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1.
Int J Mol Sci ; 22(19)2021 Sep 28.
Artigo em Inglês | MEDLINE | ID: mdl-34638787

RESUMO

High night temperatures (HNT) affect rice yield in the field and induce chlorosis symptoms in leaves in controlled chamber experiments. However, little is known about molecular changes in leaf segments under these conditions. Transcript and metabolite profiling were performed for leaf segments of six rice cultivars with different HNT sensitivity. The metabolite profile of the sheath revealed a lower metabolite abundance compared to segments of the leaf blade. Furthermore, pre-adaptation to stress under control conditions was detected in the sheath, whereas this segment was only slightly affected by HNT. No unique significant transcriptomic changes were observed in the leaf base, including the basal growth zone at HNT conditions. Instead, selected metabolites showed correlations with HNT sensitivity in the base. The middle part and the tip were most highly affected by HNT in sensitive cultivars on the transcriptomic level with higher expression of jasmonic acid signaling related genes, genes encoding enzymes involved in flavonoid metabolism and a gene encoding galactinol synthase. In addition, gene expression of expansins known to improve stress tolerance increased in tolerant and sensitive cultivars. The investigation of the different leaf segments indicated highly segment specific responses to HNT. Molecular key players for HNT sensitivity were identified.

2.
Plant Physiol ; 2021 Sep 07.
Artigo em Inglês | MEDLINE | ID: mdl-34623449

RESUMO

The high-value carotenoid astaxanthin (3,3'-dihydroxy-ß,ß-carotene-4,4'-dione) is one of the most potent antioxidants in nature. In addition to its large-scale use in fish farming, the pigment has applications as a food supplement and an active ingredient in cosmetics and in pharmaceuticals for the treatment of diseases linked to reactive oxygen species. The biochemical pathway for astaxanthin synthesis has been introduced into seed plants, which do not naturally synthesize this pigment, by nuclear and plastid engineering. The highest accumulation rates have been achieved in transplastomic plants, but massive production of astaxanthin has resulted in severe growth retardation. What limits astaxanthin accumulation levels and what causes the mutant phenotype is unknown. Here, we addressed these questions by making astaxanthin synthesis in tobacco (Nicotiana tabacum) plastids inducible by a synthetic riboswitch. We show that, already in the uninduced state, astaxanthin accumulates to similarly high levels as in transplastomic plants expressing the pathway constitutively. Importantly, the inducible plants displayed wild-type-like growth properties and riboswitch induction resulted in a further increase in astaxanthin accumulation. Our data suggest that the mutant phenotype associated with constitutive astaxanthin synthesis is due to massive metabolite turnover, and indicate that astaxanthin accumulation is limited by the sequestration capacity of the plastid.

3.
Plant J ; 2021 Sep 06.
Artigo em Inglês | MEDLINE | ID: mdl-34486764

RESUMO

13 C-Metabolic flux analysis (13 C-MFA) has greatly contributed to our understanding of plant metabolic regulation. However, the generation of detailed in vivo flux maps remains a major challenge. Flux investigations based on nuclear magnetic resonance have resolved small networks with high accuracy. Mass spectrometry (MS) approaches have broader potential, but have hitherto been limited in their power to deduce flux information due to lack of atomic level position information. Herein we established a gas chromatography (GC) coupled to MS-based approach that provides 13 C-positional labelling information in glucose, malate and glutamate (Glu). A map of electron impact (EI)-mediated MS fragmentation was created and validated by 13 C-positionally labelled references via GC-EI-MS and GC-atmospheric pressure chemical ionization-MS technologies. The power of the approach was revealed by analysing previous 13 C-MFA data from leaves and guard cells, and 13 C-HCO3 labelling of guard cells harvested in the dark and after the dark-to-light transition. We demonstrated that the approach is applicable to established GC-EI-MS-based 13 C-MFA without the need for experimental adjustment, but will benefit in the future from paired analyses by the two GC-MS platforms. We identified specific glucose carbon atoms that are preferentially labelled by photosynthesis and gluconeogenesis, and provide an approach to investigate the phosphoenolpyruvate carboxylase (PEPc)-derived 13 C-incorporation into malate and Glu. Our results suggest that gluconeogenesis and the PEPc-mediated CO2 assimilation into malate are activated in a light-independent manner in guard cells. We further highlight that the fluxes from glycolysis and PEPc toward Glu are restricted by the mitochondrial thioredoxin system in illuminated leaves.

4.
Int J Mol Sci ; 22(15)2021 Jul 29.
Artigo em Inglês | MEDLINE | ID: mdl-34360938

RESUMO

During seed germination, desiccation tolerance is lost in the radicle with progressing radicle protrusion and seedling establishment. This process is accompanied by comprehensive changes in the metabolome and proteome. Germination of Arabidopsis seeds was investigated over 72 h with special focus on the heat-stable proteome including late embryogenesis abundant (LEA) proteins together with changes in primary metabolites. Six metabolites in dry seeds known to be important for seed longevity decreased during germination and seedling establishment, while all other metabolites increased simultaneously with activation of growth and development. Thermo-stable proteins were associated with a multitude of biological processes. In the heat-stable proteome, a relatively similar proportion of fully ordered and fully intrinsically disordered proteins (IDP) was discovered. Highly disordered proteins were found to be associated with functional categories development, protein, RNA and stress. As expected, the majority of LEA proteins decreased during germination and seedling establishment. However, four germination-specific dehydrins were identified, not present in dry seeds. A network analysis of proteins, metabolites and amino acids generated during the course of germination revealed a highly connected LEA protein network.


Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis , Germinação , Proteínas de Plantas/metabolismo , Proteoma/metabolismo , Plântula/metabolismo , Arabidopsis/crescimento & desenvolvimento , Arabidopsis/metabolismo , Regulação da Expressão Gênica de Plantas , Temperatura Alta
5.
Int J Mol Sci ; 22(11)2021 Jun 07.
Artigo em Inglês | MEDLINE | ID: mdl-34200118

RESUMO

Drought represents a major abiotic stress factor negatively affecting growth, yield and tuber quality of potatoes. Quantitative trait locus (QTL) analyses were performed in cultivated potatoes for drought tolerance index DRYM (deviation of relative starch yield from the experimental median), tuber starch content, tuber starch yield, tuber fresh weight, selected transcripts and metabolites under control and drought stress conditions. Eight genomic regions of major interest for drought tolerance were identified, three representing standalone DRYM QTL. Candidate genes, e.g., from signaling pathways for ethylene, abscisic acid and brassinosteroids, and genes encoding cell wall remodeling enzymes were identified within DRYM QTL. Co-localizations of DRYM QTL and QTL for tuber starch content, tuber starch yield and tuber fresh weight with underlying genes of the carbohydrate metabolism were observed. Overlaps of DRYM QTL with metabolite QTL for ribitol or galactinol may indicate trade-offs between starch and compatible solute biosynthesis. Expression QTL confirmed the drought stress relevance of selected transcripts by overlaps with DRYM QTL. Bulked segregant analyses combined with next-generation sequencing (BSAseq) were used to identify mutations in genes under the DRYM QTL on linkage group 3. Future analyses of identified genes for drought tolerance will give a better insight into drought tolerance in potatoes.


Assuntos
Cromossomos de Plantas/genética , Secas , Genoma de Planta , Polimorfismo de Nucleotídeo Único , Locos de Características Quantitativas , Solanum tuberosum/genética , Tetraploidia , Mapeamento Cromossômico , Ligação Genética , Genômica , Fenótipo , Tubérculos/genética , Solanum tuberosum/fisiologia
6.
Int J Mol Sci ; 22(11)2021 Jun 07.
Artigo em Inglês | MEDLINE | ID: mdl-34200446

RESUMO

Ribosome biogenesis is essential for plants to successfully acclimate to low temperature. Without dedicated steps supervising the 60S large subunits (LSUs) maturation in the cytosol, e.g., Rei-like (REIL) factors, plants fail to accumulate dry weight and fail to grow at suboptimal low temperatures. Around REIL, the final 60S cytosolic maturation steps include proofreading and assembly of functional ribosomal centers such as the polypeptide exit tunnel and the P-Stalk, respectively. In consequence, these ribosomal substructures and their assembly, especially during low temperatures, might be changed and provoke the need for dedicated quality controls. To test this, we blocked ribosome maturation during cold acclimation using two independent reil double mutant genotypes and tested changes in their ribosomal proteomes. Additionally, we normalized our mutant datasets using as a blank the cold responsiveness of a wild-type Arabidopsis genotype. This allowed us to neglect any reil-specific effects that may happen due to the presence or absence of the factor during LSU cytosolic maturation, thus allowing us to test for cold-induced changes that happen in the early nucleolar biogenesis. As a result, we report that cold acclimation triggers a reprogramming in the structural ribosomal proteome. The reprogramming alters the abundance of specific RP families and/or paralogs in non-translational LSU and translational polysome fractions, a phenomenon known as substoichiometry. Next, we tested whether the cold-substoichiometry was spatially confined to specific regions of the complex. In terms of RP proteoforms, we report that remodeling of ribosomes after a cold stimulus is significantly constrained to the polypeptide exit tunnel (PET), i.e., REIL factor binding and functional site. In terms of RP transcripts, cold acclimation induces changes in RP families or paralogs that are significantly constrained to the P-Stalk and the ribosomal head. The three modulated substructures represent possible targets of mechanisms that may constrain translation by controlled ribosome heterogeneity. We propose that non-random ribosome heterogeneity controlled by specialized biogenesis mechanisms may contribute to a preferential or ultimately even rigorous selection of transcripts needed for rapid proteome shifts and successful acclimation.


Assuntos
Aclimatação , Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Temperatura Baixa , Proteoma/metabolismo , Proteínas Ribossômicas/metabolismo , Ribossomos/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Proteoma/análise , Proteínas Ribossômicas/genética , Ribossomos/genética
7.
J Exp Bot ; 72(18): 6544-6569, 2021 09 30.
Artigo em Inglês | MEDLINE | ID: mdl-34106234

RESUMO

Grapevine (Vitis vinifera) berries are extremely sensitive to infection by the biotrophic pathogen Erysiphe necator, causing powdery mildew disease with deleterious effects on grape and wine quality. The combined analysis of the transcriptome and metabolome associated with this common fungal infection has not been previously carried out in any fruit. In order to identify the molecular, hormonal, and metabolic mechanisms associated with infection, healthy and naturally infected V. vinifera cv. Carignan berries were collected at two developmental stages: late green (EL33) and early véraison (EL35). RNA sequencing combined with GC-electron impact ionization time-of-flight MS, GC-electron impact ionization/quadrupole MS, and LC-tandem MS analyses revealed that powdery mildew-susceptible grape berries were able to activate defensive mechanisms with the involvement of salicylic acid and jasmonates and to accumulate defense-associated metabolites (e.g. phenylpropanoids, fatty acids). The defensive strategies also indicated organ-specific responses, namely the activation of fatty acid biosynthesis. However, defense responses were not enough to restrict fungal growth. The fungal metabolic program during infection involves secretion of effectors related to effector-triggered susceptibility, carbohydrate-active enzymes and activation of sugar, fatty acid, and nitrogen uptake, and could be under epigenetic regulation. This study also identified potential metabolic biomarkers such as gallic, eicosanoic, and docosanoic acids and resveratrol, which can be used to monitor early stages of infection.

8.
Genes (Basel) ; 12(4)2021 03 28.
Artigo em Inglês | MEDLINE | ID: mdl-33800602

RESUMO

Potato is regarded as drought sensitive and most vulnerable to climate changes. Its cultivation in drought prone regions or under conditions of more frequent drought periods, especially in subtropical areas, requires intensive research to improve drought tolerance in order to guarantee high yields under limited water supplies. A candidate gene approach was used to develop functional simple sequence repeat (SSR) markers for association studies in potato with the aim to enhance breeding for drought tolerance. SSR primer combinations, mostly surrounding interrupted complex and compound repeats, were derived from 103 candidate genes for drought tolerance. Validation of the SSRs was performed in an association panel representing 34 mainly starch potato cultivars. Seventy-five out of 154 SSR primer combinations (49%) resulted in polymorphic, highly reproducible banding patterns with polymorphic information content (PIC) values between 0.11 and 0.90. Five SSR markers identified allelic differences between the potato cultivars that showed significant associations with drought sensitivity. In all cases, the group of drought-sensitive cultivars showed predominantly an additional allele, indicating that selection against these alleles by marker-assisted breeding might confer drought tolerance. Further studies of these differences in the candidate genes will elucidate their role for an improved performance of potatoes under water-limited conditions.


Assuntos
Repetições de Microssatélites , Solanum tuberosum/fisiologia , Estresse Fisiológico , Biologia Computacional/métodos , DNA de Plantas/genética , Secas , Estudos de Associação Genética , Melhoramento Vegetal , Solanum tuberosum/genética
9.
Plant Cell Environ ; 44(7): 2034-2048, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-33764557

RESUMO

Asymmetric warming resulting in a faster increase in night compared to day temperatures affects crop yields negatively. Physiological characterization and agronomic findings have been complemented more recently by molecular biology approaches including transcriptomic, proteomic, metabolomic and lipidomic investigations in crops exposed to high night temperature (HNT) conditions. Nevertheless, the understanding of the underlying mechanisms causing yield decline under HNT is still limited. The discovery of significant differences between HNT-tolerant and HNT-sensitive cultivars is one of the main research directions to secure continuous food supply under the challenge of increasing climate change. With this review, we provide a summary of current knowledge on the physiological and molecular basis of contrasting HNT tolerance in rice and wheat cultivars. Requirements for HNT tolerance and the special adaptation strategies of the HNT-tolerant rice cultivar Nagina-22 (N22) are discussed. Putative metabolite markers for HNT tolerance useful for marker-assisted breeding are suggested, together with future research directions aimed at improving food security under HNT conditions.

10.
Commun Biol ; 4(1): 181, 2021 02 10.
Artigo em Inglês | MEDLINE | ID: mdl-33568709

RESUMO

Protein-metabolite interactions are of crucial importance for all cellular processes but remain understudied. Here, we applied a biochemical approach named PROMIS, to address the complexity of the protein-small molecule interactome in the model yeast Saccharomyces cerevisiae. By doing so, we provide a unique dataset, which can be queried for interactions between 74 small molecules and 3982 proteins using a user-friendly interface available at https://promis.mpimp-golm.mpg.de/yeastpmi/ . By interpolating PROMIS with the list of predicted protein-metabolite interactions, we provided experimental validation for 225 binding events. Remarkably, of the 74 small molecules co-eluting with proteins, 36 were proteogenic dipeptides. Targeted analysis of a representative dipeptide, Ser-Leu, revealed numerous protein interactors comprising chaperones, proteasomal subunits, and metabolic enzymes. We could further demonstrate that Ser-Leu binding increases activity of a glycolytic enzyme phosphoglycerate kinase (Pgk1). Consistent with the binding analysis, Ser-Leu supplementation leads to the acute metabolic changes and delays timing of a diauxic shift. Supported by the dipeptide accumulation analysis our work attests to the role of Ser-Leu as a metabolic regulator at the interface of protein degradation and central metabolism.


Assuntos
Metabolismo Energético , Fosfoglicerato Quinase/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/enzimologia , Glicólise , Metaboloma , Metabolômica , Fosfoglicerato Quinase/genética , Mapas de Interação de Proteínas , Proteólise , Proteoma , Proteômica , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/genética
11.
Sci Rep ; 11(1): 4787, 2021 Feb 26.
Artigo em Inglês | MEDLINE | ID: mdl-33637852

RESUMO

Ribosome biogenesis is tightly associated to plant metabolism due to the usage of ribosomes in the synthesis of proteins necessary to drive metabolic pathways. Given the central role of ribosome biogenesis in cell physiology, it is important to characterize the impact of different components involved in this process on plant metabolism. Double mutants of the Arabidopsis thaliana cytosolic 60S maturation factors REIL1 and REIL2 do not resume growth after shift to moderate 10 [Formula: see text] chilling conditions. To gain mechanistic insights into the metabolic effects of this ribosome biogenesis defect on metabolism, we developed TC-iReMet2, a constraint-based modelling approach that integrates relative metabolomics and transcriptomics time-course data to predict differential fluxes on a genome-scale level. We employed TC-iReMet2 with metabolomics and transcriptomics data from the Arabidopsis Columbia 0 wild type and the reil1-1 reil2-1 double mutant before and after cold shift. We identified reactions and pathways that are highly altered in a mutant relative to the wild type. These pathways include the Calvin-Benson cycle, photorespiration, gluconeogenesis, and glycolysis. Our findings also indicated differential NAD(P)/NAD(P)H ratios after cold shift. TC-iReMet2 allows for mechanistic hypothesis generation and interpretation of system biology experiments related to metabolic fluxes on a genome-scale level.

12.
Sci Rep ; 11(1): 2410, 2021 01 28.
Artigo em Inglês | MEDLINE | ID: mdl-33510206

RESUMO

Arabidopsis REIL proteins are cytosolic ribosomal 60S-biogenesis factors. After shift to 10 °C, reil mutants deplete and slowly replenish non-translating eukaryotic ribosome complexes of root tissue, while controlling the balance of non-translating 40S- and 60S-subunits. Reil mutations respond by hyper-accumulation of non-translating subunits at steady-state temperature; after cold-shift, a KCl-sensitive 80S sub-fraction remains depleted. We infer that Arabidopsis may buffer fluctuating translation by pre-existing non-translating ribosomes before de novo synthesis meets temperature-induced demands. Reil1 reil2 double mutants accumulate 43S-preinitiation and pre-60S-maturation complexes and alter paralog composition of ribosomal proteins in non-translating complexes. With few exceptions, e.g. RPL3B and RPL24C, these changes are not under transcriptional control. Our study suggests requirement of de novo synthesis of eukaryotic ribosomes for long-term cold acclimation, feedback control of NUC2 and eIF3C2 transcription and links new proteins, AT1G03250, AT5G60530, to plant ribosome biogenesis. We propose that Arabidopsis requires biosynthesis of specialized ribosomes for cold acclimation.


Assuntos
Aclimatação , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Arabidopsis/fisiologia , Temperatura Baixa , Regulação da Expressão Gênica de Plantas , Ribossomos/metabolismo , Citosol/metabolismo , Mutação , Especificidade de Órgãos , Desenvolvimento Vegetal/genética , Biossíntese de Proteínas , Proteômica/métodos , Subunidades Ribossômicas/metabolismo , Transcrição Genética
13.
Curr Opin Plant Biol ; 61: 102002, 2021 06.
Artigo em Inglês | MEDLINE | ID: mdl-33497897

RESUMO

Chromatin structure influences DNA accessibility and underlying gene expression. Disturbances of chromatin structure often result in pleiotropic developmental phenotypes. Interactions between chromatin modifications and development have been the main focus of epigenetic studies. Recent years brought major advance in uncovering and understanding connections between chromatin organisation in the nucleus and metabolic processes that take place in the cytoplasm or other cellular compartments. Products of primary metabolism and cell redox states influence chromatin-modifying complexes, and chromatin modifiers in turn affect expression of metabolic genes. Current evidence indicates that complex interaction loops between these biological system layers exist. Applying interdisciplinary and holistic approaches will decipher causality and molecular mechanisms of the dynamic crosstalk between chromatin structure, metabolism and plant growth and development.


Assuntos
Cromatina , Epigênese Genética , Cromatina/genética , DNA , Desenvolvimento Vegetal/genética , Plantas/genética
14.
Plant Cell Environ ; 44(3): 915-930, 2021 03.
Artigo em Inglês | MEDLINE | ID: mdl-33190295

RESUMO

Freezing triggers extracellular ice formation leading to cell dehydration and deformation during a freeze-thaw cycle. Many plant species increase their freezing tolerance during exposure to low, non-freezing temperatures, a process termed cold acclimation. In addition, exposure to mild freezing temperatures after cold acclimation evokes a further increase in freezing tolerance (sub-zero acclimation). Previous transcriptome and proteome analyses indicate that cell wall remodelling may be particularly important for sub-zero acclimation. In the present study, we used a combination of immunohistochemical, chemical and spectroscopic analyses to characterize the cell walls of Arabidopsis thaliana and characterized a mutant in the XTH19 gene, encoding a xyloglucan endotransglucosylase/hydrolase (XTH). The mutant showed reduced freezing tolerance after both cold and sub-zero acclimation, compared to the Col-0 wild type, which was associated with differences in cell wall composition and structure. Most strikingly, immunohistochemistry in combination with 3D reconstruction of centres of rosette indicated that epitopes of the xyloglucan-specific antibody LM25 were highly abundant in the vasculature of Col-0 plants after sub-zero acclimation but absent in the XTH19 mutant. Taken together, our data shed new light on the potential roles of cell wall remodelling for the increased freezing tolerance observed after low temperature acclimation.

15.
Plant Cell Environ ; 44(3): 870-884, 2021 03.
Artigo em Inglês | MEDLINE | ID: mdl-33251628

RESUMO

Stomatal movements are enabled by changes in guard cell turgor facilitated via transient accumulation of inorganic and organic ions imported from the apoplast or biosynthesized within guard cells. Under salinity, excess salt ions accumulate within plant tissues resulting in osmotic and ionic stress. To elucidate whether (a) Na+ and Cl- concentrations increase in guard cells in response to long-term NaCl exposure and how (b) guard cell metabolism acclimates to the anticipated stress, we profiled the ions and primary metabolites of leaves, the apoplast and isolated guard cells at darkness and during light, that is, closed and fully opened stomata. In contrast to leaves, the primary metabolism of guard cell preparations remained predominantly unaffected by increased salt ion concentrations. Orchestrated reductions of stomatal aperture and guard cell osmolyte synthesis were found, but unlike in leaves, no increases of stress responsive metabolites or compatible solutes occurred. Diverging regulation of guard cell metabolism might be a prerequisite to facilitate the constant adjustment of turgor that affects aperture. Moreover, the photoperiod-dependent sucrose accumulation in the apoplast and guard cells changed to a permanently replete condition under NaCl, indicating that stress-related photosynthate accumulation in leaves contributes to the permanent closing response of stomata under stress.

16.
Elife ; 92020 08 24.
Artigo em Inglês | MEDLINE | ID: mdl-32831171

RESUMO

The promiscuous activities of enzymes provide fertile ground for the evolution of new metabolic pathways. Here, we systematically explore the ability of E. coli to harness underground metabolism to compensate for the deletion of an essential biosynthetic pathway. By deleting all threonine deaminases, we generated a strain in which isoleucine biosynthesis was interrupted at the level of 2-ketobutyrate. Incubation of this strain under aerobic conditions resulted in the emergence of a novel 2-ketobutyrate biosynthesis pathway based upon the promiscuous cleavage of O-succinyl-L-homoserine by cystathionine γ-synthase (MetB). Under anaerobic conditions, pyruvate formate-lyase enabled 2-ketobutyrate biosynthesis from propionyl-CoA and formate. Surprisingly, we found this anaerobic route to provide a substantial fraction of isoleucine in a wild-type strain when propionate is available in the medium. This study demonstrates the selective advantage underground metabolism offers, providing metabolic redundancy and flexibility which allow for the best use of environmental carbon sources.


Assuntos
Butiratos/metabolismo , Carbono-Oxigênio Liases/metabolismo , Escherichia coli/metabolismo , Deleção de Genes , Homosserina/análogos & derivados , Isoleucina/metabolismo , Escherichia coli/genética , Homosserina/metabolismo , Redes e Vias Metabólicas
17.
Front Plant Sci ; 11: 1071, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32793257

RESUMO

Climate models predict an increased likelihood of drought, demanding efficient selection for drought tolerance to maintain yield stability. Classic tolerance breeding relies on selection for yield in arid environments, which depends on yield trials and takes decades. Breeding could be accelerated by marker-assisted selection (MAS). As an alternative to genomic markers, transcript and metabolite markers have been suggested for important crops but also for orphan corps. For potato, we suggested a random-forest-based model that predicts tolerance from leaf metabolite and transcript levels with a precision of more than 90% independent of the agro-environment. To find out how the model based selection compares to yield-based selection in arid environments, we applied this approach to a population of 200 tetraploid Solanum tuberosum ssp. tuberosum lines segregating for drought tolerance. Twenty-four lines were selected into a phenotypic subpopulation (PPt) for superior tolerance based on relative tuber starch yield data from three drought stress trials. Two subpopulations with superior (MPt) and inferior (MPs) tolerance were selected based on drought tolerance predictions based on leaf metabolite and transcript levels from two sites. The 60 selected lines were phenotyped for yield and drought tolerance in 10 multi-environment drought stress trials representing typical Central European drought scenarios. Neither selection affected development or yield potential. Lines with superior drought tolerance and high yields under stress were over-represented in both populations selected for superior tolerance, with a higher number in PPt compared to MPt. However, selection based on leaf metabolites may still be an alternative to yield-based selection in arid environments as it works on leaves sampled in breeder's fields independent of drought trials. As the selection against low tolerance was ineffective, the method is best used in combination with tools that select against sensitive genotypes. Thus, metabolic and transcript marker-based selection for drought tolerance is a viable alternative to the selection on yield in arid environments.

18.
Front Plant Sci ; 11: 1118, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32793268

RESUMO

Staple crops in human and livestock diets suffer from deficiencies in certain "essential" amino acids including methionine. With the goal of increasing methionine in rice seed, we generated a pair of "Push × Pull" double transgenic lines, each containing a methionine-dense seed storage protein (2S albumin from sunflower, HaSSA) and an exogenous enzyme for either methionine (feedback desensitized cystathionine gamma synthase from Arabidopsis, AtD-CGS) or cysteine (serine acetyltransferase from E. coli, EcSAT) biosynthesis. In both double transgenic lines, the total seed methionine content was approximately 50% higher than in their untransformed parental line, Oryza sativa ssp. japonica cv. Taipei 309. HaSSA-containing rice seeds were reported to display an altered seed protein profile, speculatively due to insufficient sulfur amino acid content. However, here we present data suggesting that this may result from an overloaded protein folding machinery in the endoplasmic reticulum rather than primarily from redistribution of limited methionine from endogenous seed proteins to HaSSA. We hypothesize that HaSSA-associated endoplasmic reticulum stress results in redox perturbations that negatively impact sulfate reduction to cysteine, and we speculate that this is mitigated by EcSAT-associated increased sulfur import into the seed, which facilitates additional synthesis of cysteine and glutathione. The data presented here reveal challenges associated with increasing the methionine content in rice seed, including what may be relatively low protein folding capacity in the endoplasmic reticulum and an insufficient pool of sulfate available for additional cysteine and methionine synthesis. We propose that future approaches to further improve the methionine content in rice should focus on increasing seed sulfur loading and avoiding the accumulation of unfolded proteins in the endoplasmic reticulum. Oryza sativa ssp. japonica: urn:lsid:ipni.org:names:60471378-2.

19.
Nat Plants ; 6(8): 1031-1043, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32719473

RESUMO

The unparalleled performance of Chlorella ohadii under irradiances of twice full sunlight underlines the gaps in our understanding of how the photosynthetic machinery operates, and what sets its upper functional limit. Rather than succumbing to photodamage under extreme irradiance, unique features of photosystem II function allow C. ohadii to maintain high rates of photosynthesis and growth, accompanied by major changes in composition and cellular structure. This remarkable resilience allowed us to investigate the systems response of photosynthesis and growth to extreme illumination in a metabolically active cell. Using redox proteomics, transcriptomics, metabolomics and lipidomics, we explored the cellular mechanisms that promote dissipation of excess redox energy, protein S-glutathionylation, inorganic carbon concentration, lipid and starch accumulation, and thylakoid stacking. C. ohadii possesses a readily available capacity to utilize a sudden excess of reducing power and carbon for growth and reserve formation, and post-translational redox regulation plays a pivotal role in this rapid response. Frequently the response in C. ohadii deviated from that of model species, reflecting its life history in desert sand crusts. Comparative global and case-specific analyses provided insights into the potential evolutionary role of effective reductant utilization in this extreme resistance of C. ohadii to extreme irradiation.


Assuntos
Chlorella/metabolismo , Proteínas de Algas/metabolismo , Proteínas de Algas/fisiologia , Chlorella/fisiologia , Chlorella/efeitos da radiação , Clima Desértico , Perfilação da Expressão Gênica , Lipidômica , Metabolômica , Oxirredução/efeitos da radiação , Fotossíntese , Complexo de Proteína do Fotossistema II/metabolismo , Complexo de Proteína do Fotossistema II/fisiologia , Proteômica
20.
Methods Mol Biol ; 2156: 203-239, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32607984

RESUMO

This book chapter describes the analytical procedures required for the profiling of a metabolite fraction enriched for primary metabolites. The profiling is based on routine gas chromatography coupled to mass spectrometry (GC-MS). The generic profiling method is adapted to plant material, specifically to the analysis of plant material that was exposed to temperature stress. The method can be combined with stable isotope labeling and tracing experiments and is equally applicable to preparations of plant material and microbial photosynthetic organisms. The described methods are modular and can be multiplexed, that is, the same sample or a paired identical backup sample can be analyzed sequentially by more than one of the described procedures. The modules include rapid sampling and metabolic inactivation protocols for samples in a wide weight range, sample extraction procedures, chemical derivatization steps that are required to make the metabolite fraction amenable to gas chromatographic analysis, routine GC-MS methods, and procedures of data processing and data mining. A basic and extendable set of standardizations for metabolite recovery and retention index alignment of the resulting GC-MS chromatograms is included. The methods have two applications: (1) The rapid screening for changes of relative metabolite pools sizes under temperature stress and (2) the verification by exact quantification using GC-MS protocols that are extended by internal and external standardization.


Assuntos
Cromatografia Gasosa-Espectrometria de Massas , Metaboloma , Metabolômica , Fenômenos Fisiológicos Vegetais , Plantas/metabolismo , Temperatura , Análise de Dados , Cromatografia Gasosa-Espectrometria de Massas/métodos , Marcação por Isótopo , Metabolômica/métodos , Fenótipo , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz
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