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1.
Plant Sci ; 291: 110336, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31928684

RESUMO

Wheat grain nitrogen content displays large variations within different pearling fractions of grains because of radial gradients in the protein content. We identified how spatiotemporal mechanisms regulate this. The protein gradients emerged clearly at 19 days after anthesis, with the highest N content in aleurone and seed coat, followed by outer endosperm, whereas the lowest was in middle and inner endosperm. Laser microdissection, qRT-PCR and LC-MS were used to dissect tissue from aleurone, outer endosperm, middle endosperm, inner endosperm and transfer cells, measure gene expression and levels of free and protein-bound amino acids, respectively. The results showed that different FAA transportation pathways worked in parallel during grain filling stage while the grain protein gradient did not follow spatial expression of storage proteins. Additionally, two nitrogen (N) topdressing timings were conducted, either at the emergence of top third leaf (standard timing) or top first leaf (delayed timing), finding that delayed N topdressing enhanced both amino acids supply and protein synthesis capacity. The results provide insight into protein synthesis and amino acid transport pathways in endosperm and suggest targets for the enhancement of specialty pearled wheat with higher quality.


Assuntos
Aminoácidos/metabolismo , Endosperma/química , Proteínas de Plantas/metabolismo , Sementes/química , Triticum/genética , Endosperma/crescimento & desenvolvimento , Endosperma/metabolismo , Triticum/química , Triticum/metabolismo
2.
J Exp Bot ; 71(1): 234-246, 2020 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-31494665

RESUMO

The biosynthesis of starch granules in plant plastids is coordinated by the orchestrated action of transferases, hydrolases, and dikinases. These enzymes either contain starch-binding domain(s) themselves, or are dependent on direct interactions with co-factors containing starch-binding domains. As a means to competitively interfere with existing starch-protein interactions, we expressed the protein module Carbohydrate-Binding Motif 20 (CBM20), which has a very high affinity for starch, ectopically in barley plastids. This interference resulted in an increase in the number of starch granules in chloroplasts and in formation of compound starch granules in grain amyloplasts, which is unusual for barley. More importantly, we observed a photosystem-independent inhibition of CO2 fixation, with a subsequent reduced growth rate and lower accumulation of carbohydrates with effects throughout the metabolome, including lower accumulation of transient leaf starch. Our results demonstrate the importance of endogenous starch-protein interactions for controlling starch granule morphology and number, and plant growth, as substantiated by a metabolic link between starch-protein interactions and control of CO2 fixation in chloroplasts.

3.
J Exp Bot ; 70(17): 4521-4537, 2019 08 29.
Artigo em Inglês | MEDLINE | ID: mdl-31245808

RESUMO

To investigate the effect of high atmospheric NO concentrations on crop plants and the role of phytoglobins under these conditions, we performed a long-term study on barley 'Golden Promise' wild type (WT), class 1 phytoglobin knockdown (HvPgb1.1-) and class 1 phytoglobin overexpression (HvPgb1.1+) lines. Plants were cultivated with nitrogen-free nutrient solution during the entire growth period and were fumigated with different NO concentration (ambient, 800, 1500, and 3000 ppb). Analysis of fresh weight, stem number, chlorophyll content, and effective quantum yield of PSII showed that NO fumigation promoted plant growth and tillering significantly in the HvPgb1.1+ line. After 80 d of NO fumigation, dry matter weight, spikes number, kernel number, and plant kernel weight were significantly increased in HvPgb1.1+ plants with increasing NO concentration. In contrast, yield decreased in WT and HvPgb1.1- plants the higher the NO level. Application of atmospheric 15NO and 15NO2 demonstrated NO specificity of phytoglobins. 15N from 15NO could be detected in RNA, DNA, and proteins of barley leaves and the 15N levels were significantly higher in HvPgb1.1+ plants in comparison with HvPgb1.1- and WT plants. Our results demonstrate that overexpression of phytoglobins allows plants to more efficiently use atmospheric NO as N source.


Assuntos
Regulação da Expressão Gênica de Plantas , Hordeum/genética , Óxido Nítrico/metabolismo , Proteínas de Plantas/genética , Hordeum/metabolismo , Proteínas de Plantas/metabolismo
4.
J Exp Bot ; 70(2): 485-496, 2019 01 07.
Artigo em Inglês | MEDLINE | ID: mdl-30407538

RESUMO

Plant starch is the main energy contributor to the human diet. Its biosynthesis is catalyzed and regulated by co-ordinated actions of several enzymes. Recently, a factor termed Protein Targeting to Starch 1 (PTST1) was identified as being required for correct granule-bound starch synthase (GBSS) localization and demonstrated to be crucial for amylose synthesis in Arabidopsis. However, the function of its homologous protein in storage tissues (e.g. endosperm) is unknown. We identified a PTST1 homolog in barley and it was found to contain a crucial coiled-coil domain and carbohydrate-binding module. We demonstrated the interaction between PTST1 and GBSS1 by fluorescence resonance energy transfer (FRET) in barley endosperm. By tagging PTST1 with the fluorophore mCherry, we observed that it is localized in the stroma of barley endosperm amyloplasts. PTST1 overexpression in endosperm increased endogenous gbss1a gene expression and amylose content. Gbss1a and ptst1 mutants were generated using clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-related protein 9 (Cas9)-based targeted mutagenesis. Homozygous gbss1a mutants showed a waxy phenotype. Grains of ptst1 mutants did not accumulate any starch. These grains dried out during the desiccation stage and were unable to germinate, suggesting that PTST1 is essential for development of starchy endosperm and viable grains.


Assuntos
Endosperma/crescimento & desenvolvimento , Hordeum/metabolismo , Proteínas de Plantas/metabolismo , Amido/biossíntese , Cloroplastos/metabolismo , Grão Comestível/crescimento & desenvolvimento , Endosperma/metabolismo , Hordeum/genética , Hordeum/crescimento & desenvolvimento , Fenótipo , Folhas de Planta/metabolismo
5.
Food Chem ; 277: 135-144, 2019 Mar 30.
Artigo em Inglês | MEDLINE | ID: mdl-30502129

RESUMO

Nitrogen fertilization regimes significantly affect both grain quality and yield. Wheat plants were subjected to different application timing of topdressed nitrogen at the emergence of the top fifth (TL5), top third (TL3) and top first leaf (TL1), respectively. The iTRAQ (isobaric tag for relative and absolute quantitation) technology was adopted to obtain the complete proteome of wheat flour and to identify the differentially expressed proteins (DEPs) as regulated by nitrogen topdressing timing. Collectively, 591 proteins into 17 functional categories in flour of mature grains were identified. In comparison to TL3, 50 and 63 DEPs were identified in TL5 and TL1, respectively. Nine of the DEPs commonly dependent on nitrogen topdressing timing are the γ-gliadins or high-molecular-weight glutenin subunits. Additionally, delaying nitrogen topdressing modified the grain hardness and allergic protein content. The results suggested that altering nitrogen topdressing timing is a potential strategy for pursuing targeted processing quality of wheat flour.


Assuntos
Grão Comestível/efeitos dos fármacos , Grão Comestível/metabolismo , Glutens/metabolismo , Dureza/efeitos dos fármacos , Nitrogênio/farmacologia , Farinha/análise , Qualidade dos Alimentos , Folhas de Planta/metabolismo , Proteômica , Fatores de Tempo
6.
BMC Plant Biol ; 18(1): 353, 2018 Dec 13.
Artigo em Inglês | MEDLINE | ID: mdl-30545290

RESUMO

BACKGROUND: Nitrogen is one basic element of amino acids and grain protein in wheat. In field experiments, wheat plants were subjected to different timing of nitrogen topdressing treatments: at the stages of emergence of the top fifth leaf (TL5), top third leaf (TL3) and top first leaf (TL1) to test the regulatory effects of nitrogen topdressing timing on grain protein quality. The underlying mechanisms were elucidated by clarifying the relationship between proteolysis in vegetative organs and accumulation of amino acids in the endosperm cavity, conversion of amino acids, and storage protein synthesis in endosperm of wheat grain. RESULTS: Delayed nitrogen topdressing up-regulated gene expression related to nitrogen metabolism and protease synthesis in the flag leaf, followed by more free amino acids being transported to both the cavity and the endosperm from 7 days after anthesis (DAA) to 13 DAA in TL1. TL1 enhanced the conversion between free amino acids in endosperm and upregulated the expression of genes encoding high molecular weight (HMW) and low molecular weight (LMW) subunits and protein disulfide isomerases-like (PDIL) proteins, indicating that the synthesis and folding of glutenin were enhanched by delayed nitrogen topdressing. As a consequense, the content of glutenin macropolymers (GMP) and glutenin increased with delaying nitrogen topdressing. CONCLUSIONS: The results highlight the relationship between nitrogen remobilization and final grain protein production and suggest that the nitrogen remobilization processes could be a potential target for improving the quality of wheat grain. Additionally, specific gene expression related to nitrogen topdressing was identified, which conferred more detailed insights into underlying mechanism on the modification protein quality.


Assuntos
Aminoácidos/metabolismo , Grão Comestível/metabolismo , Nitrogênio/metabolismo , Proteínas de Plantas/metabolismo , Triticum/metabolismo , Aminoácidos/análise , Grão Comestível/química , Endosperma/química , Endosperma/metabolismo , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Nitrogênio/administração & dosagem , Folhas de Planta/metabolismo , Reação em Cadeia da Polimerase em Tempo Real
7.
Plant Sci ; 256: 1-4, 2017 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-28167022

RESUMO

Grasses such as wheat and barley constitute some of the main crops currently cultivated worldwide. Cultivation of wild cereals such as Hordeum spontaneum (wild barley) and Triticum diccocoides (wild emmer) started in southwest Asia. Despite the many studies on the origins of agriculture and plant domestication, surprisingly few studies have discussed the importance of the nutritional quality of barley and emmer wheat in their evolution. A comparison of domesticated forms of these cereals with their wild progenitors suggests an evolution towards bigger grains with higher glycemic loads in the form of higher relative starch content with lower relative protein, fiber and mineral contents. In this work we hypothesize that in addition to the simple explanation that larger grains emerged under domestication, complex and indirect effects such as increased glycemic index and sweet taste should also be taken into consideration.


Assuntos
Evolução Biológica , Dieta , Domesticação , Grão Comestível/química , Hordeum/química , Valor Nutritivo , Triticum/química , Agricultura , Produtos Agrícolas , Índice Glicêmico , Paladar
8.
Plant Cell Environ ; 40(1): 36-50, 2017 01.
Artigo em Inglês | MEDLINE | ID: mdl-27245884

RESUMO

Nitric oxide (NO) is an important signalling molecule that is involved in many different physiological processes in plants. Here, we report about a NO-fixing mechanism in Arabidopsis, which allows the fixation of atmospheric NO into nitrogen metabolism. We fumigated Arabidopsis plants cultivated in soil or as hydroponic cultures during the whole growing period with up to 3 ppmv of NO gas. Transcriptomic, proteomic and metabolomic analyses were used to identify non-symbiotic haemoglobin proteins as key components of the NO-fixing process. Overexpressing non-symbiotic haemoglobin 1 or 2 genes resulted in fourfold higher nitrate levels in these plants compared with NO-treated wild-type. Correspondingly, rosettes size and weight, vegetative shoot thickness and seed yield were 25, 40, 30, and 50% higher, respectively, than in wild-type plants. Fumigation with 250 ppbv 15 NO confirmed the importance of non-symbiotic haemoglobin 1 and 2 for the NO-fixation pathway, and we calculated a daily uptake for non-symbiotic haemoglobin 2 overexpressing plants of 250 mg N/kg dry weight. This mechanism is probably important under conditions with limited N supply via the soil. Moreover, the plant-based NO uptake lowers the concentration of insanitary atmospheric NOx, and in this context, NO-fixation can be beneficial to air quality.


Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Hemoglobinas/metabolismo , Óxido Nítrico/metabolismo , Nitrogênio/farmacologia , Simbiose , Amônia/metabolismo , Arabidopsis/efeitos dos fármacos , Arabidopsis/genética , Arabidopsis/crescimento & desenvolvimento , Fumigação , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Nitratos/metabolismo , Óxido Nítrico/farmacologia , Nitritos/metabolismo , Fenótipo , Folhas de Planta/efeitos dos fármacos , Folhas de Planta/metabolismo , Propanóis/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , S-Nitrosotióis/metabolismo
9.
Planta ; 227(4): 917-27, 2008 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-18034356

RESUMO

Plants contain three classes of hemoglobin genes of which two, class 1 and class 2, have a structure similar to classical vertebrate globins. We investigated the effect of silencing the class 1 non-symbiotic hemoglobin gene, GLB1, and the effect of overexpression of GLB1 or the class 2 non-symbiotic hemoglobin gene, GLB2, in Arabidopsis thaliana. Lines with GLB1 silencing had a significant delay of bolting and after bolting, shoots reverted to the rosette vegetative phase by formation of aerial rosettes at lateral meristems. Lines with overexpression of GLB1 or GLB2 bolted earlier than wild type plants. By germinating the lines in a medium containing the nitric oxide (NO) donor, sodium nitroprusside (SNP), it was demonstrated that both GLB1 and GLB2 promote bolting by antagonizing the effect of NO, suggesting that non-symbiotic plant hemoglobin controls bolting by scavenging the floral transition signal molecule, NO. So far, NO scavenging has only been demonstrated for class 1 non-symbiotic hemoglobins. A direct assay in Arabidopsis leaf cells shows that GLB1 as well as the class 2 non-symbiotic hemoglobin, GLB2, scavenge NO in vivo. NO has also been demonstrated to be a growth stimulating signal with an optimum at low concentrations. It was observed that overexpression of either GLB1 or GLB2 shifts the optimum for NO growth stimulation to a higher concentration. In conclusion, we have found that expression of NO scavenging plant hemoglobin is involved in the control of bolting in Arabidopsis.


Assuntos
Proteínas de Arabidopsis/genética , Arabidopsis/genética , Hemoglobinas/genética , Óxido Nítrico/metabolismo , Arabidopsis/crescimento & desenvolvimento , Arabidopsis/metabolismo , Proteínas de Arabidopsis/metabolismo , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Genótipo , Hemoglobinas/metabolismo , Nitroprussiato/farmacologia , Fenótipo , Plantas Geneticamente Modificadas , Reação em Cadeia da Polimerase Via Transcriptase Reversa
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