ABSTRACT
Covering: up to 2023Specialized metabolite (SM) modifications and/or decorations, corresponding to the addition or removal of functional groups (e.g. hydroxyl, methyl, glycosyl or acyl group) to SM structures, contribute to the huge diversity of structures, activities and functions of seed and plant SMs. This review summarizes available knowledge (up to 2023) on SM modifications in Brassicaceae and their contribution to SM plasticity. We give a comprehensive overview on enzymes involved in the addition or removal of these functional groups. Brassicaceae, including model (Arabidopsis thaliana) and crop (Brassica napus, Camelina sativa) plant species, present a large diversity of plant and seed SMs, which makes them valuable models to study SM modifications. In this review, particular attention is given to the environmental plasticity of SM and relative modification and/or decoration enzymes. Furthermore, a spotlight is given to SMs and related modification enzymes in seeds of Brassicaceae species. Seeds constitute a large reservoir of beneficial SMs and are one of the most important dietary sources, providing more than half of the world's intake of dietary proteins, oil and starch. The seed tissue- and stage-specific expressions of A. thaliana genes involved in SM modification are presented and discussed in the context of available literature. Given the major role in plant phytochemistry, biology and ecology, SM modifications constitute a subject of study contributing to the research and development in agroecology, pharmaceutical, cosmetics and food industrial sectors.
Subject(s)
Brassicaceae , Seeds , Seeds/metabolism , Seeds/chemistry , Brassicaceae/metabolism , Brassicaceae/chemistry , Molecular Structure , Plant Proteins/metabolismABSTRACT
Identifying traits that exhibit improved drought resistance is highly important to cope with the challenges of predicted climate change. We investigated the response of state transition mutants to drought. Compared with the wild type, state transition mutants were less affected by drought. Photosynthetic parameters in leaves probed by chlorophyll fluorescence confirmed that mutants possess a more reduced plastoquinone (PQ) pool, as expected due to the absence of state transitions. Seedlings of the mutants showed an enhanced growth of the primary root and more lateral root formation. The photosystem II inhibitor 3-(3,4-dichlorophenyl)-1,1-dimethylurea, leading to an oxidised PQ pool, inhibited primary root growth in wild type and mutants, while the cytochrome b6 f complex inhibitor 2,5-dibromo-3-methyl-6-isopropylbenzoquinone, leading to a reduced PQ pool, stimulated root growth. A more reduced state of the PQ pool was associated with a slight but significant increase in singlet oxygen production. Singlet oxygen may trigger a, yet unknown, signalling cascade promoting root growth. We propose that photosynthetic mutants with a deregulated ratio of photosystem II to photosystem I activity can provide a novel path for improving crop drought resistance.
Subject(s)
Photosystem II Protein Complex , Plastoquinone , Photosystem II Protein Complex/metabolism , Drought Resistance , Singlet Oxygen , Oxidation-Reduction , Photosynthesis/physiology , Chlorophyll , Electron Transport , LightABSTRACT
Strigolactones are compounds produced by plant roots in response to nutrient deficiency, acting both as local and systemic signals to control development and nutrition. Strigolactones are exuded in the rhizosphere to positively influence interactions with beneficial microbes. LC-MS/MS analysis shows that two genetically distinct grapevine rootstocks exudate one or two non-canonical strigolactones when subjected to low nitrogen conditions. Gene expression profiles and orobanche seed germination assays confirm that the biosynthesis and exudation of non-canonical compounds is the preferred pathway. The first compound, corresponding to heliolactone or 6-epi-heliolactone, is only exuded by the rootstock showing lower shoot branching and a higher level of mycorrhization with arbuscular mycorrhizal fungi. The structure of the second compound exuded by both rootstocks was identified by NMR and LC-MS/MS analysis. It is a non-canonical strigolactone, which has never been identified in another species. This first identification of a natural compound with the potential to stimulate beneficial root-microbe interactions in grapevines opens new perspectives in viticulture.
Subject(s)
Nitrogen , Plant Roots , Plant Roots/chemistry , Nitrogen/metabolism , Chromatography, Liquid , Germination/physiology , Tandem Mass Spectrometry , Lactones/chemistry , Exudates and Transudates/chemistry , Exudates and Transudates/metabolismABSTRACT
Andean potatoes (Solanum tuberosum L. ssp. andigena) are a good source of dietary antioxidant polyphenols. We have previously demonstrated that polyphenol extracts from Andean potato tubers exerted a dose-dependent cytotoxic effect in human neuroblastoma SH-SY5Y cells, being skin extracts more potent than flesh ones. In order to gain insight into the bioactivities of potato phenolics, we investigated the composition and the in vitro cytotoxic activity of total extracts and fractions of skin and flesh tubers of three Andean potato cultivars (Santa María, Waicha, and Moradita). Potato total extracts were subjected to liquid-liquid fractionation using ethyl acetate solvent in organic and aqueous fractions. We analyzed both fractions by HPLC-DAD, HPLC-ESI-MS/MS, and HPLC-HRMS. Results corroborated the expected composition of each fraction. Organic fractions were rich in hydroxycinnamic acids (principally chlorogenic acid isomers), whereas aqueous fractions contained mainly polyamines conjugated with phenolic acids, glycoalkaloids, and flavonoids. Aqueous fractions were cytotoxic against SH-SY5Y cells and even more potent than their respective total extracts. Treatment with a combination of both fractions showed a similar cytotoxic response to the corresponding extract. According to correlation studies, it is tempting to speculate that polyamines and glycoalkaloids are crucial in inducing cell death. Our findings indicate that the activity of Andean potato extracts is a combination of various compounds and contribute to the revalorization of potato as a functional food.
Subject(s)
Antineoplastic Agents , Neuroblastoma , Solanum tuberosum , Humans , Antineoplastic Agents/pharmacology , Antineoplastic Agents/metabolism , Plant Extracts/pharmacology , Plant Extracts/metabolism , Polyamines/metabolism , Polyphenols/metabolism , Solanum tuberosum/metabolism , Tandem Mass Spectrometry , MetabolomeABSTRACT
We have investigated the photophysics of aggregated lutein/violaxanthin in daffodil chromoplasts. We reveal the presence of three carotenoid aggregate species, the main one composed of a mixture of lutein/violaxanthin absorbing at 481 nm, and two secondary populations of aggregated carotenoids absorbing circa 500 and 402 nm. The major population exhibits an efficient singlet fission process, generating µs-lived triplet states on an ultrafast timescale. The structural organization of aggregated lutein/violaxanthin in daffodil chromoplasts produces well-defined electronic levels that permit the energetic pathways to be disentangled unequivocally, allowing us to propose a consistent mechanism for singlet fission in carotenoid aggregates. Transient absorption measurements on this system reveal for the first time an entangled triplet signature for carotenoid aggregates, and its evolution into dissociated triplet states. A clear picture of the carotenoid singlet fission pathway is obtained, which is usually blurred due to the intrinsic disorder of carotenoid aggregates.
Subject(s)
Fluorescent Dyes/chemistry , Lutein/chemistry , Dimerization , Kinetics , Molecular Conformation , Photochemical Processes , Plastids/chemistry , Spectrometry, Fluorescence , Xanthophylls/chemistryABSTRACT
In maize, nitrate regulates root development thanks to the coordinated action of many players. In this study, the involvement of strigolactones (SLs) and auxin as putative components of the nitrate regulation of lateral root (LR) was investigated. To this aim, the endogenous SL content of maize root in response to nitrate was assessed by liquid chromatography with tandem mass Spectrometry (LC-MS/MS) and measurements of LR density in the presence of analogues or inhibitors of auxin and SLs were performed. Furthermore, an untargeted RNA-sequencing (RNA-seq)-based approach was used to better characterize the participation of auxin and SLs to the transcriptional signature of maize root response to nitrate. Our results suggested that N deprivation induces zealactone and carlactonoic acid biosynthesis in root, to a higher extent if compared to P-deprived roots. Moreover, data on LR density led to hypothesize that the induction of LR development early occurring upon nitrate supply involves the inhibition of SL biosynthesis, but that the downstream target of SL shutdown, besides auxin, also includes additional unknown players. Furthermore, RNA-seq results provided a set of putative markers for the auxin- or SL-dependent action of nitrate, meanwhile also allowing to identify novel components of the molecular regulation of maize root response to nitrate. Globally, the existence of at least four different pathways was hypothesized: one dependent on auxin, a second one mediated by SLs, a third deriving from the SL-auxin interplay, and a last one attributable to nitrate itself through further downstream signals. Further work will be necessary to better assess the reliability of the model proposed.
Subject(s)
Heterocyclic Compounds, 3-Ring/metabolism , Indoleacetic Acids/metabolism , Lactones/metabolism , Nitrates/metabolism , Plant Roots/growth & development , Zea mays/growth & development , Gene Expression Regulation, Plant/drug effects , Germination , Hexanones/pharmacology , Nitrates/pharmacology , Nitrogen/metabolism , Orobanchaceae/metabolism , Plant Roots/drug effects , Plant Roots/metabolism , Sequence Analysis, RNA , Tandem Mass Spectrometry , Triazoles/pharmacology , Zea mays/drug effects , Zea mays/metabolismABSTRACT
The liverwort Marchantia polymorpha contains two isoforms of the plastid terminal oxidase (PTOX), an enzyme that catalyzes the reduction of oxygen to water using plastoquinol as substrate. Phylogenetic analyses showed that one isoform, here called MpPTOXa, is closely related to isoforms occurring in plants and some algae, while the other isoform, here called MpPTOXb, is closely related to the two isoforms occurring in Chlamydomonas reinhardtii. Mutants of each isoform were created in Marchantia polymorpha using CRISPR/Cas9 technology. While no obvious phenotype was found for these mutants, chlorophyll fluorescence analyses demonstrated that the plastoquinone pool was in a higher reduction state in both mutants. This was visible at the level of fluorescence measured in dark-adapted material and by post illumination fluorescence rise. These results suggest that both isoforms have a redundant function. However, when P700 oxidation and re-reduction was studied, differences between these two isoforms were observed. Furthermore, the mutant affected in MpPTOXb showed a slight alteration in the pigment composition, a higher non-photochemical quenching and a slightly lower electron transport rate through photosystem II. These differences may be explained either by differences in the enzymatic activities or by different activities attributed to preferential involvement of the two PTOX isoforms to either linear or cyclic electron flow.
Subject(s)
Chlamydomonas reinhardtii/enzymology , Hepatophyta/enzymology , Mutation , Oxidoreductases/metabolism , Photosynthetic Reaction Center Complex Proteins/metabolism , Chlamydomonas reinhardtii/genetics , Hepatophyta/genetics , Oxidation-Reduction , Oxidoreductases/genetics , Photosynthetic Reaction Center Complex Proteins/geneticsABSTRACT
Abscisic acid (ABA), a plant hormone synthesized from carotenoids, functions in seed germination and abiotic stress responses. ABA is derived from the cleavage of 9-cis-isomers of violaxanthin and neoxanthin, which are oxygenated carotenoids, also called xanthophylls. Although genes encoding enzymes responsible for most steps of the ABA biosynthesis pathway have been identified, enzymatic reactions leading to the production of these cis-isomers from trans-violaxanthin remain poorly understood. Two mutants that lack trans- and cis-neoxanthin, tomato (Solanum lycopersicum) neoxanthin-deficient1 (nxd1) and Arabidopsis (Arabidopsis thaliana) ABA-deficient4 (aba4), were identified previously, but only aba4 exhibited ABA-deficient phenotypes. No enzymatic activity was detected for ABA4 and NXD1 proteins, and their exact function remained unknown. To further investigate ABA4 and NXD1 function in Arabidopsis, we compared phenotypes of single and double mutants, and analyzed the effect of ABA4 overexpression on ABA and carotenoid accumulation in wild-type and mutant backgrounds. We provide convergent evidence that ABA4 is not only required for the formation of trans- and 9'-cis-neoxanthin from trans-violaxanthin, but also controls 9-cis-violaxanthin accumulation. While nxd1 produces high amounts of 9-cis-violaxanthin and ABA, aba4 nxd1 exhibits reduced levels in both leaves and seeds. Furthermore, ABA4 constitutive expression in nxd1 increases both 9-cis-violaxanthin and ABA accumulation. Subcellular localization of NXD1 protein in transient expression assays suggests that production of the NXD1-derived factor required for neoxanthin synthesis takes place in the cytosol. Finally, we postulate that ABA4, with additional unknown cofactor(s), is required for, or contributes to, trans-to-cis violaxanthin isomerase activity, producing both cis-xanthophyll precursors of ABA.
Subject(s)
Abscisic Acid/metabolism , Arabidopsis/genetics , Arabidopsis/metabolism , Biosynthetic Pathways/genetics , Dehydration/genetics , Dehydration/physiopathology , Xanthophylls/metabolism , Gene Expression Regulation, Plant , Genes, Plant , Genetic Variation , Genotype , Mutation , Phenotype , Stress, PhysiologicalABSTRACT
Plants produce a huge diversity of specialized metabolites (SM) throughout their life cycle that play important physiological and ecological functions. SM can protect plants and seeds against diseases, predators, and abiotic stresses, or support their interactions with beneficial or symbiotic organisms. They also have strong impacts on human nutrition and health. Despite this importance, the biosynthesis and biological functions of most of the SM remain elusive and their diversity and/or quantity have been reduced in most crops during domestication. Seeds present a large number of SM that are important for their physiological, agronomic, nutritional or industrial qualities and hence, provide interesting models for both studying biosynthesis and producing large amounts of specialized metabolites. For instance, phenolics are abundant and widely distributed in seeds. More specifically, flavonoid pathway has been instrumental for understanding environmental or developmental regulations of specialized metabolic pathways, at the molecular and cellular levels. Here, we summarize current knowledge on seed phenolics as model, and discuss how recent progresses in omics approaches could help to further characterize their diversity, regulations, and the underlying molecular mechanisms involved.
Subject(s)
Phenols/metabolism , Seeds/metabolism , Flavonoids/chemistry , Flavonoids/metabolism , Flavonoids/physiology , Metabolic Networks and Pathways , Phenols/chemistry , Seeds/physiologyABSTRACT
Abscisic acid (ABA) is an important hormone for seed development and germination whose physiological action is modulated by its endogenous levels. Cleavage of carotenoid precursors by 9-cis epoxycarotenoid dioxygenase (NCED) and inactivation of ABA by ABA 8'-hydroxylase (CYP707A) are key regulatory metabolic steps. In Arabidopsis (Arabidopsis thaliana), both enzymes are encoded by multigene families, having distinctive expression patterns. To evaluate the genome-wide impact of ABA deficiency in developing seeds at the maturation stage when dormancy is induced, we used a nced2569 quadruple mutant in which ABA deficiency is mostly restricted to seeds, thus limiting the impact of maternal defects on seed physiology. ABA content was very low in nced2569 seeds, similar to the severe mutant aba2; unexpectedly, ABA Glc ester was detected in aba2 seeds, suggesting the existence of an alternative metabolic route. Hormone content in nced2569 seeds compared with nced259 and wild type strongly suggested that specific expression of NCED6 in the endosperm is mainly responsible for ABA production. In accordance, transcriptome analyses revealed broad similarities in gene expression between nced2569 and either wild-type or nced259 developing seeds. Gene ontology enrichments revealed a large spectrum of ABA activation targets involved in reserve storage and desiccation tolerance, and repression of photosynthesis and cell cycle. Proteome and metabolome profiles in dry nced2569 seeds, compared with wild-type and cyp707a1a2 seeds, also highlighted an inhibitory role of ABA on remobilization of reserves, reactive oxygen species production, and protein oxidation. Down-regulation of these oxidative processes by ABA may have an essential role in dormancy control.
Subject(s)
Abscisic Acid/metabolism , Arabidopsis/growth & development , Arabidopsis/metabolism , Genomics , Seeds/growth & development , Seeds/metabolism , Arabidopsis/genetics , Arabidopsis Proteins/metabolism , Biosynthetic Pathways/genetics , Cell Cycle , Desiccation , Gene Expression Regulation, Plant , Metabolome , Mutation/genetics , Oxidation-Reduction , Photosynthesis , Plant Dormancy/genetics , RNA, Messenger/genetics , RNA, Messenger/metabolism , Seeds/genetics , Signal Transduction/genetics , Transcriptome/geneticsABSTRACT
Dormancy is an adaptive trait that blocks seed germination until the environmental conditions become favorable for subsequent vegetative plant growth. Seed dormancy is defined as the inability to germinate in favorable conditions. Dormancy is alleviated during after-ripening, a dry storage period, during which dormant (D) seeds unable to germinate become non-dormant (ND), able to germinate in a wide range of environmental conditions. The treatment of dormant seeds with ethylene (D/ET) promotes seed germination, and abscisic acid (ABA) treatment reduces non-dormant (ND/ABA) seed germination in sunflowers (Helianthus annuus). Metabolomic and transcriptomic studies have been performed during imbibition to compare germinating seeds (ND and D/ET) and low-germinating seeds (D and ND/ABA). A PCA analysis of the metabolites content showed that imbibition did not trigger a significant change during the first hours (3 and 15 h). The metabolic changes associated with germination capacity occurred at 24 h and were related to hexoses, as their content was higher in ND and D/ET and was reduced by ABA treatment. At the transcriptional level, a large number of genes were altered oppositely in germinating, compared to the low-germinating seeds. The metabolomic and transcriptomic results were integrated in the interpretation of the processes involved in germination. Our results show that ethylene treatment triggers molecular changes comparable to that of after-ripening treatment, concerning sugar metabolism and ABA signaling inhibition.
Subject(s)
Ethylenes/metabolism , Germination , Helianthus/growth & development , Plant Growth Regulators/metabolism , Seeds/growth & development , Gene Expression Regulation, Plant , Helianthus/genetics , Helianthus/metabolism , Metabolome , Plant Dormancy , Seeds/genetics , Seeds/metabolism , TranscriptomeABSTRACT
The demethylation of lignin in ionic liquids (ILs) was investigated by using pure lignin model monomers and dimers together with dioxane-isolated lignins from poplar, miscanthus, and maize. Different methylimidazolium ILs were compared and the samples were treated with two different heating processes: microwave irradiation and conventional heating in a sealed tube. The conversion yield and influence of the treatment on the lignin structure were assessed by 31 Pâ NMR spectroscopy, size-exclusion chromatography, and thioacidolysis. The acidic methylimidazolium IL [HMIM]Br was shown to be an effective combination of solvent and reagent for the demethylation and depolymerization of lignin. The relatively mild reaction conditions, the clean work-up, and the ability to reuse the IL makes the described procedure an attractive and new green method for the conversion of lignin to produce phenol-rich lignin oligomers.
Subject(s)
Green Chemistry Technology/methods , Imidazoles/chemistry , Indicators and Reagents/chemistry , Ionic Liquids/chemistry , Lignin/chemistry , Chromatography, Gel , Demethylation , Hydrolysis , Magnetic Resonance Spectroscopy/methods , Microwaves , Poaceae/chemistry , Polymerization , Populus/chemistry , Zea mays/chemistryABSTRACT
INTRODUCTION: Strigolactones (SLs) are important plant hormones. They are difficult to analyse because they occur in very small concentrations especially in comparison with other plant hormones and other substances can interfere with their detection. OBJECTIVE: To develop a procedure for the extraction, purification and quantification of SLs from plant roots. METHODOLOGY: Samples were prepared by extraction of plant root tissues with ethyl acetate. Then the extracts were further purified with silica column chromatography. The natural SLs in the final extracts were quantified using novel deuterium labelled SLs. The results of the methodology were compared with those of the procedure of Yoneyama and coworkers. RESULTS: This procedure required about 1-g root samples to detect and quantify simultaneously the SLs (orobanchyl acetate and fabacyl acetate) concentration with high reliability. CONCLUSION: A method was developed for determining endogenous fabacyl acetate and orobanchyl acetate in plant tissue based on novel deuterium labelled standards. A method of orobanchol quantification using a synthetic SL GR24 as internal standard was proposed. Copyright © 2017 John Wiley & Sons, Ltd.
Subject(s)
Chromatography, Liquid/methods , Deuterium , Isotope Labeling , Lactones/chemistry , Spectrometry, Mass, Electrospray Ionization/methods , Tandem Mass Spectrometry/methods , Pisum sativum/chemistry , Plant Roots/chemistry , Reproducibility of ResultsABSTRACT
The photoactive Orange Carotenoid Protein (OCP) photoprotects cyanobacteria cells by quenching singlet oxygen and excess excitation energy. Its N-terminal domain is the active part of the protein, and the C-terminal domain regulates the activity. Recently, the characteristics of a family of soluble carotenoid-binding proteins (Helical Carotenoid Proteins [HCPs]), paralogs of the N-terminal domain of OCP, were described. Bioinformatics studies also revealed the existence of genes coding for homologs of CTD. Here, we show that the latter genes encode carotenoid proteins (CTDHs). This family of proteins contains two subgroups with distinct characteristics. One CTDH of each clade was further characterized, and they proved to be very good singlet oxygen quenchers. When synthesized in Escherichia coli or Synechocystis PCC 6803, CTDHs formed dimers that share a carotenoid molecule and are able to transfer their carotenoid to apo-HCPs and apo-OCP. The CTDHs from clade 2 have a cysteine in position 103. A disulfide bond is easily formed between the monomers of the dimer preventing carotenoid transfer. This suggests that the transfer of the carotenoid could be redox regulated in clade 2 CTDH. We also demonstrate here that apo-OCPs and apo-CTDHs are able to take the carotenoid directly from membranes, while HCPs are unable to do so. HCPs need the presence of CTDH to become holo-proteins. We propose that, in cyanobacteria, the CTDHs are carotenoid donors to HCPs.
Subject(s)
Bacterial Proteins/chemistry , Bacterial Proteins/metabolism , Carotenoids/metabolism , Sequence Homology, Amino Acid , Synechocystis/metabolism , Amino Acid Sequence , Apoproteins/chemistry , Apoproteins/metabolism , Canthaxanthin/metabolism , Consensus Sequence , Escherichia coli/metabolism , Evolution, Molecular , Fluorescence , Models, Biological , Models, Molecular , Phylogeny , Protein Binding , Protein Domains , Protein Multimerization , Spectrum AnalysisABSTRACT
Indole glucosinolates (IGs) are plant secondary metabolites that are derived from the amino acid tryptophan. The product of Arabidopsis (Arabidopsis thaliana) IG core biosynthesis, indol-3-ylmethyl glucosinolate (I3M), can be modified by hydroxylation and subsequent methoxylation of the indole ring in position 1 (1-IG modification) or 4 (4-IG modification). Products of the 4-IG modification pathway mediate plant-enemy interactions and are particularly important for Arabidopsis innate immunity. While CYP81Fs encoding cytochrome P450 monooxygenases and IGMTs encoding indole glucosinolate O-methyltransferases have been identified as key genes for IG modification, our knowledge about the IG modification pathways is not complete. In particular, it is unknown which enzyme is responsible for methyl transfer in the 1-IG modification pathway and whether this pathway plays a role in defense, similar to 4-IG modification. Here, we analyze two Arabidopsis transfer DNA insertion lines with targeted metabolomics. We show that biosynthesis of 1-methoxyindol-3-ylmethyl glucosinolate (1MOI3M) from I3M involves the predicted unstable intermediate 1-hydroxyindol-3-ylmethyl glucosinolate (1OHI3M) and that IGMT5, a gene with moderate similarity to previously characterized IGMTs, encodes the methyltransferase that is responsible for the conversion of 1OHI3M to 1MOI3M. Disruption of IGMT5 function increases resistance against the root-knot nematode Meloidogyne javanica and suggests a potential role for the 1-IG modification pathway in Arabidopsis belowground defense.
Subject(s)
Arabidopsis Proteins/metabolism , Arabidopsis/enzymology , Biosynthetic Pathways , Glucosinolates/biosynthesis , Methyltransferases/metabolism , Animals , Arabidopsis/genetics , Arabidopsis/parasitology , DNA, Bacterial/genetics , Disease Resistance/genetics , Gene Expression Regulation, Plant , Genes, Plant , Mass Spectrometry , Metabolome/genetics , Methylation , Mutagenesis, Insertional/genetics , Mutation/genetics , Plant Diseases/genetics , Plant Diseases/microbiology , Plant Diseases/parasitology , Plant Roots/enzymology , Plant Roots/genetics , Plant Tumors/parasitology , Promoter Regions, Genetic/genetics , Tylenchoidea/physiologyABSTRACT
Using a phylogenomic approach, we have identified and subclassified a new family of carotenoid-binding proteins. These proteins have sequence homology to the N-terminal domain (NTD) of the Orange Carotenoid Protein (OCP), and are referred to as Helical Carotenoid Proteins (HCPs). These proteins comprise at least nine distinct clades and are found in diverse organisms, frequently as multiple paralogs representing the distinct clades. These seem to be out-paralogs maintained from ancient duplications associated with subfunctionalization. All of the HCPs share conservation of the residues for carotenoid binding, and we confirm that carotenoid binding is a fundamental property of HCPs. We solved two crystal structures of the Nostoc sp. PCC 7120 HCP1 protein, each binding a different carotenoid, suggesting that the proteins flexibly bind a range of carotenoids. Based on a comprehensive phylogenetic analysis, we propose that one of the HCP subtypes is likely the evolutionary ancestor of the NTD of the OCP, which arose following a domain fusion event. However, we predict that the majority of HCPs have functions distinct from the NTD of the OCP. Our results demonstrate that the HCPs are a new family of functionally diverse carotenoid-binding proteins found among ecophysiologically diverse cyanobacteria.
Subject(s)
Bacterial Proteins/metabolism , Carotenoids/metabolism , Cyanobacteria/metabolism , Bacterial Proteins/classification , Bacterial Proteins/genetics , Cyanobacteria/genetics , Evolution, Molecular , PhylogenyABSTRACT
The photoactive Orange Carotenoid Protein (OCP) is involved in cyanobacterial photoprotection. Its N-terminal domain (NTD) is responsible for interaction with the antenna and induction of excitation energy quenching, while the C-terminal domain is the regulatory domain that senses light and induces photoactivation. In most nitrogen-fixing cyanobacterial strains, there are one to four paralogous genes coding for homologs to the NTD of the OCP. The functions of these proteins are unknown. Here, we study the expression, localization, and function of these genes in Anabaena sp. PCC 7120. We show that the four genes present in the genome are expressed in both vegetative cells and heterocysts but do not seem to have an essential role in heterocyst formation. This study establishes that all four Anabaena NTD-like proteins can bind a carotenoid and the different paralogs have distinct functions. Surprisingly, only one paralog (All4941) was able to interact with the antenna and to induce permanent thermal energy dissipation. Two of the other Anabaena paralogs (All3221 and Alr4783) were shown to be very good singlet oxygen quenchers. The fourth paralog (All1123) does not seem to be involved in photoprotection. Structural homology modeling allowed us to propose specific features responsible for the different functions of these soluble carotenoid-binding proteins.
Subject(s)
Anabaena/metabolism , Bacterial Proteins/genetics , Bacterial Proteins/metabolism , Anabaena/genetics , Bacterial Proteins/chemistry , Carotenoids/metabolism , Electron Spin Resonance Spectroscopy , Escherichia coli/genetics , Fluorescence , Gene Expression Regulation, Bacterial , Green Fluorescent Proteins/genetics , Green Fluorescent Proteins/metabolism , Phycobilisomes/chemistry , Phycobilisomes/metabolism , Protein DomainsABSTRACT
Cell wall remodeling is an essential mechanism for the regulation of plant growth and architecture, and xyloglucans (XyGs), the major hemicellulose, are often considered as spacers of cellulose microfibrils during growth. In the seed, the activity of cell wall enzymes plays a critical role in germination by enabling embryo cell expansion leading to radicle protrusion, as well as endosperm weakening prior to its rupture. A screen for Arabidopsis (Arabidopsis thaliana) mutants affected in the hormonal control of germination identified a mutant, xyl1, able to germinate on paclobutrazol, an inhibitor of gibberellin biosynthesis. This mutant also exhibited reduced dormancy and increased resistance to high temperature. The XYL1 locus encodes an α-xylosidase required for XyG maturation through the trimming of Xyl. The xyl1 mutant phenotypes were associated with modifications to endosperm cell wall composition that likely impact on its resistance, as further demonstrated by the restoration of normal germination characteristics by endosperm-specific XYL1 expression. The absence of phenotypes in mutants defective for other glycosidases, which trim Gal or Fuc, suggests that XYL1 plays the major role in this process. Finally, the decreased XyG abundance in hypocotyl longitudinal cell walls of germinating embryos indicates a potential role in cell wall loosening and anisotropic growth together with pectin de-methylesterification.
Subject(s)
Arabidopsis/growth & development , Arabidopsis/metabolism , Glucans/metabolism , Xylans/metabolism , Arabidopsis/genetics , Arabidopsis Proteins/genetics , Arabidopsis Proteins/metabolism , Cell Wall/metabolism , Endosperm/growth & development , Endosperm/metabolism , Gene Expression Regulation, Developmental , Gene Expression Regulation, Plant , Genes, Plant , Germination/drug effects , Germination/genetics , Germination/physiology , Mutation , Plants, Genetically Modified , Protein Processing, Post-Translational , Seeds/growth & development , Seeds/metabolism , Triazoles/pharmacology , Xylosidases/genetics , Xylosidases/metabolismABSTRACT
The plastid terminal oxidase PTOX is a plastohydroquinone:oxygen oxidoreductase that is important for carotenoid biosynthesis and plastid development. Its role in photosynthesis is controversially discussed. Under a number of abiotic stress conditions, the protein level of PTOX increases. PTOX is thought to act as a safety valve under high light protecting the photosynthetic apparatus against photodamage. However, transformants with high PTOX level were reported to suffer from photoinhibition. To analyze the effect of PTOX on the photosynthetic electron transport, tobacco expressing PTOX-1 from Chlamydomonas reinhardtii (Cr-PTOX1) was studied by chlorophyll fluorescence, thermoluminescence, P700 absorption kinetics and CO2 assimilation. Cr-PTOX1 was shown to compete very efficiently with the photosynthetic electron transport for PQH2 . High pressure liquid chromatography (HPLC) analysis confirmed that the PQ pool was highly oxidized in the transformant. Immunoblots showed that, in the wild-type, PTOX was associated with the thylakoid membrane only at a relatively alkaline pH value while it was detached from the membrane at neutral pH. We present a model proposing that PTOX associates with the membrane and oxidizes PQH2 only when the oxidation of PQH2 by the cytochrome b6 f complex is limiting forward electron transport due to a high proton gradient across the thylakoid membrane.