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Copper homeostasis and the effects of copper deficiency on tomato plants and fruit quality

Periodic Reporting for period 1 - TOMACOP (Copper homeostasis and the effects of copper deficiency on tomato plants and fruit quality)

Periodo di rendicontazione: 2019-09-01 al 2021-08-31

Copper (Cu) is a vital micronutrient acting as a double-edged sword in living beings because it is an essential redox-active cofactor in biological processes but is toxic when in excess. Suboptimal Cu levels in human diet can cause impaired neurological development and metabolic disorders. In plants, Cu plays important roles in key processes. Consequently, plants are also sensitive to Cu bioavailability in the soil. Thus, low Cu levels may result in impaired pollen development and viability and regulation of responses to iron deficiency, but its toxicity causes DNA damage, chlorosis and root growth inhibition, among other symptoms. Since plant nutritional deficiencies or excesses are transferred to consumers, deciphering the regulatory mechanisms underlying Cu uptake and distribution to edible products is crucial to prevent deficient or toxic Cu levels in horticultural crops that may ultimately affect human health. Furthermore, in Europe, around 20% of the arable land is classified as Cu deficient, which has been compensated by using Cu-enriched fertilizers. However, the EU warns that this practice implies high environmental costs and compromises food security for consumers.
TOMACOP aimed to study the effects of Cu deficient availability in the soil on plant growth and development and on fruit nutritional status and quality by using tomato (Solanum lycopersicum) as experimental system. Major results indicate that deficient Cu availability had detrimental consequences on plant growth and yield and reduced the micronutritional value, marketability and postharvest quality of the fruit. The characterization of Cu homeostasis-related components and the identification of tissue-specificities in the molecular mechanisms regulating Cu uptake in this species has provided important clues for future biotechnological improvements aimed to solve the challenge facing EU agriculture.
We evaluated the physiological and biochemical changes in the plant as consequence of plant growth under suboptimal Cu availability throughout in vitro, hydroponics and greenhouse assays. Root and shoot lengths were significantly reduced, as well as the dry weight and biomass per plant. Also the number of flowers and the total yield was diminished as consequence of Cu deficiency conditions. In terms of external fruit quality and nutritional status, deficient Cu availability did not alter color evolution during the fruit ripening process but advanced fruit softening rate and increased internal acidity after harvest. In turn, this stress increased the antioxidant capacity of the fruit, probably due to increased vitamin C content, but had little effect on total phenolic, flavonoids and lycopene contents. Also, the incidence of fruit cracking and the susceptibility to pathogen infection increased in the fruit harvested from plants grown under Cu deficient conditions. Last, the contents of important micronutrients for human health, such as Cu and Fe, were changed in response to Cu deficiency stress conditions. These results have been divided in two different manuscripts and presented in the International Symposiums and in several diffusion seminars.
We also studied the molecular mechanisms underlying the Cu deficiency stress response in tomato plants and fruit. Six members of the copper transporters family were identified (SlCOPT1-6) and their secondary and tertiary structures, as well as the potential interaction network and gene expression patters were analyzed. Also, we assayed their functionality through complementation expression assays in yeast. These results point altogether that SlCOPT1 and SlCOPT2 are completely functional and the most ubiquitously expressed COPTs in the plant and fruit tissues. On the other hand, the expression of SlCOPT3 and SlCOPT5 is specialized in stem and fruit tissues. These results are already published (10.1016/j.ijbiomac.2021.10.032) and have been presented in the 6th ABS International Conference through an invited speech. In parallel, we have compared the transcriptome of different tissues (root, stem, leaf and fruit) of plants grown under Cu sufficiency and deficiency conditions. New generation sequencing results uncovered a set of common responses mainly directed to increase the Cu uptake in the root and to enhance its mobilization to upper parts of the plant.The major SlCOPT carrying out such functions was SlCOPT2, which point out this Cu transporter as the most plausible target for biotechnological improvement of Cu intake and distribution in tomato plants through genome-editing technologies. These results will be published in two independent manuscripts. On the other hand, the comparative transcriptomic analysis of the fruit harvested at the red ripe (commercial) ripening stage has revealed that this organ is not regulating a high number of biological processes in response to the Cu deficiency stress. These data are very valuable since they have allowed identifying fruit-specificities in the regulation of Cu homeostasis-related genes. This has a promising biotechnological potential in order to improve/optimize the micronutritional content of this product, which might has an impact on human diet and hence in human health.
This proposal deals with a subject of importance for worldwide Agriculture and Food Science using cutting-edge technologies developed during this research. On one hand, this project provides results that will contribute to optimize the tomato growing conditions and hence to reduce the loss of fruit nutritional and external quality and, consequently, the loss of economic value of fresh fruit. On the other hand, results obtained so far establish basic knowledge in tomato plants and fruit for the future development of biotechnological strategies aimed to enhance Cu deficiency/toxicity resilience. Also, these results open new research lines focusing on disease resistance by improving Cu content in the fruit, which will lead to a reduction in the use of pesticides, and to a better understanding of the mechanistic of fruit cracking incidence, which will impact fruit marketability and food waste. Both applied and basic research components are integral to those strategies that the EU are currently promoting, as reflected in Horizon 2020 designating the food security, sustainable agriculture and resources efficiency as key societal challenges.
Furthermore, translation of basic research into the breeding of Cu stress-tolerant crops is a very important challenge for food security under global conditions where Cu is either a nutrient lacking in the soil or a toxic element accumulated as consequence of irresponsible use of fertilizers. Because the global population continues to increase, and longevity has also been rapidly increasing, it is not only important for human health to increase food production but also to improve food quality. The successful application of basic knowledge from plant science and genomics will be crucial for future food security and agroindustry in the world. Such translational research for the sustainable production of more healthy and better foods is a visible contribution of plant science not only to European but also to worldwide society.
In silico analysis
Hydroponic assays
In vitro assays
Transcriptomic results