Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 2.507
Filtrar
1.
Int J Mol Sci ; 22(15)2021 Jul 24.
Artigo em Inglês | MEDLINE | ID: mdl-34360673

RESUMO

Auxin is a key regulator of plant development affecting the formation and maturation of reproductive structures. The apoplastic route of auxin transport engages influx and efflux facilitators from the PIN, AUX and ABCB families. The polar localization of these proteins and constant recycling from the plasma membrane to endosomes is dependent on Rab-mediated vesicular traffic. Rab proteins are anchored to membranes via posttranslational addition of two geranylgeranyl moieties by the Rab Geranylgeranyl Transferase enzyme (RGT), which consists of RGTA, RGTB and REP subunits. Here, we present data showing that seed development in the rgtb1 mutant, with decreased vesicular transport capacity, is disturbed. Both pre- and post-fertilization events are affected, leading to a decrease in seed yield. Pollen tube recognition at the stigma and its guidance to the micropyle is compromised and the seed coat forms incorrectly. Excess auxin in the sporophytic tissues of the ovule in the rgtb1 plants leads to an increased tendency of autonomous endosperm formation in unfertilized ovules and influences embryo development in a maternal sporophytic manner. The results show the importance of vesicular traffic for sexual reproduction in flowering plants, and highlight RGTB1 as a key component of sporophytic-filial signaling.


Assuntos
Arabidopsis/enzimologia , Sementes/enzimologia , Transdução de Sinais , Alquil e Aril Transferases/metabolismo , Alquil e Aril Transferases/fisiologia , Arabidopsis/crescimento & desenvolvimento , Arabidopsis/metabolismo , Transporte Biológico , Regulação da Expressão Gênica de Plantas , Ácidos Indolacéticos/metabolismo , Mutação , Tubo Polínico/fisiologia , Sementes/crescimento & desenvolvimento , Sementes/metabolismo
2.
Int J Mol Sci ; 22(16)2021 Aug 20.
Artigo em Inglês | MEDLINE | ID: mdl-34445686

RESUMO

Quinoin is a type 1 ribosome-inactivating protein (RIP) we previously isolated from the seeds of pseudocereal quinoa (Chenopodium quinoa) and is known as a functional food for its beneficial effects on human health. As the presence of RIPs in edible plants could be potentially risky, here we further characterised biochemically the protein (complete amino acid sequence, homologies/differences with other RIPs and three-dimensional homology modeling) and explored its possible defensive role against pathogens. Quinoin consists of 254 amino acid residues, without cysteinyl residues. As demonstrated by similarities and homology modeling, quinoin preserves the amino acid residues of the active site (Tyr75, Tyr122, Glu177, Arg180, Phe181 and Trp206; quinoin numbering) and the RIP-fold characteristic of RIPs. The polypeptide chain of quinoin contains two N-glycosylation sites at Asn115 and Asp231, the second of which appears to be linked to sugars. Moreover, by comparative MALDI-TOF tryptic peptide mapping, two differently glycosylated forms of quinoin, named pre-quinoin-1 and pre-quinoin-2 (~0.11 mg/100 g and ~0.85 mg/100 g of seeds, respectively) were characterised. Finally, quinoin possesses: (i) strong antiviral activity, both in vitro and in vivo towards Tobacco Necrosis Virus (TNV); (ii) a growth inhibition effect on the bacterial pathogens of plants; and (iii) a slight antifungal effect against two Cryphonectria parasitica strains.


Assuntos
Chenopodium quinoa/enzimologia , Saporinas/metabolismo , Sequência de Aminoácidos/genética , Chenopodium quinoa/metabolismo , Proteínas de Plantas/metabolismo , Inibidores da Síntese de Proteínas/farmacologia , Ribossomos/metabolismo , Saporinas/fisiologia , Sementes/enzimologia , Homologia de Sequência de Aminoácidos
3.
Int J Mol Sci ; 22(15)2021 Jul 29.
Artigo em Inglês | MEDLINE | ID: mdl-34360902

RESUMO

Acyl-CoA:lysophosphatidylethanolamine acyltransferases (LPEATs) are known as enzymes utilizing acyl-CoAs and lysophospholipids to produce phosphatidylethanolamine. Recently, it has been discovered that they are also involved in the growth regulation of Arabidopsis thaliana. In our study we investigated expression of each Camelina sativa LPEAT isoform and their behavior in response to temperature changes. In order to conduct a more extensive biochemical evaluation we focused both on LPEAT enzymes present in microsomal fractions from C. sativa plant tissues, and on cloned CsLPEAT isoforms expressed in yeast system. Phylogenetic analyses revealed that CsLPEAT1c and CsLPEAT2c originated from Camelina hispida, whereas other isoforms originated from Camelina neglecta. The expression ratio of all CsLPEAT1 isoforms to all CsLPEAT2 isoforms was higher in seeds than in other tissues. The isoforms also displayed divergent substrate specificities in utilization of LPE; CsLPEAT1 preferred 18:1-LPE, whereas CsLPEAT2 preferred 18:2-LPE. Unlike CsLPEAT1, CsLPEAT2 isoforms were specific towards very-long-chain fatty acids. Above all, we discovered that temperature strongly regulates LPEATs activity and substrate specificity towards different acyl donors, making LPEATs sort of a sensor of external thermal changes. We observed the presented findings not only for LPEAT activity in plant-derived microsomal fractions, but also for yeast-expressed individual CsLPEAT isoforms.


Assuntos
Aciltransferases/metabolismo , Camellia/enzimologia , Camellia/genética , Fosfatidiletanolaminas/metabolismo , Proteínas de Plantas/metabolismo , Sementes/enzimologia , Temperatura , Acil Coenzima A/metabolismo , Aciltransferases/genética , Camellia/classificação , Camellia/crescimento & desenvolvimento , Resposta ao Choque Frio , DNA de Plantas/genética , Ativação Enzimática , Resposta ao Choque Térmico , Isoenzimas/genética , Microssomos/enzimologia , Filogenia , Proteínas de Plantas/genética , Sementes/crescimento & desenvolvimento , Especificidade por Substrato
4.
J Plant Physiol ; 265: 153494, 2021 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-34454370

RESUMO

Raffinose, stachyose and verbascose form the three major members of the raffinose family oligosaccharides (RFO) accumulated during seed development. Raffinose synthase (RS; EC 2.4.1.82) and stachyose synthase (STS; EC 2.4.1.67) have been associated with raffinose and stachyose synthesis, but the precise mechanism for verbascose synthesis is not well understood. In this study, full-length RS (2.7 kb) and STS (2.6 kb) clones were isolated by screening a cDNA library prepared from developing lentil seeds (18, 20, 22 and 24 days after flowering [DAF]) to understand the roles of RS and STS in RFO accumulation in developing lentil seeds. The nucleotide sequences of RS and STS genes were similar to those reported for Pisum sativum. Patterns of transcript accumulation, enzyme activities and RFO concentrations were also comparable to P. sativum. However, during lentil seed development raffinose, stachyose and verbascose accumulation corresponded to transcript accumulation for RS and STS, with peak transcript abundance occurring at about 22-24 DAF, generally followed by a sequential increase in raffinose, stachyose and verbascose concentrations followed by a steady level thereafter. Enzyme activities for RS, STS and verbascose synthase (VS) also indicated a sudden increase at around 24-26 DAF, but with an abrupt decline again coinciding with the subsequent steady state increase in the RFO. Galactan:galactan galactosyl transferase (GGT), the galactinol-independent pathway enzyme, however, exhibited steady increase in activity from 24 DAF onwards before abruptly decreasing at 34 DAF. Although GGT activity was detected, isolation of a GGT sequence from the cDNA library was not successful.


Assuntos
Galactosiltransferases/genética , Galactosiltransferases/metabolismo , Lens (Planta)/enzimologia , Lens (Planta)/genética , Oligossacarídeos/biossíntese , Rafinose/biossíntese , Sementes/crescimento & desenvolvimento , Regulação da Expressão Gênica de Plantas , Genes de Plantas , Lens (Planta)/crescimento & desenvolvimento , Oligossacarídeos/genética , Rafinose/genética , Sementes/enzimologia , Sementes/genética
5.
Food Chem ; 361: 130031, 2021 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-34058661

RESUMO

Pearl millet is considered as 'nutri-cereal' because of high nutrient density of the seeds. The grain has limited use because of low keeping quality of the flour due to the activities of rancidity causing enzymes like lipase, lox, pox and PPO. Among all the enzymes, lipase is most notorious because of its robust nature and high activity under different conditions. we have identified 2180 putative transcripts showing homology with different variants of lipase precursor through transcriptome data mining (NCBI BioProject acc. no. PRJNA625418). Lipase plays dual role of facilitating the germination of seeds and deteriorating the quality of the pearl millet flour through hydrolytic rancidity. Different physiochemical methods like heat treatment, micro oven, hydrothermal, etc. have been developed to inhibit lipase activity in pearl millet flour. There is further need to develop improved processing technologies to inhibit the hydrolytic and oxidative rancidity in the floor with enhanced shelf-life.


Assuntos
Armazenamento de Alimentos , Germinação , Lipase/metabolismo , Pennisetum/enzimologia , Sementes/enzimologia , Farinha , Manipulação de Alimentos , Lipase/fisiologia , Pennisetum/fisiologia , Sementes/fisiologia
6.
Plant Cell Rep ; 40(7): 1215-1228, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-34028583

RESUMO

KEY MESSAGE: Among the 113 lipases present in rice genome, bran and endosperm-specific lipases were identified and lipase activity for one of the selected lipase gene is demonstrated in yeast. Rice bran is nutritionally superior than endosperm as it has major reservoirs of various minerals, vitamins, essential mineral oils and other bioactive compounds, however it is often under-utilized as a food product due to bran instability after milling. Various hydrolytic enzymes, such as lipases, present in bran causes degradation of the lipids present and are responsible for the bran instability. Here, in this study, we have systematically analyzed the 113 lipase genes present in rice genome, and identified 21 seed-specific lipases. By analyzing the expression of these genes in different seed tissues during seed development, we have identified three bran-specific and three endosperm-specific lipases, and one lipase which expresses in both bran and endosperm tissues. Further analysis of these genes during seed maturation and seed germination revealed that their expression increases during seed maturation and decreases during seed germination. Finally, we have shown the lipase activity for one of the selected genes, LOC_Os05g30900, in heterologous system yeast. The bran-specific lipases identified in this study would be very valuable for engineering designer rice varieties having increased bran stability in post-milling.


Assuntos
Lipase/genética , Lipase/metabolismo , Oryza/enzimologia , Simulação por Computador , Perfilação da Expressão Gênica , Regulação Enzimológica da Expressão Gênica , Regulação da Expressão Gênica de Plantas , Germinação , Oryza/genética , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Sementes/enzimologia , Sementes/genética , Sementes/crescimento & desenvolvimento , Leveduras/genética , Leveduras/metabolismo
7.
BMC Plant Biol ; 21(1): 245, 2021 May 29.
Artigo em Inglês | MEDLINE | ID: mdl-34051740

RESUMO

BACKGROUND: Arabinogalactan-proteins (AGPs) are heavily glycosylated with type II arabinogalactan (AG) polysaccharides attached to hydroxyproline residues in their protein backbone. Type II AGs are necessary for plant growth and critically important for the establishment of normal cellular functions. Despite the importance of type II AGs in plant development, our understanding of the underlying role of these glycans/sugar residues in mucilage formation and seed coat epidermal cell development is poorly understood and far from complete. One such sugar residue is the glucuronic acid residues of AGPs that are transferred onto AGP glycans by the action of ß-glucuronosyltransferase genes/enzymes. RESULTS: Here, we have characterized two ß-glucuronosyltransferase genes, GLCAT14A and GLCAT14C, that are involved in the transfer of ß-glucuronic acid (GlcA) to type II AGs. Using a reverse genetics approach, we observed that glcat14a-1 mutants displayed subtle alterations in mucilage pectin homogalacturonan (HG) compared to wild type (WT), while glcat14a-1glcat14c-1 mutants displayed much more severe mucilage phenotypes, including loss of adherent mucilage and significant alterations in cellulose ray formation and seed coat morphology. Monosaccharide composition analysis showed significant alterations in the sugar amounts of glcat14a-1glcat14c-1 mutants relative to WT in the adherent and non-adherent seed mucilage. Also, a reduction in total mucilage content was observed in glcat14a-1glcat14c-1 mutants relative to WT. In addition, glcat14a-1glcat14c-1 mutants showed defects in pectin formation, calcium content and the degree of pectin methyl-esterification (DM) as well as reductions in crystalline cellulose content and seed size. CONCLUSIONS: These results raise important questions regarding cell wall polymer interactions and organization during mucilage formation. We propose that the enzymatic activities of GLCAT14A and GLCAT14C play partially redundant roles and are required for the organization of the mucilage matrix and seed size in Arabidopsis thaliana. This work brings us a step closer towards identifying potential gene targets for engineering plant cell walls for industrial applications.


Assuntos
Arabidopsis/enzimologia , Galactanos/metabolismo , Glucuronosiltransferase/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Cálcio/metabolismo , Parede Celular/metabolismo , Celulose/metabolismo , Esterificação , Glucuronosiltransferase/genética , Monossacarídeos/metabolismo , Pectinas/metabolismo , Fenótipo , Polissacarídeos/metabolismo , Sementes/enzimologia , Sementes/genética
8.
Int J Mol Sci ; 22(5)2021 Mar 06.
Artigo em Inglês | MEDLINE | ID: mdl-33800919

RESUMO

Trypsin inhibitors (TI), a common anti-nutritional factor in soybean, prevent animals' protein digestibility reducing animal growth performance. No commercial soybean cultivars with low or null concentration of TI are available. The availability of a high throughput genotyping assay will be beneficial to incorporate the low TI trait into elite breeding lines. The aim of this study is to develop and validate a breeder friendly Kompetitive Allele Specific PCR (KASP) assay linked to low Kunitz trypsin inhibitor (KTI) in soybean seeds. A total of 200 F3:5 lines derived from PI 547656 (low KTI) X Glenn (normal KTI) were genotyped using the BARCSoySNP6K_v2 Beadchip. F3:4 and F3:5 lines were grown in Blacksburg and Orange, Virginia in three years, respectively, and were measured for KTI content using a quantitative HPLC method. We identified three SNP markers tightly linked to the major QTL associated to low KTI in the mapping population. Based on these SNPs, we developed and validated the KASP assays in a set of 93 diverse germplasm accessions. The marker Gm08_44814503 has 86% selection efficiency for the accessions with low KTI and could be used in marker assisted breeding to facilitate the incorporation of low KTI content in soybean seeds.


Assuntos
Genes de Plantas , Melhoramento Vegetal , Reação em Cadeia da Polimerase/métodos , Polimorfismo de Nucleotídeo Único , Locos de Características Quantitativas , Sementes/enzimologia , Soja/genética , Inibidor da Tripsina de Soja de Kunitz/genética , Alelos , Cromatografia Líquida de Alta Pressão/métodos , DNA de Plantas/análise , DNA de Plantas/genética , Ligação Genética , Fenótipo , Folhas de Planta/química , Soja/enzimologia , Inibidor da Tripsina de Soja de Kunitz/análise
9.
Int J Mol Sci ; 22(6)2021 Mar 11.
Artigo em Inglês | MEDLINE | ID: mdl-33799521

RESUMO

Climate change, environmental pollution and pathogen resistance to available chemical agents are part of the problems that the food industry has to face in order to ensure healthy food for people and livestock. One of the promising solutions to these problems is the use of cold atmospheric pressure plasma (CAPP). Plasma is suitable for efficient surface decontamination of seeds and food products, germination enhancement and obtaining higher yields in agricultural production. However, the plasma effects vary due to plasma source, treatment conditions and seed type. In our study, we tried to find the proper conditions for treatment of barley grains by diffuse coplanar surface barrier discharge, in which positive effects of CAPP, such as enhanced germination or decontamination effects, would be maximized and harmful effects, such as oxidation and genotoxic potential, minimized. Besides germination parameters, we evaluated DNA damage and activities of various germination and antioxidant enzymes in barley seedlings. Plasma exposure resulted in changes in germination parameters and enzyme activities. Longer exposures had also genotoxic effects. As such, our findings indicate that appropriate plasma exposure conditions need to be carefully optimized in order to preserve germination, oxidation balance and genome stability, should CAPP be used in agricultural practice.


Assuntos
Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Germinação/efeitos dos fármacos , Hordeum/efeitos dos fármacos , Gases em Plasma/farmacologia , Plântula/efeitos dos fármacos , Sementes/efeitos dos fármacos , Dano ao DNA , DNA de Plantas/genética , DNA de Plantas/metabolismo , Hordeum/enzimologia , Hordeum/genética , Hordeum/crescimento & desenvolvimento , Oxirredução , Estresse Oxidativo , Peroxidase/genética , Peroxidase/metabolismo , Raízes de Plantas , Brotos de Planta , Plântula/enzimologia , Plântula/genética , Plântula/crescimento & desenvolvimento , Sementes/enzimologia , Sementes/genética , Sementes/crescimento & desenvolvimento , Superóxido Dismutase/genética , Superóxido Dismutase/metabolismo
10.
Plant Cell Physiol ; 62(5): 798-814, 2021 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-33693904

RESUMO

In Arabidopsis thaliana, two genes encode the E2 subunit of the 2-oxoglutarate dehydrogenase (2-OGDH), a multimeric complex composed of three subunits. To functionally characterize the isoforms of E2 subunit, we isolated Arabidopsis mutant lines for each gene encoding the E2 subunit and performed a detailed molecular and physiological characterization of the plants under controlled growth conditions. The functional lack of expression of E2 subunit isoforms of 2-OGDH increased plant growth, reduced dark respiration and altered carbohydrate metabolism without changes in the photosynthetic rate. Interestingly, plants from e2-ogdh lines also exhibited reduced seed weight without alterations in total seed number. We additionally observed that downregulation of 2-OGDH activity led to minor changes in the levels of tricarboxylic acid cycle intermediates without clear correlation with the reduced expression of specific E2-OGDH isoforms. Furthermore, the e2-ogdh mutant lines exhibited a reduction by up to 25% in the leaf total amino acids without consistent changes in the amino acid profile. Taken together, our results indicate that the two isoforms of E2 subunit play a similar role in carbon-nitrogen metabolism, in plant growth and in seed weight.


Assuntos
Arabidopsis/fisiologia , Carbono/metabolismo , Complexo Cetoglutarato Desidrogenase/metabolismo , Nitrogênio/metabolismo , Arabidopsis/crescimento & desenvolvimento , Regulação para Baixo , Regulação da Expressão Gênica de Plantas , Germinação , Complexo Cetoglutarato Desidrogenase/genética , Fotossíntese , Filogenia , Subunidades Proteicas , Plântula/genética , Plântula/crescimento & desenvolvimento , Plântula/metabolismo , Sementes/enzimologia , Sementes/crescimento & desenvolvimento
11.
BMC Plant Biol ; 21(1): 58, 2021 Jan 22.
Artigo em Inglês | MEDLINE | ID: mdl-33482732

RESUMO

BACKGROUND: Physical seed dormancy is an important trait in legume domestication. Although seed dormancy is beneficial in wild ecosystems, it is generally considered to be an undesirable trait in crops due to reduction in yield and / or quality. The physiological mechanism and underlying genetic factor(s) of seed dormancy is largely unknown in several legume species. Here we employed an integrative approach to understand the mechanisms controlling physical seed dormancy in common bean (Phaseolus vulgaris L.). RESULTS: Using an innovative CT scan imaging system, we were able to track water movements inside the seed coat. We found that water uptake initiates from the bean seed lens. Using a scanning electron microscopy (SEM) we further identified several micro-cracks on the lens surface of non-dormant bean genotypes. Bulked segregant analysis (BSA) was conducted on a bi-parental RIL (recombinant inbred line) population, segregating for seed dormancy. This analysis revealed that the seed water uptake is associated with a single major QTL on Pv03. The QTL region was fine-mapped to a 118 Kb interval possessing 11 genes. Coding sequence analysis of candidate genes revealed a 5-bp insertion in an ortholog of pectin acetylesterase 8 that causes a frame shift, loss-of-function mutation in non-dormant genotype. Gene expression analysis of the candidate genes in the seed coat of contrasting genotypes indicated 21-fold lower expression of pectin acetylesterase 8 in non-dormant genotype. An analysis of mutational polymorphism was conducted among wild and domesticated beans. Although all the wild beans possessed the functional allele of pectin acetylesterase 8, the majority (77%) of domesticated beans had the non-functional allele suggesting that this variant was under strong selection pressure through domestication. CONCLUSIONS: In this study, we identified the physiological mechanism of physical seed dormancy and have identified a candidate allele causing variation in this trait. Our findings suggest that a 5-bp insertion in an ortholog of pectin acetylesterase 8 is likely a major causative mutation underlying the loss of seed dormancy during domestication. Although the results of current study provide strong evidences for the role of pectin acetylesterase 8 in seed dormancy, further confirmations seem necessary by employing transgenic approaches.


Assuntos
Cromossomos de Plantas/genética , Esterases/metabolismo , Phaseolus/genética , Dormência de Plantas/genética , Locos de Características Quantitativas/genética , Mapeamento Cromossômico , Produtos Agrícolas , Domesticação , Ecossistema , Esterases/genética , Genótipo , Microscopia Eletrônica de Varredura , Mutagênese Insercional , Phaseolus/enzimologia , Phaseolus/fisiologia , Phaseolus/ultraestrutura , Fenótipo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Sementes/enzimologia , Sementes/genética , Sementes/fisiologia , Sementes/ultraestrutura , Água/metabolismo
12.
BMC Plant Biol ; 21(1): 50, 2021 Jan 19.
Artigo em Inglês | MEDLINE | ID: mdl-33468064

RESUMO

BACKGROUND: Simmondsia chinensis (jojoba) is the only plant known to store wax esters instead of triacylglycerols in its seeds. Wax esters are composed of very-long-chain monounsaturated fatty acids and fatty alcohols and constitute up to 60% of the jojoba seed weight. During jojoba germination, the first step of wax ester mobilization is catalyzed by lipases. To date, none of the jojoba lipase-encoding genes have been cloned and characterized. In this study, we monitored mobilization of storage reserves during germination of jojoba seeds and performed detailed characterization of the jojoba lipases using microsomal fractions isolated from germinating seeds. RESULTS: During 26 days of germination, we observed a 60-70% decrease in wax ester content in the seeds, which was accompanied by the reduction of oleosin amounts and increase in glucose content. The activity of jojoba lipases in the seed microsomal fractions increased in the first 50 days of germination. The enzymes showed higher activity towards triacylglycerols than towards wax esters. The maximum lipase activity was observed at 60 °C and pH around 7 for triacylglycerols and 6.5-8 for wax esters. The enzyme efficiently hydrolyzed various wax esters containing saturated and unsaturated acyl and alcohol moieties. We also demonstrated that jojoba lipases possess wax ester-synthesizing activity when free fatty alcohols and different acyl donors, including triacylglycerols and free fatty acids, are used as substrates. For esterification reactions, the enzyme utilized both saturated and unsaturated fatty alcohols, with the preference towards long chain and very long chain compounds. CONCLUSIONS: In in vitro assays, jojoba lipases catalyzed hydrolysis of triacylglycerols and different wax esters in a broad range of temperatures. In addition, the enzymes had the ability to synthesize wax esters in the backward reaction. Our data suggest that jojoba lipases may be more similar to other plant lipases than previously assumed.


Assuntos
Caryophyllales/enzimologia , Lipase/metabolismo , Proteínas de Plantas/metabolismo , Sementes/metabolismo , Triglicerídeos/metabolismo , Caryophyllales/metabolismo , Ésteres/química , Ésteres/metabolismo , Germinação , Hidrólise , Lipase/química , Lipídeos/análise , Lipídeos/química , Microssomos/efeitos dos fármacos , Microssomos/enzimologia , Microssomos/metabolismo , Orlistate/farmacologia , Proteínas de Plantas/química , Sementes/enzimologia , Especificidade por Substrato , Temperatura , Triglicerídeos/química , Ceras/química , Ceras/metabolismo
13.
PLoS One ; 16(1): e0246095, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33508026

RESUMO

Chapatti (unleavened flatbread) is a staple food in northern India and neighboring countries but the genetics behind its processing quality are poorly understood. To understand the genes determining chapatti quality, differentially expressed genes were selected from microarray data of contrasting chapatti cultivars. From the gene and trait association studies, a null allele of granule bound starch synthase (GBSS; Wx-B1) was found to be associated with low amylose content and good chapatti quality. For validation, near-isogenic lines (NILs) of this allele were created by marker assisted backcross (MAB) breeding. Background screening indicated 88.2 to 96.7% background recovery in 16 selected BC3F5 NILs. Processing quality and sensory evaluation of selected NILs indicated improvement in chapatti making quality. Traits that showed improvement were mouthfeel, tearing strength and softness indicating that the Wx-B1 may be one of the major genes controlling chapatti softness.


Assuntos
Alelos , Amilose , Pão , Proteínas de Plantas , Sementes , Sintase do Amido , Triticum , Amilose/genética , Amilose/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Sementes/enzimologia , Sementes/genética , Sintase do Amido/genética , Sintase do Amido/metabolismo , Triticum/enzimologia , Triticum/genética
14.
Int J Mol Sci ; 22(1)2021 Jan 02.
Artigo em Inglês | MEDLINE | ID: mdl-33401671

RESUMO

Beech seeds are produced irregularly, and there is a need for long-term storage of these seeds for forest management practices. Accumulated reactive oxygen species broadly oxidize molecules, including amino acids, such as methionine, thereby contributing to decreased seed viability. Methionine oxidation can be reversed by the activity of methionine sulfoxide reductases (Msrs), which are enzymes involved in the regulation of many developmental processes and stress responses. Two types of Msrs, MsrB1 and MsrB2, were investigated in beech seeds to determine their abundance and localization. MsrB1 and MsrB2 were detected in the cortical cells and the outer area of the vascular cylinder of the embryonic axes as well as in the epidermis and parenchyma cells of cotyledons. The abundances of MsrB1 and MsrB2 decreased during long-term storage. Ultrastructural analyses have demonstrated the accumulation of these proteins in protein storage vacuoles and in the cytoplasm, especially in close proximity to the cell membrane. In silico predictions of possible Msr interactions supported our findings. In this study, we investigate the contribution of MsrB1 and MsrB2 locations in the regulation of seed viability and suggest that MsrB2 is linked with the longevity of beech seeds via association with proper utilization of storage material.


Assuntos
Cotilédone/metabolismo , Fagus/metabolismo , Metionina Sulfóxido Redutases/metabolismo , Sementes/crescimento & desenvolvimento , Sementes/metabolismo , Sobrevivência Celular/fisiologia , Biologia Computacional , Cotilédone/citologia , Fagus/citologia , Fagus/embriologia , Fagus/crescimento & desenvolvimento , Imunofluorescência , Imuno-Histoquímica , Ligação Proteica , Sementes/citologia , Sementes/enzimologia
15.
J Agric Food Chem ; 69(4): 1206-1213, 2021 Feb 03.
Artigo em Inglês | MEDLINE | ID: mdl-33481586

RESUMO

Starch biosynthesis in cereal crops is a complex pathway regulated by multiple starch synthetic enzymes. Starch synthase IIa (SSIIa) is well-known to be one of the major starch synthases and is very important in amylopectin biosynthesis. It has significant effects on grain composition and kernel traits. However, there are few reports on the association of natural variation of SSIIa in barley and grain composition and characteristics. In this work, two SSIIa isoforms were first identified as SSIIaH and SSIIaL by one-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis, mass spectrometry, and Western blotting. Sequence analysis of the SSIIa gene demonstrated that a 33 bp insertion coding a peptide of APPSSVVPAKK caused different SSIIa, e.g., SSIIaH and SSIIaL. Based on this molecular difference, a polymerase chain reaction marker was developed, which could be used to screen different SSIIa genotypes easily. Kernel hardness of SSIIaL genotypes was significantly higher than that of SSIIaH Chinese barley cultivars. The proportion of SSIIaL genotypes was extremely low in Australian barley cultivars (5/24) and much higher in Tibetan hull-less barley cultivars (46/74), consistent with the end-use requirements of barley grain. This study provided new information in barley endosperm starch synthesis and indicated that it is valuable for choosing the preferred SSIIa genotype according to the end-use requirements.


Assuntos
Hordeum/enzimologia , Proteínas de Plantas/metabolismo , Sementes/química , Sintase do Amido/metabolismo , Sequência de Aminoácidos , Amilopectina/química , Amilopectina/metabolismo , Austrália , Hordeum/química , Hordeum/genética , Proteínas de Plantas/genética , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Sementes/enzimologia , Sementes/genética , Amido/química , Amido/metabolismo , Sintase do Amido/genética
16.
Int J Biol Macromol ; 171: 539-549, 2021 Feb 28.
Artigo em Inglês | MEDLINE | ID: mdl-33434550

RESUMO

The present study aimed to investigate the biological functions of germinated M. oleifera seed proteins and to identify the identity of milk-clotting proteases. A total of 963 proteins were identified, and those with molecular weights between 10 and 30 kDa were most abundant. The identified proteins were mainly involved in energy-associated catalytic activity and metabolic processes, and carbohydrate and protein metabolisms. The numbers of proteins associated with the hydrolytic and catalytic activities were higher than the matured dry M. oleifera seeds reported previously. Of the identified proteins, proteases were mainly involved in the milk-clotting activity. Especially, a cysteine peptidase with a molecular mass of 17.727 kDa exhibiting hydrolase and peptidase activities was purified and identified. The identified cysteine peptidase was hydrophilic, and its secondary structure consisted of 27.60% alpha helix, 9.20% beta fold, and 63.20% irregular curl; its tertiary structure was also constructed using M. oleifera seed 2S protein as the protein template. The optimal pH and temperature of the purified protease were pH 4.0 and 60 °C, respectively. The protease had high acidic stability and good thermostability, thus could potentially be applied in the dairy industry.


Assuntos
Caseínas/efeitos dos fármacos , Cisteína Proteases/isolamento & purificação , Moringa oleifera/enzimologia , Peptídeo Hidrolases/isolamento & purificação , Proteínas de Plantas/isolamento & purificação , Sementes/enzimologia , Sequência de Aminoácidos , Técnicas de Química Analítica , Cisteína Proteases/metabolismo , Estabilidade Enzimática , Ontologia Genética , Germinação , Temperatura Alta , Concentração de Íons de Hidrogênio , Interações Hidrofóbicas e Hidrofílicas , Modelos Moleculares , Peso Molecular , Moringa oleifera/química , Moringa oleifera/genética , Peptídeo Hidrolases/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Conformação Proteica , Proteoma
17.
Int J Biol Macromol ; 172: 452-463, 2021 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-33454325

RESUMO

Lipases play a crucial role in the life cycle of seed plants and the oil content of the seed is highly regulated by the lipase activity. Hence, understanding the role of lipases during germination and post-germination will provide insights into lipid mobilization. However, to date, no lipase gene has been identified in seeds except, Sugar-dependent-1 in Arabidopsis. Hence, in the present study, we employed a functional proteomic approach for the identification of seed-specific lipase. Activity-Based Proteome Profiling (ABPP) of Arabidopsis mature and germinating seeds revealed the expression of a functional serine hydrolase exclusively during germination. The mass-spectrometry analysis reveals the identity and amino acid sequence of the protein correspond to AT4G28520 gene, a canonical 12S Seed Storage Protein (SSP). Interestingly, the identified SSP was a proteoform of AT4G28520 (SL-AT4G28520) and exhibited >90% identity with the canonical AT4G28520 (FL-AT4G28520). Heterologous expression and enzyme assays indicated that SL-AT4G28520 protein indeed possesses monoacylglycerol lipase activity, while the FL-AT4G28520 protein didn't exhibit any detectable activity. Functional proteomics and lipidomics analysis demonstrated a catalytic function of this SSP. Collectively, this is the first report, which suggests that SL-AT4G28520 encodes a lipase, and the activity is depending on the physiological condition.


Assuntos
Regulação da Expressão Gênica de Plantas , Metabolismo dos Lipídeos/genética , Monoacilglicerol Lipases/metabolismo , Monoglicerídeos/metabolismo , Proteínas de Armazenamento de Sementes/metabolismo , Sementes/enzimologia , Sequência de Aminoácidos , Arabidopsis , Sítios de Ligação , Clonagem Molecular , Escherichia coli/genética , Escherichia coli/metabolismo , Expressão Gênica , Perfilação da Expressão Gênica , Vetores Genéticos/química , Vetores Genéticos/metabolismo , Germinação/fisiologia , Isoenzimas/genética , Isoenzimas/metabolismo , Cinética , Monoacilglicerol Lipases/genética , Ligação Proteica , Proteômica/métodos , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Proteínas de Armazenamento de Sementes/genética , Sementes/genética , Sementes/crescimento & desenvolvimento , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Especificidade por Substrato , Termodinâmica
18.
Plant Cell Environ ; 44(1): 68-87, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-32974958

RESUMO

Superoxide dismutases (SODs) are enzymes detoxifying superoxide to hydrogen peroxide while temporal developmental expression and subcellular localisation are linked to their functions. Therefore, we aimed here to reveal in vivo developmental expression, subcellular, tissue- and organ-specific localisation of iron superoxide dismutase 1 (FSD1) in Arabidopsis using light-sheet and Airyscan confocal microscopy. FSD1-GFP temporarily accumulated at the site of endosperm rupture during seed germination. In emerged roots, it showed the highest abundance in cells of the lateral root cap, columella, and endodermis/cortex initials. The largest subcellular pool of FSD1-GFP was localised in the plastid stroma, while it was also located in the nuclei and cytosol. The majority of the nuclear FSD1-GFP is immobile as revealed by fluorescence recovery after photobleaching. We found that fsd1 knockout mutants exhibit reduced lateral root number and this phenotype was reverted by genetic complementation. Mutant analysis also revealed a requirement for FSD1 in seed germination during salt stress. Salt stress tolerance was coupled with the accumulation of FSD1-GFP in Hechtian strands and superoxide removal. It is likely that the plastidic pool is required for acquiring oxidative stress tolerance in Arabidopsis. This study suggests new developmental and osmoprotective functions of SODs in plants.


Assuntos
Proteínas de Arabidopsis/metabolismo , Superóxido Dismutase/metabolismo , Proteínas de Arabidopsis/fisiologia , Imunofluorescência , Germinação , Microscopia , Microscopia Confocal , Raízes de Plantas/enzimologia , Raízes de Plantas/metabolismo , Plantas Geneticamente Modificadas , Reação em Cadeia da Polimerase em Tempo Real , Sementes/enzimologia , Sementes/metabolismo , Estresse Fisiológico , Superóxido Dismutase/fisiologia
19.
Appl Biochem Biotechnol ; 193(2): 389-404, 2021 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-33009584

RESUMO

Proteolytic enzymes are widely distributed in nature, playing essential roles in important biological functions. Recently, the use of plant proteases at the industrial level has mainly increased in the food industry (e.g., cheesemaking, meat tenderizing, and protein hydrolysate production). Current technological and scientific advances in the detection and characterization of proteolytic enzymes have encouraged the search for new natural sources. Thus, this work aimed to explore the milk-clotting and proteolytic properties of different tissues of Vallesia glabra. Aqueous extracts from the leaves, fruits, and seeds of V. glabra presented different protein profiles, proteolytic activity, and milk-clotting activity. The milk-clotting activity increased with temperature (30-65 °C), but this activity was higher in leaf (0.20 MCU/mL) compared with that in fruit and seed extracts (0.12 and 0.11 MCU/mL, respectively) at 50 °C. Proteolytic activity in the extracts assayed at different pH (2.5-12.0) suggested the presence of different types of active proteases, with maximum activity at acidic conditions (4.0-4.5). Inhibitory studies indicated that major activity in V. glabra extracts is related to cysteine proteases; however, the presence of serine, aspartic, and metalloproteases was also evident. The hydrolytic profile of caseins indicated that V. glabra leaves could be used as a rennet substitute in cheesemaking, representing a new and promising source of proteolytic enzymes.


Assuntos
Apocynaceae/enzimologia , Leite/química , Peptídeo Hidrolases/química , Folhas de Planta/enzimologia , Proteínas de Plantas/química , Proteólise , Sementes/enzimologia , Animais , Concentração de Íons de Hidrogênio
20.
Plant Mol Biol ; 105(3): 247-262, 2021 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-33089420

RESUMO

KEY MESSAGE: Two OsDGAT1 genes showed the ability to restore TAG and LB synthesis in yeast H1246. Alterations in the N-terminal region of OsDGAT1-1 gene revealed its regulatory role in gene function. Accumulation of triacylglycerol (TAG) or oil in vegetative tissues has emerged as a promising approach to meet the global needs of food, feed, and fuel. Rice (Oryza sativa) has been recognized as an important cereal crop containing nutritional rice bran oil with high economic value for renewable energy production. To identify the key component involved in storage lipid biosynthesis, two type-1 diacylglycerol acyltransferases (DGAT1) from rice were characterized for its in vivo function in the H1246 (dga1, lro1, are1 and are2) yeast quadruple mutant. The ectopic expression of rice DGAT1 (designated as OsDGAT1-1 and OsDGAT1-2) genes restored the capability of TAG synthesis and lipid body (LB) formation in H1246. OsDGAT1-1 showed nearly equal substrate preferences to C16:0-CoA and 18:1-CoA whereas OsDGAT1-2 displayed substrate selectivity for C16:0-CoA over 18:1-CoA, indicating that these enzymes have contrasting substrate specificities. In parallel, we have identified the intrinsically disordered region (IDR) at the N-terminal domains of OsDGAT1 proteins. The regulatory role of the N-terminal domain was dissected. Single point mutations at the phosphorylation sites and truncations of the N-terminal region highlighted reduced lipid accumulation capabilities among different OsDGAT1-1 variants.


Assuntos
Diacilglicerol O-Aciltransferase/genética , Oryza/enzimologia , Oryza/genética , Saccharomyces cerevisiae/metabolismo , Sementes/enzimologia , Sementes/genética , Triglicerídeos/metabolismo , Sequência de Aminoácidos , Diacilglicerol O-Aciltransferase/química , Diacilglicerol O-Aciltransferase/metabolismo , Diglicerídeos/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Regulação da Expressão Gênica de Plantas , Proteínas Intrinsicamente Desordenadas/química , Proteínas Intrinsicamente Desordenadas/metabolismo , Isoenzimas/genética , Isoenzimas/metabolismo , Gotículas Lipídicas/metabolismo , Mutação/genética , Fosforilação , Filogenia , Proteínas de Plantas/química , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Domínios Proteicos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Especificidade por Substrato
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA
...