via GSH coordinated intermediaries of the iron–sulfur cluster assembly machinery .Our radiotracer experiments using 35S-GSH and the ferric reductase assay in opt3-2 roots show that leaf-to-leaf movement of GSH was unaffected and that foliar application of GSH does not suppress the constitutive Fe-deficiency response in opt3-2. These results suggest that shoot-to-root transport of GSH alone has little effect on the long-distance signaling of the Fe status in Arabidopsis.Watercress is a leafy-green crop in the Brassicaceae family, consumed widely across the world for its peppery taste and known to be the most nutrient dense salad leaf . The peppery taste is the result of high concentrations of glucosinolates – phytochemicals which can be hydrolyzed to isothiocyanates upon plant tissue damage, such as chewing, known for their potent anticancer , anti-inflammatory , and antioxidant effects that are beneficial to human health. Although ITCs are the main products of digestion depending on pH, metal ions, and other epithio specifier proteins, nitriles can also be formed through GLS break-down and they too may have chemo preventitive properties . Watercress is high value horticultural crop. A specialty leafy vegetable, with a growing area of 282 ha in the US, with 75 ha of production in California, compared to 58 ha in the UK . It is also a high-value horticultural crop in the UK, with the market value of £8.90 per kg compared to £4.97 per kg for mixed baby leaf salad bags and represents a total value of £15 million per year . Watercress is traditionally grown in outdoor aquatic systems, but there is increasing interest in its suitability for indoor hydroponic systems, such as in vertical farms .
VF utilize hydroponic or aeroponic systems that allow plant stacking in multiple vertical or horizontal layers increasing the effective use of space and other resources, particularly water . Indoor vertical agriculture is well-suited to the production of leafy greens. Their fast growth rate, high harvest index, low photosynthetic energy demand and compact shape make them ideal for indoor farming technologies . VFs have multi-layered indoor crop production space with the use of artificial lights and soilless cultivation systems. With the capacity to control lighting, ventilation, irrigation, nutrient levels, and abiotic stress, VFs offer the potential of high and predictable yields and uniform produce alongside reduced water use and often no pesticide applications whatsoever . The future of indoor food production is likely to include other high-value horticultural crops such as several leafy greens, culinary herbs, strawberries, and flowers. Breeding targets for these crops include short life cycles, low energy demands, improved yield , small root systems, as well novel sensory and nutritional profiles . VF systems are gaining traction for commercial scale cultivation, partly due to their ability to deliver locally-grown food to urban areas, with lower environmental costs and also to deliver food in locations where fresh produce cannot be easily grown . These systems also offer a unique opportunity to tailor crop characteristics to changing consumer preferences by altering environmental conditions such as light quality for example , where blue light has been used to increase the glucosinolate content of several Brassica species including, pak choi and watercress . Here we investigate differences in yield, morphology, and glucosinolate content of watercress grown under three different cultivation systems . This research provides foundation information to suggest that high yield watercress crop production is possible in vertical farming systems and that watercress quality may be further enhanced for improved anti-cancer characteristics. A commercial watercress line with low heterozygosity and high vigor was cultivated from seeds germinated in plug trays until roots were well-established ,30 planter pot and then transplanted into one of three experimental sites. Seeds were germinated in a controlled-environment greenhouse and planted on Pro-Mix HP , 80:20 Baltic white: Baltic black peat , or rock wool in plug trays. From November 2020 to January 2021, this commercial line was grown in the UCD indoor VF .
This indoor farm has four walls of growth panels, hanging side by side with emitters at the top to supply water and nutrients. Light treatment 1 and light treatment 2 were applied after transplanting and stopped one day before harvesting. Growth light intensity was at 220 µmol/m2 /s photosynthetic photon flux density with both red and blue LEDs, 38 µmol/m2 /s under blue only and 187 µmol/m2 /s under red light only. Fertilizer containing macro and micro-nutrients was dosed to maintain an electrical conductivity at 400–800 μ. Fresh above ground weight and main stem diameter of each individual plant was recorded. Images of dissected plants were analyzed using ImageJ version 1.52d for leaf area. The shoot was bagged and oven-dried at 80 ◦C for 48 h for dry weight analysis. Glucosinolates were measured as previously described . Individual desulfo-GLSs within each sample were separated and detected by HPLC-DAD, identified, quantified by comparison to standard curves from purified compounds, and further normalized to equalize the fresh weight. Pure aromatic glucosinolate known to be in watercress was obtained from Extrasynthese to add to the existing collection of standards. We hypothesized that optimized light and temperature regimes in the VF system was likely to result in higher yield and glucosinolate content, as well as less variability between plants due to tightly controlled conditions. Comparing across growth environments, four morphological traits were grouped by locations. Plants showed high plasticity between environments. Watercress plants exhibited considerably different growth morphologies depending on the growing system . Despite being the system currently used for commercial cultivation, watercress yield was lowest in the UK trial; fresh weight was significantly higher in the CA and VF trials . Plants grown in CA, however had less palatable “woody” growth types, with smaller leaves, illustrated by a significantly higher dry weight compared to UK and VF . Stem diameter was not affected by growth environments . Most importantly, the harvestable part of the plant, leaf area, differed significantly between environments , with the smallest leaves produced in CA and the largest in the VF trials: mean individual leaf area for CA, UK VFBlue20 and VFBlue30 was 1.33 ± 0.46, 4.69 ± 0.23, 8.25 ± 0.33, and 8.75 ± 0.33, respectively. Increasing blue light in the VF treatments had no impact on morphology. Also, not all morphological traits changed unidirectionally . Glucosinolate composition and quantity was significantly impacted by growth environments. Short chain aliphatic GLSs and longer chain aliphatic GLSs were all found at higher concentrations when grown in the VF compared to the UK or CA. Low concentrations of Indole GLSs were found in all watercress , CA , VFBlue20 , and VFBlue30 . Conversely, the most abundant aromatic GLS compound in watercress, gluconasturiin, had mean concentrations of 0.56 ± 0.17 nmol/mg , 1.71 ± 0.17 nmol/mg , 1.98 ± 0.12 nmol/mg , and 2.18 ± 0.12 . The UK-grown watercress had the lowest PEGLS concentration compared to the other sites. GLS quality was altered in the red:blue light treatment of the VF, and a new compound PBGLS , an alkylglucosinolate homologous to PEGLS with a longer alkyl chain, was detected in the VF that was not apparent for outdoor grown watercress in either CA or the UK .
Interestingly, between the two light regimes of the VF trials, neither GLS quality nor quantity differed, but total GLS was altered between UK, CA, and VFBlue30 treatment, indicating that the GLS quantity profile changes under distinctive environments . We have shown that the quality and yield of the leafy green salad crop watercress can be significantly improved by growth in an indoor vertical hydroponic system, enriched in blue light. The CDC ranked watercress as the most nutrient dense crop based on the content of 17 nutrients that are associated with reducing chronic disease risk . Our results show the yield and nutrient content of watercress can be enhanced even further by utilizing a novel vertical indoor growing environment other than the current commercial system used in the UK. Yield increases may be explained by the ability to tightly control environmental conditions in the VF that generate a consistent optimal nutrient and temperature environment. The increase in glucosinolate content from UK to CA is probably explained by heat stress in CA, with the maximum temperature recorded at the CA site at 43.8 ◦C compared to 30.9 ◦C for the UK. Glucosinolate accumulation is associated with improved heat and drought stress tolerance in Arabidopsis and increases in GLSs areobserved in heat-stressed Brassica rapa . Increases observed in GLS content in VF can be explained by prolonged blue light exposure and a longer growth period . The mechanism of different LEDs on GLS biosynthesis regulations still remain unclear, but a short-duration blue light photoperiod increased the total aliphatic GLSs in broccoli . A similar result from a genome wide association mapping of Arabidopsis also revealed that blue light controlled GLS accumulation by altering the PHOT1/PHOT2 blue light receptors . Increasing blue light in the VF increased total GLSs content and although not statistically significant, it confirms the study by Chen et al. that showed increased GLSs content with increased blue light. Rosa et al. showed that GLS concentrations are more sensitive to the effect of temperature than of photoperiod and this is consistent with our results in total GLSs between the UK and CA sites. Our results support the idea that indoor farm cultivation is effective in promoting health-beneficial chemical properties. Watercress produced PBGLS in both the VF treatments, but this compound was not detected in either the UK or CA trials. PBGLS strengthens the nutrient profile of watercress. PEITC derived from PEGLS has already been proven to be an extremely effective naturally-occurring dietaryisothiocyanates against cancer .