Crocus sativus L. Ecotypes from Mediterranean Countries: Phenological, Morpho-Productive, Qualitative and Genetic Traits

The characterization of C. sativus ecotypes is of great interest for preserving them from a possible genetic erosion due to the decrease of European cultivation surface. In this study, we evaluated four ecotypes from Italy (Sardinia and Abruzzo), Spain (Castilla-La Mancha), and Greece (Kozani) in order to detect the existence of variability and promote the biodiversity of this crop. Thirty-one traits related to saffron flowering, flower morphology, production of spice and daughter corms, vegetative development (leaf and corm traits), and spice quality, were evaluated. In addition, a genetic analysis through three PCR-based approaches, SSRs, RAPD, and SRAP was assessed. Results highlighted a phenotypic variation among ecotypes during two consecutive years. All the studied parameters were influenced by the ecotype except for the stamen length, color coordinates of tepals, leaf length, and leaf number per plant. Sardinia had a longer flowering interval, earlier flowering, and higher spice yield and quality than the other corm origins. The maximum values of morphological traits, such as stigma length, dry weight of stigmas, tepals, flowers and leaves, leaf area, and daughter corm weight were observed in the Abruzzo ecotype. Principal component analysis (PCA) showed a clear separation among ecotypes, in which Sardinia and Spain showed more similarities than Abruzzo and Kozani. Significant negative correlation was found between days to flower with stigma yield and quality. However, we could not find molecular markers discriminating among corm origins. In conclusion, this study suggests the importance of C. sativus ecotypes as precious source of biodiversity and bioactive compounds, and of their enhancement as fundamental prerequisite for a sustainable development strategy and as an agricultural diversification opportunity for growers

Arabidopsis ILITHYIA protein is necessary for proper chloroplast biogenesis and root development independent of eIF2alpha phosphorylation

[EN] One of the main mechanisms blocking translation after stress situations is mediated by phosphorylation of the alpha-subunit of the eukaryotic initiation factor 2 (eIF2), performed in Arabidopsis by the protein kinase GCN2 which interacts and is activated by ILITHYIA(ILA). ILA is involved in plant immunity and its mutant lines present phenotypes not shared by the gcn2 mutants. The functional link between these two genes remains elusive in plants. In this study, we show that, although both ILA and GCN2 genes are necessary to mediate eIF2 alpha phosphorylation upon treatments with the aromatic amino acid biosynthesis inhibitor glyphosate, their mutants develop distinct root and chloroplast phenotypes. Electron microscopy experiments reveal that ila mutants, but not gcn2, are affected in chloroplast biogenesis, explaining the macroscopic phenotype previously observed for these mutants. ila3 mutants present a complex transcriptional reprogramming affecting defense responses, photosynthesis and protein folding, among others. Double mutant analyses suggest that ILA has a distinct function which is independent of GCN2 and eIF2 alpha phosphorylation. These results suggest that these two genes may have common but also distinct functions in Arabidopsis.Microarray experiments were done in the Genomics Facility of the IBMCP. MTH was supported by the Austrian Science Found (FWF) projectF03707. This work has been supported by the Spanish Ministry for Science and Education (Plan Nacional 2008-2011).Faus, I.; Niñoles Rodenes, R.; Kesari, V.; Llabata, P.; Tam, E.; Nebauer, SG.; Santiago, J.... (2018). Arabidopsis ILITHYIA protein is necessary for proper chloroplast biogenesis and root development independent of eIF2alpha phosphorylation. Journal of Plant Physiology. 224:173-182. https://doi.org/10.1016/j.jplph.2018.04.003S17318222

Rootstock alleviates PEG-induced water stress in grafted pepper seedlings: Physiological responses

Recent studies have shown that tolerance to abiotic stress, including water stress, is improved by grafting. In a previous work, we took advantage of the natural variability of Capsicum spp. and selected accessions tolerant and sensitive to water stress as rootstocks. The behavior of commercial cultivar 'Verset' seedlings grafted onto the selected rootstocks at two levels of water stress provoked by adding 3.5 and 7% PEG (polyethylene glycol) was examined over 14 days. The objective was to identify the physiological traits responsible for the tolerance provided by the rootstock in order to determine if the tolerance is based on the maintenance of the water relations under water stress or through the activation of protective mechanisms. To achieve this goal, various physiological parameters were measured, including: water relations; proline accumulation; gas exchange; chlorophyll fluorescence; nitrate reductase activity; and antioxidant capacity. Our results indicate that the effect of water stress on the measured parameters depends on the duration and intensity of the stress level, as well as the rootstock used. Under control conditions (0% PEG) all plant combinations showed similar values for all measured parameters. In general terms, PEG provoked a strong decrease in the gas exchange parameters in the cultivar grafted onto the sensitive accessions, as also observed in the ungrafted plants. This effect was related to lower relative water content in the plants, provoked by an inefficient osmotic adjustment that was dependent on reduced proline accumulation. At the end of the experiment, chronic photoinhibition was observed in these plants. However, the plants grafted onto the tolerant rootstocks, despite the reduction in photosynthetic rate, maintained the protective capacity of the photosynthetic machinery mediated by osmotic adjustment (based on higher proline content). In addition, water stress limited uptake and further NO3- transfer to the leaves. Increased nitrate reductase activity in the roots was observed, mainly in plants grafted onto the sensitive rootstocks, as well as the ungrafted plants, and this was associated with the lessened flux to the leaves. This study suggests that PEG-induced water stress can be partially alleviated by using tolerant accessions as rootstocks. (C) 2014 Elsevier GmbH. All rights reserved

Flower induction and development in saffron: Timing and hormone signalling pathways

[EN] The demand for saffron is expected to rise in the coming years due to its nutraceutical and medicinal properties. To cope with this, it will be necessary to develop a mechanised production of saffron. Upgrading the production methods requires accurate control of the flowering time in this species. Nevertheless, little is known about the control of flowering time in Crocus sativus L. The aim of this study is to gain insight into the floral induction regulatory networks operating in this species. A transcriptomic analysis was performed from saffron main buds in different stages of development. The identification of putative integrators of flowering time signals, like FT, as well as meristem identity genes, such as LFY and TFL1, permitted the definition of the time of flowering induction of the buds, being able to use them as molecular markers. The identification of the transcripts encoded by a DROOPING LEAF-like (DL) gene is of particular relevance because this gene might be a novel factor for carpel specification in saffron. To elucidate the hormonal signalling networks working during flower induction, transcriptomic data were used, and the content of IAA, ABA and gibberellins was determined in competent and non-competent buds to flower, during the saffron life cycle. Our results suggested that ABA might be negatively regulating corm dormancy release, but its involvement in flower induction cannot be ruled out. ABI5 and the mediator of ABA regulated dormancy gene MARD1, could be key players of this pathway. In addition, a drop in GA4 levels may also be a necessary, but insufficient, condition for floral induction and development. DELLA, TFL1 and PIF3 genes might be involved in the gibberellin pathway. Notably, IAA seems to be a positive regulator of the process, involving MP/ARF5 and ANT genes in the pathway. Taken together, these results pave the way to the unveiling of the regulatory networks controlling the vegetative-to-reproductive phase change in saffron.The activities of this study have been supported by a project funded by the "Ministerio de Ciencia, Innovacion y Universidades de Espana" [AGL2016-77078-R].Renau-Morata, B.; Nebauer, SG.; García-Carpintero, V.; Cañizares Sales, J.; Minguet, E.; De Los Mozos, M.; Molina Romero, RV. (2021). Flower induction and development in saffron: Timing and hormone signalling pathways. Industrial Crops and Products. 164:1-19. https://doi.org/10.1016/j.indcrop.2021.113370S11916

Evaluation of some pepper genotypes as rootstocks in water stress conditions

Water stress is a major environmental factor that limits crop production and it is important to develop crop varieties with higher yield under water scarcity. Increased pepper tolerance to water stress through grafting onto robust rootstocks could be an optimal alternative in the context of environmentally friendly agriculture. Our work evaluated the behaviour of 18 pepper genotypes during vegetative and reproductive stages under water stress in order to select tolerant genotypes to be used as rootstocks for pepper cultivation. The pepper tolerance screening was based on photosynthetic parameters. The genotypes Atlante, C-40, Serrano, PI-152225, ECU-973, BOL-58 and NuMex Conquistador were revealed as the most tolerant genotypes to water stress because they maintained net photosynthetic rate levels under water stress conditions. The selected genotypes were validated as rootstocks on a pepper cultivar in terms of productivity under severe water stress. Plants grafted onto cvs Atlante, PI-152225 and ECU-973 showed higher marketable yields when compared with ungrafted cultivar

Phosphoglycerate Kinases Are Co-Regulated to Adjust Metabolism and to Optimize Growth

[EN] In plants, phosphoglycerate kinase (PGK) converts 1,3-bisphosphoglycerate into 3-phosphoglycerate in glycolysis but also participates in the reverse reaction in gluconeogenesis and the Calvin-Benson cycle. In the databases, we found three genes that encode putative PGKs. Arabidopsis (Arabidopsis thaliana) PGK1 was localized exclusively in the chloroplasts of photosynthetic tissues, while PGK2 was expressed in the chloroplast/plastid of photosynthetic and nonphotosynthetic cells. PGK3 was expressed ubiquitously in the cytosol of all studied cell types. Measurements of carbohydrate content and photosynthetic activities in PGK mutants and silenced lines corroborated that PGK1 was the photosynthetic isoform, while PGK2 and PGK3 were the plastidial and cytosolic glycolytic isoforms, respectively. The pgk1.1 knockdown mutant displayed reduced growth, lower photosynthetic capacity, and starch content. The pgk3.2 knockout mutant was characterized by reduced growth but higher starch levels than the wild type. The pgk1.1 pgk3.2 double mutant was bigger than pgk3.2 and displayed an intermediate phenotype between the two single mutants in all measured biochemical and physiological parameters. Expression studies in PGK mutants showed that PGK1 and PGK3 were down-regulated in pgk3.2 and pgk1.1, respectively. These results indicate that the down-regulation of photosynthetic activity could be a plant strategy when glycolysis is impaired to achieve metabolic adjustment and optimize growth. The double mutants of PGK3 and the triose-phosphate transporter (pgk3.2 tpt3) displayed a drastic growth phenotype, but they were viable. This implies that other enzymes or nonspecific chloroplast transporters could provide 3-phosphoglycerate to the cytosol. Our results highlight both the complexity and the plasticity of the plant primary metabolic network.This work has been funded by the Spanish Government and the European Union: FEDER/ BFU2012-31519 and FEDER/ BFU2015-64204R, FPI fellowship to S.R.-T., and the Valencian Regional Government: PROMETEO II/2014/052.Rosa-Tellez, S.; Anoman, A.; Flores-Tornero, M.; Toujani, W.; Alseek, S.; Fernie, A.; Nebauer, SG.... (2018). Phosphoglycerate Kinases Are Co-Regulated to Adjust Metabolism and to Optimize Growth. PLANT PHYSIOLOGY. 176(2):1182-1198. https://doi.org/10.1104/pp.17.01227S11821198176

The targeted overexpression of SlCDF4 in the fruit enhances tomato size and yield involving gibberellin signalling

[EN] Tomato is one of the most widely cultivated vegetable crops and a model for studying fruit biology. Although several genes involved in the traits of fruit quality, development and size have been identified, little is known about the regulatory genes controlling its growth. In this study, we characterized the role of the tomato SlCDF4 gene in fruit development, a cycling DOF-type transcription factor highly expressed in fruits. The targeted overexpression of SlCDF4 gene in the fruit induced an increased yield based on a higher amount of both water and dry matter accumulated in the fruits. Accordingly, transcript levels of genes involved in water transport and cell division and expansion during the fruit enlargement phase also increased. Furthermore, the larger amount of biomass partitioned to the fruit relied on the greater sink strength of the fruits induced by the increased activity of sucrose-metabolising enzymes. Additionally, our results suggest a positive role of SlCDF4 in the gibberellin-signalling pathway through the modulation of GA(4) biosynthesis. Finally, the overexpression of SlCDF4 also promoted changes in the profile of carbon and nitrogen compounds related to fruit quality. Overall, our results unveil SlCDF4 as a new key factor controlling tomato size and composition.Renau-Morata, B.; Carrillo, L.; Cebolla Cornejo, J.; Molina Romero, RV.; Martí-Renau, R.; Domínguez-Figueroa, J.; Vicente-Carbajosa, J.... (2020). The targeted overexpression of SlCDF4 in the fruit enhances tomato size and yield involving gibberellin signalling. Scientific Reports. 10(1):1-14. https://doi.org/10.1038/s41598-020-67537-x1141011FAO. Crops production database. FAOSTAT. Latest update: 04/03/2020. Food and Agriculture Organization of the United Nations. Rome https://www.fao.org/faostat (2018).Willcox, J. K., Catignani, G. L. & Lazarus, S. Tomatoes and cardiovascular health. Crit. Rev. Food Sci. 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The Arabidopsis Transcription Factor CDF3 Is Involved in Nitrogen Responses and Improves Nitrogen Use Efficiency in Tomato

Nitrate is an essential macronutrient and a signal molecule that regulates the expression of multiple genes involved in plant growth and development. Here, we describe the participation of Arabidopsis DNA binding with one finger (DOF) transcription factor CDF3 in nitrate responses and shows that CDF3 gene is induced under nitrate starvation. Moreover, knockout cdf3 mutant plants exhibit nitrate-dependent lateral and primary root modifications, whereas CDF3 overexpression plants show increased biomass and enhanced root development under both nitrogen poor and rich conditions. Expression analyses of 35S::CDF3 lines reveled that CDF3 regulates the expression of an important set of nitrate responsive genes including, glutamine synthetase-1, glutamate synthase-2, nitrate reductase-1, and nitrate transporters NRT2.1, NRT2.4, and NRT2.5 as well as carbon assimilation genes like PK1 and PEPC1 in response to N availability. Consistently, metabolite profiling disclosed that the total amount of key N metabolites like glutamate, glutamine, and asparagine were higher in CDF3-overexpressing plants, but lower in cdf3-1 in N limiting conditions. Moreover, overexpression of CDF3 in tomato increased N accumulation and yield efficiency under both optimum and limiting N supply. These results highlight CDF3 as an important regulatory factor for the nitrate response, and its potential for improving N use efficiency in crops

Ectopic expression of the AtCDF1 transcription factor in potato enhances tuber starch and amino acid contents and yield under open field conditions

Introduction Cycling Dof transcription factors (CDFs) have been involved in different aspects of plant growth and development. In Arabidopsis and tomato, one member of this family (CDF1) has recently been associated with the regulation of primary metabolism and abiotic stress responses, but their roles in crop production under open field conditions remain unknown. Methods In this study, we compared the growth, and tuber yield and composition of plants ectopically expressing the CDF1 gene from Arabidopsis under the control of the 35S promoter with wild-type (WT) potato plants cultured in growth chamber and open field conditions. Results In growth chambers, the 35S::AtCDF1 plants showed a greater tuber yield than the WT by increasing the biomass partition for tuber development. Under field conditions, the ectopic expression of CDF1 also promoted the sink strength of the tubers, since 35S::AtCDF1 plants exhibited significant increases in tuber size and weight resulting in higher tuber yield. A metabolomic analysis revealed that tubers of 35S::AtCDF1 plants cultured under open field conditions accumulated higher levels of glucose, starch and amino acids than WT tubers. A comparative proteomic analysis of tubers of 35S::AtCDF1 and WT plants cultured under open field conditions revealed that these changes can be accounted for changes in the expression of proteins involved in energy production and different aspects of C and N metabolism. Discussion The results from this study advance our collective understanding of the role of CDFs and are of great interest for the purposes of improving the yield and breeding of crop plants

Ongoing Evolution in the Genus Crocus: Diversityof Flowering Strategies on the Way to Hysteranthy

[EN] Species of the genus Crocus are found over a wide range of climatic areas. In natural habitats, these geophytes diverge in the flowering strategies. This variability was assessed by analyzing the flowering traits of the Spanish collection of wild crocuses, preserved in the Bank of Plant Germplasm of Cuenca. Plants of the seven Spanish species were analyzed both in their natural environments (58 native populations) and in common garden experiments (112 accessions). Differences among species observed in the native habitats were maintained under uniform environmental conditions, suggesting a genetic basis for flowering mechanisms. Two eco-morphological types, autumn- and spring-flowering species, share similar patterns of floral induction and differentiation period in summer. The optimal temperature for this process was 23 degrees C for both types. Unlike Irano-Turanian crocuses, spring-flowering Spanish species do not require low winter temperatures for flower elongation. Hysteranthous crocuses flower in autumn prior to leaf elongation. We conclude that the variability in flowering traits in crocuses is related to the genetic and environmental regulation of flower primordia differentiation and elongation prior to emergence above the soil surface. The elucidation of the physiological differences between eco-morphological types of crocuses: synanthous with cold requirements and synanthous and hysteranthous without cold requirements, unlocks a new approach to the flowering evolution of geophytes in Mediterranean regions. Crocus species can serve both as a new model in the study of the molecular basis of hysteranthy and for the purposes of developing the molecular markers for desirable flowering traits.The collection activities of plant materials included in this study were mainly supported by successive Spanish research projects funded by the "Instituto Nacional de Investigacion y Tecnologia Agraria y Alimentaria" [INIA RF2004-00032-C03, INIA RF2008-00012-C03, INIA RF2011-00005-C03], co-funded by the European Regional Development Fund (ERDF-FEDER), and also by means of the European Action 018 Agri Gen Res (CrocusBank). The activities of PhD. TPF were supported by a pre-doctoral grant from the "Instituto Nacional de Investigacion y Tecnologia Agraria y Alimentaria" within the framework of the project INIA RF2011-0005-C03-01. The preservation of these materials in the facilities of the Bank of Plant Germplasm of Cuenca (CIAF Albaladejito -IRIAF), as part of the Spanish Germplasm Collection of Saffron and other Crocus, is currently supported by the Spanish National Program for Conservation and Utilization of Plant Genetic Resources for Agriculture and Food (action INIA RFP2014-00012). Some activities have been also funded by the "Ministerio de Ciencia, Innovacion y Universidades de Espana" [AGL2016-77078-R].Pastor-Férriz, T.; De-Los-Mozos-Pascual, M.; Renau-Morata, B.; Nebauer, SG.; Sanchís, E.; Busconi, M.; Fernández, J.... (2021). Ongoing Evolution in the Genus Crocus: Diversityof Flowering Strategies on the Way to Hysteranthy. Plants. 10(3):1-18. https://doi.org/10.3390/plants1003047711810