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1.
Food Chem ; 308: 125663, 2020 Mar 05.
Artigo em Inglês | MEDLINE | ID: mdl-31655474

RESUMO

Apple exocarp was used to investigate the effect of acibenzolar-S-methyl (ASM) and dehydroepiandrosterone (DHEA) treatments on reaction oxygen species (ROS) metabolism. The results indicated that ASM enhanced the hydrogen peroxide (H2O2) content, the activities of superoxide dismutase (SOD), catalase (CAT), peroxidase (POD), ascorbate peroxidase (APX), monodehydroascorbate reductase (MDHAR), glutathione reductase (GR) and glucose-6-phosphate dehydrogenase (G6PDH). ASM also increased the contents of ascorbic acid (AsA), reduced glutathione (GSH) and nicotinamide ademine dinucleotidephosphate (NADPH), MdSOD and MdAPX expression, but decreased MdMDHAR and dehydroascorbate reductase (MdDHAR) expression. DHEA suppressed H2O2 accumulation and POD, APX, MDHAR, G6PDH activities, but increased SOD, CAT and GR activities compared to the control. ASM and DHEA treatments suppressed the contents of AsA, GSH and NADPH, and expression of MdSOD, MdAPX and MdMDHAR. These results suggest that DHEA treatment prevented ROS metabolism induced by ASM which showed the important role of G6PDH in maintaining redox homeostasis in apple exocarp.


Assuntos
Glucosefosfato Desidrogenase/metabolismo , Malus/enzimologia , Espécies Reativas de Oxigênio/metabolismo , Ascorbato Peroxidases/metabolismo , Ácido Ascórbico/metabolismo , Catalase/metabolismo , Glutationa/metabolismo , Glutationa Redutase/metabolismo , Peróxido de Hidrogênio/metabolismo , NADH NADPH Oxirredutases/metabolismo , Oxirredução , Oxirredutases/metabolismo , Superóxido Dismutase/metabolismo , Tiadiazóis/metabolismo
2.
Plant Cell Physiol ; 60(10): 2129-2140, 2019 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-31165159

RESUMO

Apple ring rot is a severe disease that affects the yield and quality of apple fruits worldwide. However, the underlying molecular mechanism that involved in this process still remains largely unexplored. Here, we report that apple POZ/BTB CONTAINING-PROTEIN 1 (MdPOB1), a BTB-BACK domain E3 ligase protein, functions to suppress apple pathogen defense against Botryosphaeria dothidea (B. dothidea). Both in vitro and in vivo assays indicated that MdPOB1 interacted directly with and degraded apple U-box E3 ligase MdPUB29, a well-established positive regulator of plant innate immunity, through the ubiquitin/26S proteasome pathway. A series of transgenic analyses in apple fruits demonstrated that MdPOB1 affected apple pathogen defense against B. dothidea at least partially, if not completely, via regulating MdPUB29. Additionally, it was found that the apple pathogen defense against B. dothidea was correlated with the H2O2 contents and the relative expression of salicylic acid (SA) synthesis- and SA signaling-related genes, which might be regulated via degradation of MdPUB29 by MdPOB1. Overall, our findings provide new insights into the mechanism of the MdPOB1 modulation of apple ring rot resistance, which occur by directly regulating potential downstream target protein MdPUB29 for proteasomal degradation in apple.


Assuntos
Ascomicetos/fisiologia , Resistência à Doença/genética , Malus/genética , Doenças das Plantas/imunologia , Proteínas de Plantas/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Frutas/enzimologia , Frutas/genética , Frutas/imunologia , Frutas/microbiologia , Peróxido de Hidrogênio/metabolismo , Malus/enzimologia , Malus/imunologia , Malus/microbiologia , Doenças das Plantas/microbiologia , Proteínas de Plantas/genética , Domínios Proteicos , Proteólise , Ácido Salicílico/metabolismo , Ubiquitina-Proteína Ligases/genética , Ubiquitinação
3.
Food Chem ; 289: 657-663, 2019 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-30955661

RESUMO

Polyphenol oxidase from Granny Smith apples was purified and characterized in both its soluble form (sPPO) and its membrane-bound form (mPPO). Both forms were purified by temperature-induced phase partitioning, precipitation with ammonium sulfate, and ion exchange chromatography. The specific activity of mPPO was 19.17 times that of sPPO. The optimum pH and temperature for both forms were 7.0 and 35 °C when catechol was the substrate. The Michaelis constant and maximum reaction rate for sPPO were 34.1 mM and 500 U/mL/min, whereas those for mPPO were 53 mM and 10,000 U/mL/min, respectively. The enzymes exhibited diphenolase activity, and their affinity was highest for catechol (sPPO) and 4-methylcatechol (mPPO). Inhibitors of sPPO and mPPO included ascorbic acid, glutathione, and l-cysteine. However, ethylenediaminetetraacetic acid increased the activity of mPPO. Purified sPPO was dimeric with a molecular weight of 31 kDa, whereas mPPO was monomeric with an estimated molecular weight of 65 kDa.


Assuntos
Catecol Oxidase/metabolismo , Frutas/enzimologia , Malus/enzimologia , Ácido Ascórbico/metabolismo , Catecóis/química , Cisteína/metabolismo , Ácido Edético/metabolismo , Glutationa/metabolismo , Concentração de Íons de Hidrogênio , Peso Molecular , Proteínas de Plantas/metabolismo , Especificidade por Substrato , Temperatura
4.
Plant Physiol Biochem ; 139: 141-151, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30889479

RESUMO

Proanthocyanidins (PAs) from plants are a nutritionally valuable component of the human diet and play important roles in defense against pests and diseases. PAs are products of the flavonoid pathway, which also leads to the production of anthocyanins and flavonols. The enzymes leucoanthocyanidin reductase (LAR) and anthocyanidin reductase (ANR) are involved in PA biosynthesis. The PA biosynthetic pathway has been characterized in several plant species, but the relationship between its expression and PA accumulation in Malus crabapple remains unclear. Here, we cloned the LAR genes MrLAR1, 2, and the ANR genes MrANR1, 2, from the red leaved Malus crabapple cultivar 'Royalty'. The contents of PAs and the expression levels of the LAR and ANR genes were investigated in different organs of the two crabapple cultivars. The transcript levels of two LAR genes and two ANR genes correlated with the contents of the catechin and epicatechin, which are proanthocyanidin precursors. Over-expression of the MrLAR1, 2 and MrANR1, 2 in tobacco (Nicotiana tabacum) promoted the accumulation of PAs, while transient silencing of their expression in crabapple resulted in reduced PA levels. In addition, a negative correlation between quercetin, anthocyanin, and PA biosynthesis was also found during crabapple leaf and fruit peel development. We also found that MrLAR1 and 2 may contribute to epicatechin biosynthesis. In summary, the LAR and ANR genes are critical factors in PA biosynthesis, and there is competition between the quercetin, anthocyanin, and PA biosynthetic pathways during leaf and fruit peel development in Malus crabapple.


Assuntos
Antocianinas/metabolismo , Malus/metabolismo , NADH NADPH Oxirredutases/genética , Proteínas de Plantas/genética , Proantocianidinas/biossíntese , Catequina/metabolismo , Cromatografia Líquida de Alta Pressão , Clonagem Molecular , Perfilação da Expressão Gênica , Genes de Plantas/genética , Malus/enzimologia , Malus/genética , Redes e Vias Metabólicas/genética , NADH NADPH Oxirredutases/metabolismo , Proteínas de Plantas/metabolismo , Plantas Geneticamente Modificadas , Reação em Cadeia da Polimerase em Tempo Real , Análise de Sequência de DNA , Tabaco
5.
Carbohydr Polym ; 209: 338-349, 2019 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-30732816

RESUMO

The effect of wheat straw arabinoxylan (AX) and ß-glucan stearic acid ester (SABG) composite coating on the quality and storage life of apple (Royal Delicious) was studied at 22 °C (±2) with relative humidity of 65% and 85% for 60 days. Fresh fruits were coated with surface coatings of AX-SABG, shellac in the concentration range of 1-4%. Application of both AX-SABG (1-4%) and shellac (1-4%) coatings was found to significantly reduce weight loss, respiration rate, fruit softening process, ripening index, color degradation and polyphenol oxidase activity compared to control during the storage period of more than 30 days. However, an AX-SABG coating was more effective in reducing fruit decay and loss of aroma volatiles followed by shellac coated apples; the un-coated apples being showing maximum quality deterioration. These findings confirmed the potential benefits of applying AX-SABG coating to extend the shelf life and quality of apples especially during transportation and storage.


Assuntos
Ésteres/química , Qualidade dos Alimentos , Malus/efeitos dos fármacos , Ácidos Esteáricos/química , Xilanos/farmacologia , beta-Glucanas/química , beta-Glucanas/farmacologia , Fenômenos Biomecânicos/efeitos dos fármacos , Catecol Oxidase/metabolismo , Respiração Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Cor , Conservação de Alimentos , Frutas/efeitos dos fármacos , Frutas/metabolismo , Malus/citologia , Malus/enzimologia , Malus/metabolismo , Odorantes/análise , Peroxidase/metabolismo
6.
Planta ; 249(4): 1177-1188, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30603792

RESUMO

MAIN CONCLUSION: MdPUB29 is a positive regulator of the defense response to the fungal pathogen Botryosphaeria dothidea possibly by directly regulating the salicylic acid (SA) content as well as SA synthesis-related and signaling-related gene transcription. In plants, ubiquitin E3 ligases containing a U-box domain (PUBs, Plant U-box E3 ubiquitin ligase) have been identified as key regulators of fundamental cellular processes, such as cellular growth, development, and apoptosis, as well as biotic and abiotic stress responses. However, the function of PUBs in apple ring rot remains elusive. Here, we isolated the U-box E3 ligase MdPUB29 from the apple cultivar 'Royal Gala' and characterized its function in plant pathogen defense against Botryosphaeria dothidea. qRT-PCR showed that the expression of MdPUB29 was significantly induced in apple fruits after B. dothidea infection. Overexpression of the MdPUB29 gene in apple calli increased the resistance to B. dothidea infection. In contrast, silencing MdPUB29 in apple calli resulted in reduced resistance. Ectopic expression of MdPUB29 in Arabidopsis also exhibited enhanced resistance to B. dothidea infection compared to that of the wild-type (Col) control. In addition, it was found that the increase of plant pathogen defense was correlated with the increased salicylic acid (SA) content, as well as SA synthesis-related and signaling-related gene transcription in comparison to the wild type. We elucidated the mechanism by which MdPUB29 elevates plant pathogen defense against B. dothidea possibly by regulating the SA pathway.


Assuntos
Ascomicetos , Malus/genética , Doenças das Plantas/microbiologia , Imunidade Vegetal/genética , Proteínas de Plantas/genética , Ubiquitina-Proteína Ligases/genética , Clorofila/metabolismo , Clonagem Molecular , Regulação da Expressão Gênica de Plantas , Glucanos/metabolismo , Malus/enzimologia , Malus/imunologia , Malus/microbiologia , Doenças das Plantas/imunologia , Reguladores de Crescimento de Planta/metabolismo , Proteínas de Plantas/fisiologia , Espécies Reativas de Oxigênio/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Ácido Salicílico/metabolismo , Ubiquitina-Proteína Ligases/fisiologia
7.
J Sci Food Agric ; 99(4): 1828-1833, 2019 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-30255560

RESUMO

BACKGROUND: During the storage of apples, apple softening is one of the main problems. Sodium silicate has been used to enhance disease resistance and maintain quality of fruits. In the present study, apple fruit (cv. Golden delicious) were treated with 100 mmol L-1 sodium silicate for 10 min and stored at 20 °C to investigate its effects on weight loss, flesh firmness, and the activity of cell wall-degrading enzymes. RESULTS: The results indicated that 100 mmol L-1 of sodium silicate treatment delayed the increase of weight loss and decrease of the flesh firmness in apples. Sodium silicate treatment also suppressed the activity of polygalacturonic acid transeliminase and pectin methyltranseliminase, pectin methylgalacturonase, polygalacturonase, cellulase and ß-galactosidase in the fruit. CONCLUSIONS: Delaying apple softening by sodium silicate treatment is closely related to the inhibition of the activity of cell wall-degrading enzymes and weight loss. © 2018 Society of Chemical Industry.


Assuntos
Parede Celular/enzimologia , Conservantes de Alimentos/farmacologia , Frutas/efeitos dos fármacos , Malus/química , Proteínas de Plantas/metabolismo , Silicatos/farmacologia , Parede Celular/metabolismo , Celulase/metabolismo , Conservação de Alimentos , Frutas/química , Frutas/enzimologia , Frutas/metabolismo , Malus/efeitos dos fármacos , Malus/enzimologia , Malus/metabolismo , Metiltransferases/metabolismo , Pectinas/metabolismo , Controle de Qualidade
8.
Planta ; 249(3): 677-691, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30357505

RESUMO

MAIN CONCLUSION: This manuscript describes the cloning and functional characterization of a biphenyl phytoalexin biosynthetic gene, 3,5-dihydroxybiphenyl O-methyltransferase from elicitor-treated cell cultures of scab resistant apple cultivar 'Florina'. Apples belong to the subtribe Malinae of the Rosaceae family. Biphenyls and dibenzofurans are the specialized phytoalexins of Malinae, of which aucuparin is the most widely distributed biphenyl. The precursor of aucuparin, 3,5-dihydroxybiphenyl, is a benzoate-derived polyketide, which is formed by the sequential condensation of three molecules of malonyl-CoA and one molecule of benzoyl-CoA in a reaction catalyzed by biphenyl synthase (BIS). This 3,5-dihydroxybiphenyl then undergoes sequential 5-O-methylation, 4-hydroxylation, and finally 3-O-methylation to form aucuparin. A cDNA encoding O-methyltransferase (OMT) was isolated and functionally characterized from the cell cultures of scab-resistant apple cultivar 'Florina' (Malus domestica cultivar 'Florina'; MdOMT) after treatment with elicitor prepared from the apple scab causing fungus Venturia inaequalis. MdOMT catalyzed the regiospecific O-methylation of 3,5-dihydroxybiphenyl at the 5-position to form 3-hydroxy-5-methoxybiphenyl. The enzyme showed absolute substrate preference for 3,5-dihydroxybiphenyl. The elicitor-treated apple cell cultures showed transient increases in the MdOMT (GenBank ID MF740747) and MdBIS3 (GenBank ID JQ390523) transcript levels followed by the accumulation of biphenyls (aucuparin and noraucuparin) and dibenzofuran (eriobofuran) phytoalexins. MdOMT fused with N- and C-terminal yellow fluorescent protein showed cytoplasmic localization in the epidermis of Nicotiana benthamiana leaves. In scab inoculated greenhouse-grown 'Florina' plants, the expression of MdOMT was transiently induced in the stem followed by the accumulation of biphenyl phytoalexins.


Assuntos
Malus/enzimologia , Metiltransferases/metabolismo , Sesquiterpenos/metabolismo , Células Cultivadas , Clonagem Molecular , Malus/citologia , Malus/genética , Malus/metabolismo , Redes e Vias Metabólicas , Metiltransferases/genética , Metiltransferases/fisiologia , Filogenia , Alinhamento de Sequência , Especificidade por Substrato
9.
Phytochemistry ; 157: 135-144, 2019 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-30399496

RESUMO

The UDP-glycosyltransferase UGT88F subfamily has been described first in Malus x domestica with the characterization of UGT88F1. Up to now UGT88F1 was one of the most active UGT glycosylating dihydrochalcones in vitro. The involvement of UGT88F1 in phloridzin (phloretin 2'-O-glucoside) synthesis, the main apple tree dihydrochalcone, was further confirmed in planta. Since the characterization of UGT88F1, this new UGT subfamily has been poorly studied probably because it seemed restricted to Maloideae. In the present study, we investigate the apple tree genome to identify and biochemically characterize the whole UGT88F subfamily. The apple tree genome contains five full-length UGT88F genes out of which three newly identified members (UGT88F6, UGT88F7 and UGT88F8) and a pseudogene. These genes are organized into two genomic clusters resulting from the recent global genomic duplication event in the apple tree. We show that recombinant UGT88F8 protein specifically glycosylates phloretin in the 2'OH position to synthetize phloridzin in vitro and was therefore named UDP-glucose: phloretin 2'-O-glycosyltransferase. The Km values of UGT88F8 are 7.72 µM and 10.84 µM for phloretin and UDP-glucose respectively and are in the same range as UGT88F1 catalytic parameters thus constituting two isoforms. Co-expression patterns of both UGT88F1 and UGT88F8 argue for a redundant function in phloridzin biosynthesis in planta. Contrastingly, recombinant UGT88F6 protein is able to glycosylate in vitro a wide range of flavonoids including flavonols, flavones, flavanones, chalcones and dihydrochalcones, although flavonols are the preferred substrates, e.g. Km value for kaempferol is 2.1 µM. Depending on the flavonoid, glycosylation occurs at least on the 3-OH and 7-OH positions. Therefore UGT88F6 corresponds to an UDP-glucose: flavonoid 3/7-O-glycosyltransferase. Finally, a molecular modeling study highlights a very high substitution rate of residues in the acceptor binding pocket between UGT88F8 and UGT88F6 which is responsible for the enzymes divergence in substrate and regiospecificity, despite an overall high protein homology.


Assuntos
Genômica , Glicosiltransferases/genética , Glicosiltransferases/metabolismo , Malus/enzimologia , Malus/genética , Genoma de Planta/genética , Glicosiltransferases/química , Concentração de Íons de Hidrogênio , Cinética , Modelos Moleculares , Conformação Proteica , Temperatura
10.
Ecotoxicol Environ Saf ; 168: 230-240, 2019 Jan 30.
Artigo em Inglês | MEDLINE | ID: mdl-30388541

RESUMO

Cadmium (Cd) induces cell death in plant roots. Mitogen-activated protein kinase (MAPK) plays a role in the regulation of cell death induced by Cd in plant roots. In this study, MhMAPK4 was isolated from the roots of Malus hupehensis. Subcellular localization showed that the MhMAPK4 protein was located in the cell membrane and cytoplasm and is a transmembrane protein that is characterized by hydrophily. The expression of MhMAPK4 in the roots of M. hupehensis was up-regulated by Cd sulfate and Cd chloride. Phenotypic comparison under Cd stress showed that the growth of wild-type (WT) tobacco was lower than the transgenic lines overexpressing MhMAPK4. The fresh weight and the root length of WT also was lower than that of the transgenic tobacco. The net Cd2+ influx in the tobacco roots was decreased by the overexpression of MhMAPK4, as was root Cd accumulation. The recovery time of the Cd2+ influx to stable state in the transgenic tobacco was also shorter than the WT. The expression of iron-regulated transporter 1 (NtIRT1) and natural resistance associated macrophage protein 5 (NtNRAMP5) was relatively low in the transgenic lines under Cd stress. Cell death and apoptosis in the tobacco roots was reduced following the overexpression of MhMAPK4. The activity of vacuolar processing enzyme (VPE) and the transcript level of VPE in the transgenic tobacco was lower than that of WT under Cd stress. In addition, the electrolyte leakage and malondialdehyde and hydrogen peroxide contents in the transgenic tobacco were lower than those of WT, whereas the antioxidant enzyme activity and expression were higher. These results suggest that MhMAPK4 regulates Cd accumulation by mediating Cd2+ uptake by the roots, and controls Cd-caused cell death by adjusting VPE activity.


Assuntos
Cádmio/toxicidade , Morte Celular/efeitos dos fármacos , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Malus/enzimologia , Proteínas de Plantas/metabolismo , Raízes de Plantas/efeitos dos fármacos , Tabaco/efeitos dos fármacos , Sequência de Aminoácidos , Proteínas de Transporte de Cátions/genética , Proteínas de Transporte de Cátions/metabolismo , Clonagem Molecular , MAP Quinases Reguladas por Sinal Extracelular/genética , Regulação da Expressão Gênica de Plantas , Peróxido de Hidrogênio/metabolismo , Malondialdeído/metabolismo , Malus/genética , Proteínas de Plantas/genética , Raízes de Plantas/metabolismo , Plantas Geneticamente Modificadas/efeitos dos fármacos , Plantas Geneticamente Modificadas/metabolismo , Estresse Fisiológico/efeitos dos fármacos , Estresse Fisiológico/genética , Tabaco/metabolismo
11.
Plant Physiol ; 179(1): 88-106, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30333149

RESUMO

SIZ1 (a SIZ/PIAS-type SUMO E3 ligase)-mediated small ubiquitin-like modifier (SUMO) modification of target proteins is important for various biological processes related to abiotic stress resistance in plants; however, little is known about its role in resistance toward iron (Fe) deficiency. Here, the SUMO E3 ligase MdSIZ1 was shown to be involved in the plasma membrane (PM) H+-ATPase-mediated response to Fe deficiency. Subsequently, a basic helix-loop-helix transcription factor, MdbHLH104 (a homolog of Arabidopsis bHLH104 in apple), which acts as a key component in regulating PM H+-ATPase-mediated rhizosphere acidification and Fe uptake in apples (Malus domestica), was identified as a direct target of MdSIZ1. MdSIZ1 directly sumoylated MdbHLH104 both in vitro and in vivo, especially under conditions of Fe deficiency, and this sumoylation was required for MdbHLH104 protein stability. Double substitution of K139R and K153R in MdbHLH104 blocked MdSIZ1-mediated sumoylation in vitro and in vivo, indicating that the K139 and K153 residues were the principal sites of SUMO conjugation. Moreover, the transcript level of the MdSIZ1 gene was substantially induced following Fe deficiency. MdSIZ1 overexpression exerted a positive influence on PM H+-ATPase-mediated rhizosphere acidification and Fe uptake. Our findings reveal an important role for sumoylation in the regulation of PM H+-ATPase-mediated rhizosphere acidification and Fe uptake during Fe deficiency in plants.


Assuntos
Ferro/metabolismo , Malus/enzimologia , ATPases Translocadoras de Prótons/metabolismo , Ubiquitinas/fisiologia , Membrana Celular/metabolismo , Malus/metabolismo , RNA Mensageiro/metabolismo , Rizosfera , Sumoilação , Ubiquitinas/genética , Ubiquitinas/metabolismo
12.
J Plant Physiol ; 232: 216-225, 2019 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-30537609

RESUMO

In plants, SIZ1 regulates abiotic and biotic stress responses by promoting the SUMOylation of proteins. The apple MdSIZ1 protein has conserved domains similar to those of Arabidopsis AtSIZ1. Real-time fluorescent quantitative analysis showed that MdSIZ1 gene expression was induced by phosphate-deficient conditions. In addition, the level of SUMOylation was also significantly increased under these conditions. The MYB transcription factor MdPHR1 might be a target for the SUMO protein, which is a phosphorus starvation-dependent protein. Subsequently, an MdSIZ1 expression vector was constructed and transformed in Arabidopsis mutant siz1-2 and apple callus. The MdSIZ1 transgenic Arabidopsis partially complemented the defect phenotype of siz1-2 under phosphate-deficient conditions. The survival rate, length of primary root, and number or density of lateral roots were similar between the transgenic lines and wild type (WT). Under phosphate-deficient conditions, the SUMO conjugate and fresh weight of the MdSIZ1 transgenic apple callus were improved compared with WT. The MdSIZ1 transgenic apple callus grew under phosphate-deficient conditions, whereas the MdSIZ1 sense apple callus did not. Therefore, MdSIZ1 is involved in the regulation of the phosphate-deficiency response in apple.


Assuntos
Ligases/fisiologia , Malus/fisiologia , Fosfatos/deficiência , Proteínas de Plantas/fisiologia , Arabidopsis , Regulação da Expressão Gênica de Plantas , Ligases/metabolismo , Malus/enzimologia , Malus/metabolismo , Proteínas de Plantas/metabolismo , Plantas Geneticamente Modificadas , Sumoilação
13.
J Sci Food Agric ; 99(4): 1519-1524, 2019 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-30142245

RESUMO

BACKGROUND: Apple (cv. Ralls) fruit were treated with 0.1 g L-1 acibenzolar-S-methyl (ASM) for 10 min to evaluate the changes in enzyme activity and gene expression in the sucrose metabolism during storage at 20 °C with 30%-40% relative humidity. RESULTS: The results showed that sucrose phosphate synthase (SPS) and sucrose synthase synthesis (SS-s) activity was enhanced by ASM in apple fruit during the entire storage period. Sucrose synthase-cleavage (SS-c) and neutral invertase (NI) activity was suppressed by ASM treatment but acid invertase (AI) activity was increased in the middle period after ASM treatment. Acibenzolar-S-methyl treatment also significantly inhibited SPS and NI gene expression in apple fruit during storage. However, SS gene expression increased in the ASM-treated apple fruit. High levels of expression of the fructokinase (FK) and hexokinase (HK) genes were observed during the middle storage period in the ASM-treated fruit. CONCLUSION: Taken together, these results suggest that ASM delays the senescence of apple fruit by regulating the sugar metabolism. © 2018 Society of Chemical Industry.


Assuntos
Conservantes de Alimentos/farmacologia , Frutas/efeitos dos fármacos , Malus/metabolismo , Sacarose/metabolismo , Tiadiazóis/farmacologia , Frutas/enzimologia , Frutas/crescimento & desenvolvimento , Frutas/metabolismo , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Glucosiltransferases/genética , Glucosiltransferases/metabolismo , Malus/efeitos dos fármacos , Malus/enzimologia , Malus/crescimento & desenvolvimento , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo
14.
Food Chem ; 274: 415-421, 2019 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-30372959

RESUMO

Photodynamic treatment (PDT) is an innovative technology with non-thermal and environmentally sound merits, but the evaluation on the storage qualities of fresh produce was scarce. In this study, the effects of curcumin-based PDT on the quality of fresh-cut 'Fuji' apple slices during storage at 4 °C were investigated. The impacts on the survival of Escherichia coli, color and weight loss were examined under different curcumin concentrations, illumination time or incubation time. Curcumin-based photodynamic inactivation of E. coli on the surface of apple slices reached 0.95 log. Curcumin-based PDT was proven to prevent browning and weight loss. Additionally, PDT significantly reduced the activity of polyphenol oxidase and peroxidases to 48% and 51%, respectively. Moreover, there were few negative changes in total phenolic, ascorbic acid content and anti-oxidant activity of the treated apples. These results indicated that curcumin-based PDT was a viable and promising non-thermal technology to preserve the quality of fresh produce.


Assuntos
Curcumina/farmacologia , Qualidade dos Alimentos , Armazenamento de Alimentos , Malus/efeitos dos fármacos , Malus/efeitos da radiação , Cor , Escherichia coli/efeitos dos fármacos , Escherichia coli/fisiologia , Escherichia coli/efeitos da radiação , Malus/enzimologia , Malus/microbiologia , Fotoquimioterapia
15.
J Agric Food Chem ; 66(51): 13473-13482, 2018 Dec 26.
Artigo em Inglês | MEDLINE | ID: mdl-30512945

RESUMO

Organic acid is an important indicator of fruit quality, and malate is the predominant organic acid in apple fruit. However, the regulation of malate metabolism in postharvest fruit is rarely reported. Here, we found that, compared with a control treatment, a 10 mM γ-aminobutyric acid (GABA) treatment remarkably delayed the loss of tiftratable acidity and malate and increased the succinate and oxalate contents in "Cripps Pink" fruit stored in polyethylene bags at room temperature. The higher malate levels in GABA-treated fruit were accompanied by higher activities of cytosolic nicotinamide adenine dinucleotide-dependent malate dehydrogenase (cyNAD-MDH) and phosphoenolpyruvate carboxylase (PEPC) but lower cytosolic NAD phosphate-dependent malic enzyme (cyNADP-ME) and phosphoenolpyruvate carboxykinase (PEPCK) activities than those seen in control fruit. Notably, ethylene production was significantly reduced by GABA treatment, paralleling the downregulation of MdACS, MdACO, and MdERF expression. Meanwhile, GABA treatment also enhanced the activity of the GABA shunt and promoted the accumulation of GABA. This study provides new insights into the regulation of malate metabolism and reports for the first time the possible interplay between GABA and ethylene signaling pathways in apple fruit during postharvest storage.


Assuntos
Etilenos/biossíntese , Conservação de Alimentos/métodos , Conservantes de Alimentos/farmacologia , Frutas/efeitos dos fármacos , Malatos/metabolismo , Ácido gama-Aminobutírico/farmacologia , Frutas/enzimologia , Frutas/genética , Frutas/metabolismo , Malato Desidrogenase/genética , Malato Desidrogenase/metabolismo , Malus/efeitos dos fármacos , Malus/enzimologia , Malus/genética , Malus/metabolismo , Fosfoenolpiruvato Carboxiquinase (ATP)/genética , Fosfoenolpiruvato Carboxiquinase (ATP)/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo
16.
Plant Physiol Biochem ; 133: 81-91, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30391815

RESUMO

Tyrosine aminotransferase (TAT, EC 2.6.1.5) is the first key enzyme that catalyzes the reversible interconversion of tyrosine and 4-hydroxyphenylpyruvate in the tyrosine-derived pathway for syntheses of important secondary metabolites and compounds. Although plant TAT genes have been proposed to be important in response to abiotic stress, there is little information about TAT genes in woody perennial tree species, especially in economic fruit trees. Based on TAT domain searching, sequence homology screening and phylogenetic analysis, we identified four TATs in apple genome. Then, we carried out a detailed phylogenetic analysis of TAT genes from multi-species, focusing on apple (Malus domestica). The result showed that the TAT family comprises three major classes corresponding to genes from angiosperms, mammals, and bacteria. Angiosperm TAT genes could be further divided into six subclasses. Analysis of intron-exon structure revealed that the typical TAT gene contains six introns and seven exons, with exons of similar size at each exon location. Promoter analysis showed that the 5'-flanking region of apple MdTATs contain multiple cis-acting elements including those implicated in light, biotic stress, abiotic stress, and hormone response. MdTATs were expressed to various levels in all apple structures and organs evaluated, and showed distinct expression patterns under water deficit stress. Ectopic expression of MdTAT2 in Arabidopsis or over-expression of MdTAT2 in apple callus tissue conferred enhanced tolerance to drought and osmotic stress. Collectively, these results suggest a role for TAT genes in drought and osmotic stresses and provide valuable information for further research of TAT genes and their function in plants.


Assuntos
Genoma de Planta , Genômica , Malus , Pressão Osmótica , Proteínas de Plantas , Tirosina Transaminase , Desidratação/genética , Desidratação/metabolismo , Malus/enzimologia , Malus/genética , Proteínas de Plantas/biossíntese , Proteínas de Plantas/genética , Tirosina Transaminase/biossíntese , Tirosina Transaminase/genética
17.
Int J Mol Sci ; 19(11)2018 Oct 24.
Artigo em Inglês | MEDLINE | ID: mdl-30356028

RESUMO

Malate dehydrogenase plays crucial roles in energy homeostasis, plant development and cold and salt tolerance, as it mediates the reversible conversion of malate to oxaloacetate. However, the evolutionary pattern of MDH genes in apple remains elusive. In this study, a total of 20 MDH genes were identified from the "Golden Delicious" apple draft genome. We revealed the physiological and biochemical properties, gene structure, and conserved motifs of MdMDH genes. Chromosomal localization and Ka/Ks ratio analysis of MdMDH genes revealed different selective pressures acted on duplicated MdMDH genes. Exploration of the phylogenetic relationships revealed six clades and similar frequencies between old and recent duplications, and significant differences in the evolutionary rates of the MDH gene family were observed. One MdMDH gene, MDP0000807458, which was highly expressed during apple fruit development and flower bud differentiation, was under positive selection. Thus, we speculated that MDP0000807458 is a likely candidate gene involved in regulation of flower bud differentiation and organic acid metabolism in apple fruits. This study provides a foundation for improved understanding of the molecular evolution of MdMDH genes and further facilitates the functional analysis of MDP0000807458 to unravel its exact role in flower bud differentiation and organic acid metabolism.


Assuntos
Evolução Molecular , Malato Desidrogenase/genética , Malus/genética , Proteínas de Plantas/genética , Genoma de Planta , Malato Desidrogenase/metabolismo , Malus/classificação , Malus/enzimologia , Filogenia , Proteínas de Plantas/metabolismo , Seleção Genética
18.
ACS Synth Biol ; 7(10): 2391-2402, 2018 10 19.
Artigo em Inglês | MEDLINE | ID: mdl-30216049

RESUMO

α-Amyrin is a plant-derived pentacyclic triterpenoid, with a lot of important physiological and pharmacological activities. The formation of α-amyrin from (3 S)-2,3-oxidosqualene is catalyzed by α-amyrin synthase (α-AS), a member of the oxidosqualene cyclase (OSC) protein family. However, α-amyrin is not yet commercially developed due to its extremely low productivity in plants. The engineered Saccharomyces cerevisiae with efficient α-amyrin production pathway could be used as an alternative and sustainable solution to produce α-amyrin from renewable raw materials. To efficiently improve α-amyrin production in S. cerevisiae, we identified two α-ASs, EjAS and MdOSC1 from Eriobotrya japonica and Malus × domestica, respectively, through strict bioinformatics screening criteria and phylogenetic analysis. The specific activities of purified EjAS and MdOSC1 were 0.0032 and 0.0293 µmol/min/mg, respectively. EjAS produced α-amyrin and ß-amyrin at a ratio of 17:3, MdOSC1 produced α-amyrin, ß-amyrin and lupeol at a ratio of 86:13:1, indicating MdOSC1 had significantly higher specific activity and higher ratio of α-amyrin than EjAS. Furthermore, MdOSC1 was introduced into S. cerevisiae combining with the increased supply of (3 S)-2,3-oxidosqualene to achieve the encouraging α-amyrin production, and the titer of α-amyrin achieved 11.97 ± 0.61 mg/L, 5.8 folds of the maximum production reported.


Assuntos
Transferases Intramoleculares/genética , Ácido Oleanólico/análogos & derivados , Proteínas de Plantas/genética , Saccharomyces cerevisiae/metabolismo , Sequência de Aminoácidos , Eriobotrya/enzimologia , Cromatografia Gasosa-Espectrometria de Massas , Transferases Intramoleculares/classificação , Transferases Intramoleculares/metabolismo , Malus/enzimologia , Engenharia Metabólica/métodos , Ácido Oleanólico/análise , Ácido Oleanólico/biossíntese , Filogenia , Proteínas de Plantas/classificação , Proteínas de Plantas/metabolismo , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/química , Proteínas Recombinantes/isolamento & purificação , Alinhamento de Sequência , Esqualeno/análogos & derivados , Esqualeno/metabolismo
19.
Plant Physiol Biochem ; 132: 320-332, 2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30248518

RESUMO

Long-chain acyl-CoA synthetases (LACSs) are members of the acyl-activating enzyme superfamily that have important roles in lipid synthesis and storage, fatty acid catabolism, vectorial acylation, and synthesis of cutin and wax. Here, 11 apple MdLACS genes were identified based on the Malus × domestica reference genome, clustered into six groups and mapped to ten chromosomes. Multiple sequence alignment and conserved motifs analyses showed that the sequences of the AtLACS and MdLACS proteins were highly conserved. A cis-element analysis in the promoter regions of the MdLACS genes revealed various elements related to stress responsiveness and plant hormones. Subsequently, expression analysis demonstrated that the MdLACS genes had different expression profiles in different tissues in response to various abiotic stresses. To further study the function of MdLACS genes in apple, MdLACS1 was isolated to identify its basic function, which the function of MdLACS1 in response to apple abiotic stress resistance was determined by the transgenic method. The results showed the MdLACS1 enhanced tolerance to polyethylene glycol, salt, and abscisic acid in the apple callus, suggesting that MdLACS1 is an important regulator in response to abiotic stresses. Finally, the functional interoperability network among the MdLACS proteins was predicted and analyzed, which could the understanding of the possible interactions among proteins and genes regulatory networks concerned with wax biosynthesis and regulatory mechanisms in response to abiotic stresses in apple.


Assuntos
Coenzima A Ligases/genética , Regulação da Expressão Gênica de Plantas , Genoma de Planta , Malus/enzimologia , Malus/genética , Estresse Fisiológico/genética , Ácido Abscísico/farmacologia , Motivos de Aminoácidos , Sequência de Aminoácidos , Arabidopsis/metabolismo , Cromossomos de Plantas/genética , Coenzima A Ligases/química , Coenzima A Ligases/metabolismo , Sequência Conservada/genética , Evolução Molecular , Ácidos Graxos/biossíntese , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Genes de Plantas , Filogenia , Proteínas de Plantas/química , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Plantas Geneticamente Modificadas , Polietilenoglicóis/farmacologia , Regiões Promotoras Genéticas/genética , Mapas de Interação de Proteínas/efeitos dos fármacos , Cloreto de Sódio/farmacologia , Estresse Fisiológico/efeitos dos fármacos , Ceras/metabolismo
20.
Food Chem ; 269: 638-643, 2018 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-30100483

RESUMO

This work describes a TLC-coupled bioautographic assay suitable for the separation and detection of apple polyphenol oxidase (PPO) inhibitors from natural extracts. PPO was immobilised in agar containing l-DOPA as substrate and 3-methyl-2-benzothiazolinone hydrazone hydrochloride (MBTH) to enhance colour development. The inhibition was detected as white spots on reddish background. Minimum amount of PPO inhibitors detected was 0.0125 µg of 4-hexylresorcinol, 0.025 µg of ascorbic acid, 0.5 µg of cysteine and 1 µg of kojic acid. The assay was compatible with normal and reverse phase TLC systems and allows detecting compounds that directly had action on the enzyme as well as agents that could convert quinones back to their reduced form. The chromatographic run evidenced the different nature of enzymatic browning inhibitory compounds from garlic and onion extracts. Using natural enzymes will provide a fast and cheap alternative for target specific exploration of natural enzymatic inhibitors.


Assuntos
Catecol Oxidase/química , Inibidores Enzimáticos/química , Malus/enzimologia , Ácido Ascórbico/química , Catecol Oxidase/isolamento & purificação , Hexilresorcinol/química , Reação de Maillard
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