A root specific induction of carotenoid biosynthesis contributes to ABA production upon salt stress in arabidopsis

Abscisic acid (ABA) is a hormone that plays a vital role in mediating abiotic stress responses in plants. Salt exposure induces the synthesis of ABA through the cleavage of carotenoid precursors (xanthophylls), which are found at very low levels in roots. Here we show that de novo ABA biosynthesis in salt-treated Arabidopsis thaliana roots involves an organ-specific induction of the carotenoid biosynthetic pathway. Upregulation of the genes encoding phytoene synthase (PSY) and other enzymes of the pathway producing ABA precursors was observed in roots but not in shoots after salt exposure. A pharmacological block of the carotenoid pathway substantially reduced ABA levels in stressed roots, confirming that an increase in carotenoid accumulation contributes to fuel hormone production after salt exposure. Treatment with exogenous ABA was also found to upregulate PSY expression only in roots, suggesting an organ-specific feedback regulation of the carotenoid pathway by ABA. Taken together, our results show that the presence of high concentrations of salt in the growth medium rapidly triggers a root-specific activation of the carotenoid pathway, probably to ensure a proper supply of ABA precursors required for a sustained production of the hormone

Making extra room for carotenoids in plant cells: new opportunities for biofortification

[EN] Plant carotenoids are essential for photosynthesis and photoprotection and provide colors in the yellow to red range to non-photosynthetic organs such as petals and ripe fruits. They are also the precursors of biologically active molecules not only in plants (including hormones and retrograde signals) but also in animals (including retinoids such as vitamin A). A carotenoid-rich diet has been associated with improved health and cognitive capacity in humans, whereas the use of carotenoids as natural pigments is widespread in the agrofood and cosmetic industries. The nutritional and economic relevance of carotenoids has spurred a large number of biotechnological strategies to enrich plant tissues with carotenoids. Most of such approaches to alter carotenoid contents in plants have been focused on manipulating their biosynthesis or degradation, whereas improving carotenoid sink capacity in plant tissues has received much less attention. Our knowledge on the molecular mechanisms influencing carotenoid storage in plants has substantially grown in the last years, opening new opportunities for carotenoid biofortification. Here we will review these advances with a particular focus on those creating extra room for carotenoids in plant cells either by promoting the differentiation of carotenoid-sequestering structures within plastids or by transferring carotenoid production to the cytosol.We greatly thank Carmen and Pilar Torres-Montilla for their collaboration and help in the design and execution of the figures. We also thank Luca Morelli and BioRender.com for some of the images. Work in our lab is funded by Spanish grants BIO2017-84041-P and PID2020-115810GB-I00 from the Agencia Estatal de Investigacion (AEI) and 202040E299 from Consejo Superior de Investigaciones Cientificas (CSIC) to MRC. STM was supported by PhD fellowship FPU16/04054 from the Spanish Ministerio de Educacion y CulturaTorres-Montilla, S.; Rodriguez-Concepcion, M. (2021). Making extra room for carotenoids in plant cells: new opportunities for biofortification. Progress in Lipid Research. 84:1-9. https://doi.org/10.1016/j.plipres.2021.101128S198

Tomato fruit carotenoid biosynthesis is adjusted to actual ripening progression by a light-dependent mechanism

Carotenoids are isoprenoid compounds that are essential for plants to protect the photosynthetic apparatus against excess light. They also function as health-promoting natural pigments that provide colors to ripe fruit, promoting seed dispersal by animals. Work in Arabidopsis thaliana unveiled that transcription factors of the phytochrome-interacting factor (PIF) family regulate carotenoid gene expression in response to environmental signals (i.e. light and temperature), including those created when sunlight reflects from or passes though nearby vegetation or canopy (referred to as shade). Here we show that PIFs use a virtually identical mechanism to modulate carotenoid biosynthesis during fruit ripening in tomato (Solanum lycopersicum). However, instead of integrating environmental information, PIF-mediated signaling pathways appear to fulfill a completely new function in the fruit. As tomatoes ripen, they turn from green to red due to chlorophyll breakdown and carotenoid accumulation. When sunlight passes through the flesh of green fruit, a self-shading effect within the tissue maintains high levels of PIFs that directly repress the master gene of the fruit carotenoid pathway, preventing undue production of carotenoids. This effect is attenuated as chlorophyll degrades, causing degradation of PIF proteins and boosting carotenoid biosynthesis as ripening progresses. Thus, shade signaling components may have been co-opted in tomato fruit to provide information on the actual stage of ripening (based on the pigment profile of the fruit at each moment) and thus finely coordinate fruit color change. We show how this mechanism may be manipulated to obtain carotenoid-enriched fruits.Peer ReviewedPostprint (published version

Nutritional Enrichment of Plant Leaves by Combining Genes Promoting Tocopherol Biosynthesis and Storage

The enrichment of plant tissues in tocochromanols (tocopherols and tocotrienols) is an important biotechnological goal due to their vitamin E and antioxidant properties. Improvements based on stimulating tocochromanol biosynthesis have repeatedly been achieved, however, enhancing sequestering and storage in plant plastids remains virtually unexplored. We previously showed that leaf chloroplasts can be converted into artificial chromoplasts with a proliferation of plastoglobules by overexpression of the bacterial crtB gene. Here we combined coexpression of crtB with genes involved in tocopherol biosynthesis to investigate the potential of artificial leaf chromoplasts for vitamin E accumulation in Nicotiana benthamiana leaves. We show that this combination improves tocopherol levels compared to controls without crtB and confirm that VTE1, VTE5, VTE6 and tyrA genes are useful to increase the total tocopherol levels, while VTE4 further leads to enrichment in alpha-tocopherol (the tocochromanol showing highest vitamin E activity). Additionally, we show that treatments that further promote plastoglobule formation (e.g., exposure to intense light or dark-induced senescence) result in even higher improvements in the tocopherol content of the leaves. An added advantage of our strategy is that it also results in increased levels of other related plastidial isoprenoids such as carotenoids (provitamin A) and phylloquinones (vitamin K1)

Differential Subplastidial Localization and Turnover of Enzymes Involved in Isoprenoid Biosynthesis in Chloroplasts

Plastidial isoprenoids are a diverse group of metabolites with roles in photosynthesis, growth regulation, and interaction with the environment. The methylerythritol 4-phosphate (MEP) pathway produces the metabolic precursors of all types of plastidial isoprenoids. Proteomics studies in Arabidopsis thaliana have shown that all the enzymes of the MEP pathway are localized in the plastid stroma. However, immunoblot analysis of chloroplast subfractions showed that the first two enzymes of the pathway, deoxyxylulose 5-phosphate synthase (DXS) and reductoisomerase (DXR), can also be found in non-stromal fractions. Both transient and stable expression of GFP-tagged DXS and DXR proteins confirmed the presence of the fusion proteins in distinct subplastidial compartments. In particular, DXR-GFP was found to accumulate in relatively large vesicles that could eventually be released from chloroplasts, presumably to be degraded by an autophagy-independent process. Together, we propose that protein-specific mechanisms control the localization and turnover of the first two enzymes of the MEP pathway in Arabidopsis chloroplasts

Intercropping maize with tomato plants improved the yield and fruit quality of tomato plants grown under salinity stress

Climate change represents a significant challenge for agriculture and food security. This phenomenon contributes substantially to food insecurity by increasing the frequency and severity of abiotic stresses such as salinity. Salt stress can unfavorably influence plant development and efficiency of numerous crops, particularly in arid and semi-arid zones. Different strategies and procedures can be utilized to moderate the negative impact of intemperate salt concentration within the soil. Here, the saltiness resistance of 2 Solanum lycopersicum L. varieties (Karima and Jade) grown in an intercropping system with maize was assessed by measuring fruit production and quality. Our results show that chlorophyll a was higher in the intercropping tomato than in the monoculture plants in both control (T0 = 0 mM NaCl) and NaCl-treated varieties (T1 = 125 mM NaCl). Moreover, in intercropping systems, the variety of Karima was less affected by NaCl treatment. However, for Jade variety, its intercropping with maize increased its production under both normal and salinity stress conditions. Regarding fruit quality, pH value of Karima was higher in intercropped plants under NaCl treatment, while the Brix value was less affected by NaCl and intercropping conditions. Finally, intercropping practices significantly influenced Na+ and K+ accumulation and Karima variety showed the capacity to accumulate more K+ and less Na+

The intrinsic chaperone network of Arabidopsis stem cells confers protection against proteotoxic stress

The biological purpose of plant stem cells is to maintain themselves while providing new pools of differentiated cells that form organs and rejuvenate or replace damaged tissues. Protein homeostasis or proteostasis is required for cell function and viability. However, the link between proteostasis and plant stem cell identity remains unknown. In contrast to their differentiated counterparts, we find that root stem cells can prevent the accumulation of aggregated proteins even under proteotoxic stress conditions such as heat stress or proteasome inhibition. Notably, root stem cells exhibit enhanced expression of distinct chaperones that maintain proteome integrity. Particularly, intrinsic high levels of the T-complex protein-1 ring complex/chaperonin containing TCP1 (TRiC/CCT) complex determine stem cell maintenance and their remarkable ability to suppress protein aggregation. Overexpression of CCT8, a key activator of TRiC/CCT assembly, is sufficient to ameliorate protein aggregation in differentiated cells and confer resistance to proteotoxic stress in plants. Taken together, our results indicate that enhanced proteostasis mechanisms in stem cells could be an important requirement for plants to persist under extreme environmental conditions and reach extreme long ages. Thus, proteostasis of stem cells can provide insights to design and breed plants tolerant to environmental challenges caused by the climate change

Synthetic conversion of leaf chloroplasts into carotenoid-rich plastids reveals mechanistic basis of natural chromoplast development

[EN] Plastids, the defining organelles of plant cells, undergo physiological and morphological changes to fulfill distinct biological functions. In particular, the differentiation of chloroplasts into chromoplasts results in an enhanced storage capacity for carotenoids with industrial and nutritional value such as beta-carotene (provitamin A). Here, we show that synthetically inducing a burst in the production of phytoene, the first committed intermediate of the carotenoid pathway, elicits an artificial chloroplast-to-chromoplast differentiation in leaves. Phytoene overproduction initially interferes with photosynthesis, acting as a metabolic threshold switch mechanism that weakens chloroplast identity. In a second stage, phytoene conversion into downstream carotenoids is required for the differentiation of chromoplasts, a process that involves a concurrent reprogramming of nuclear gene expression and plastid morphology for improved carotenoid storage. We hence demonstrate that loss of photosynthetic competence and enhanced production of carotenoids are not just consequences but requirements for chloroplasts to differentiate into chromoplasts.We greatly thank Jaume F. Martinez-Garcia and Ralf Welsch for fruitful discussions on the manuscript; Ralf Welsch and Li Li for providing seeds of the Arabidopsis ccd1 ccd4 and ator atorl mutants, respectively; Juan Jose Lopez-Moya and Maria Luisa Domingo-Calap for the gift of the HcProWMV-pGWB702 vector; and M. Rosa Rodriguez-Goberna for excellent technical support. The help of Marti Bernardo, Fidel Lozano, Lidia Jimenez, and members of the CRAG core facilities is also appreciated. This work was funded by the European Regional Development Fund and Spanish Agencia Estatal de Investigacion Grants BIO2017-84041-P, BIO2017-83184-R, BIO2017-90877-REDT, BES-2017-080652, and AGL2017-85563-C2-1-R; Ministry of Education, Culture and Sports Grants AP2012-3751 and FPU16/04054; and Generalitat de Catalunya Grant 2017SGR-710. We also thank the financial support of the European Union's Horizon 2020 (EU-H2020) COST Action CA15136 (EuroCaroten) and Marie S. Curie Action (MSCA) 753301 (Arcatom), the Severo Ochoa Programme for Centres of Excellence in R&D 2016-2019 Grant SEV-2015-0533 and the Generalitat de Catalunya CERCA Programme (to CRAG). B.L. is supported by grants from the CSIRO Synthetic Biology Future Science Platform and Macquarie University. L.M. is supported by La Caixa Foundation PhD INPhINIT Fellowship LCF/BQ/IN18/11660004, which received funding from the EU-H2020 through MSCA Grant 713673. A.R.F. is supported by Deutsche Forschungsgemeinschaft Grant DFG TRR 175.Llorente, B.; Torres-Montilla, S.; Morelli, L.; Florez-Sarasa, I.; Matus, JT.; Ezquerro, M.; D'andrea, L.... (2020). Synthetic conversion of leaf chloroplasts into carotenoid-rich plastids reveals mechanistic basis of natural chromoplast development. Proceedings of the National Academy of Sciences of the United States of America (Online). 117(35):21796-21803. https://doi.org/10.1073/pnas.2004405117S21796218031173

Metabolic crosstalk between hydroxylated monoterpenes and salicylic acid in tomato defense response against bacteria

[EN] Hydroxylated monoterpenes (HMTPs) are differentially emitted by tomato (Solanum lycopersicum) plants resisting bacterial infection. We have studied the defensive role of these volatiles in the tomato response to bacteria, whose main entrance is through stomatal apertures. Treatments with some HMTPs resulted in stomatal closure and pathogenesis-related protein 1 (PR1) induction. Particularly, alpha-terpineol induced stomatal closure in a salicylic acid (SA) and abscisic acid-independent manner and conferred resistance to bacteria. Interestingly, transgenic tomato plants overexpressing or silencing the monoterpene synthase MTS1, which displayed alterations in the emission of HMTPs, exhibited changes in the stomatal aperture but not in plant resistance. Measures of both 2-C-methyl-D-erythritol-2,4-cyclopyrophosphate (MEcPP) and SA levels revealed competition for MEcPP by the methylerythritol phosphate (MEP) pathway and SA biosynthesis activation, thus explaining the absence of resistance in transgenic plants. These results were confirmed by chemical inhibition of the MEP pathway, which alters MEcPP levels. Treatments with benzothiadiazole (BTH), a SA functional analog, conferred enhanced resistance to transgenic tomato plants overexpressing MTS1. Additionally, these MTS1 overexpressors induced PR1 gene expression and stomatal closure in neighboring plants. Our results confirm the role of HMTPs in both intra- and interplant immune signaling and reveal a metabolic crosstalk between the MEP and SA pathways in tomato plants.; Hydroxylated monoterpenes play an important role in both intra- and interplant immune signaling in tomato against bacterial infection.This work was supported by grant PID2020-116765RB-I00 funded by MCIN/AEI/10.13039/501100011033/ and grant PROMETEU/2021/056 by Generalitat Valenciana. C.P. was a recipient of a predoctoral contract of the Generalitat Valenciana (ACIF/2019/187), and J.P.-P. is a recipient of a JAEINT_21_02081 of the Consejo Superior de Investigaciones Cientificas and a predoctoral contract of the Ministerio de Universidades (FPU21/00259).Pérez, J.; Minguillón, S.; Kabbas-Piñango, E.; Payá, C.; Campos, L.; Rodriguez-Concepcion, M.; Espinosa-Ruiz, A.... (2024). Metabolic crosstalk between hydroxylated monoterpenes and salicylic acid in tomato defense response against bacteria. Plant Physiology. 195(3):2323-2338. https://doi.org/10.1093/plphys/kiae14823232338195

Metabolic crosstalk between hydroxylated monoterpenes and salicylic acid in tomato defense response against bacteria

Hydroxylated monoterpenes (HMTPs) are differentially emitted by tomato (Solanum lycopersicum) plants resisting bacterial infection. We have studied the defensive role of these volatiles in the tomato response to bacteria, whose main entrance is through stomatal apertures. Treatments with some HMTPs resulted in stomatal closure and pathogenesis-related protein 1 (PR1) induction. Particularly, α-terpineol induced stomatal closure in a salicylic acid (SA) and abscisic acid-independent manner and conferred resistance to bacteria. Interestingly, transgenic tomato plants overexpressing or silencing the monoterpene synthase MTS1, which displayed alterations in the emission of HMTPs, exhibited changes in the stomatal aperture but not in plant resistance. Measures of both 2-C-methyl-D-erythritol-2,4-cyclopyrophosphate (MEcPP) and SA levels revealed competition for MEcPP by the methylerythritol phosphate (MEP) pathway and SA biosynthesis activation, thus explaining the absence of resistance in transgenic plants. These results were confirmed by chemical inhibition of the MEP pathway, which alters MEcPP levels. Treatments with benzothiadiazole (BTH), a SA functional analog, conferred enhanced resistance to transgenic tomato plants overexpressing MTS1. Additionally, these MTS1 overexpressors induced PR1 gene expression and stomatal closure in neighboring plants. Our results confirm the role of HMTPs in both intra- and interplant immune signaling and reveal a metabolic crosstalk between the MEP and SA pathways in tomato plants