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Expression Patterns of Genes Involved in Ascorbate-Glutathione Cycle in Aphid-Infested Maize (Zea mays L.) Seedlings.

ABSTRACT: Reduced forms of ascorbate (AsA) and glutathione (GSH) are among the most important non-enzymatic foliar antioxidants in maize (Zea mays L.). The survey was aimed to evaluate impact of bird cherry-oat aphid (Rhopalosiphum padi L.) or grain aphid (Sitobion avenae F.) herbivory on expression of genes related to ascorbate-glutathione (AsA-GSH) cycle in seedlings of six maize varieties (Ambrozja, Nana, Tasty Sweet, Touran, Waza, Z?ota Kar?owa), differing in resistance to the cereal aphids. Relative expression of sixteen maize genes encoding isoenzymes of ascorbate peroxidase (APX1, APX2, APX3, APX4, APX5, APX6, APX7), monodehydroascorbate reductase (MDHAR1, MDHAR2, MDHAR3, MDHAR4), dehydroascorbate reductase (DHAR1, DHAR2, DHAR3) and glutathione reductase (GR1, GR2) was quantified. Furthermore, effect of hemipterans' attack on activity of APX, MDHAR, DHAR and GR enzymes, and the content of reduced and oxidized ascorbate and glutathione in maize plants were assessed. Seedling leaves of more resistant Z. mays varieties responded higher elevations in abundance of target transcripts. In addition, earlier and stronger aphid-triggered changes in activity of APX, MDHAR, DHAR and GR enzymes, and greater modulations in amount of the analyzed antioxidative metabolites were detected in foliar tissues of highly resistant Ambrozja genotype in relation to susceptible Tasty Sweet plants.

SUBMITTER: Sytykiewicz H

PROVIDER: S-EPMC4813132 | biostudies-literature | 2016 Feb

REPOSITORIES: biostudies-literature

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Expression Patterns of Genes Involved in Ascorbate-Glutathione Cycle in Aphid-Infested Maize (Zea mays L.) Seedlings.

Sytykiewicz Hubert H

International journal of molecular sciences 20160223 3

Reduced forms of ascorbate (AsA) and glutathione (GSH) are among the most important non-enzymatic foliar antioxidants in maize (Zea mays L.). The survey was aimed to evaluate impact of bird cherry-oat aphid (Rhopalosiphum padi L.) or grain aphid (Sitobion avenae F.) herbivory on expression of genes related to ascorbate-glutathione (AsA-GSH) cycle in seedlings of six maize varieties (Ambrozja, Nana, Tasty Sweet, Touran, Waza, Złota Karłowa), differing in resistance to the cereal aphids. Relative ...[more]

PMID: 26907270

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Project description:The isoenzyme glutathione S-transferase (GST) I from maize (Zea mays) was cloned and expressed in Escherichia coli, and its catalytic mechanism was investigated by site-directed mutagenesis and dynamic studies. The results showed that the enzyme promotes proton dissociation from the GSH thiol and creates a thiolate anion with high nucleophilic reactivity by lowering the pK(a) of the thiol from 8.7 to 6.2. Steady-state kinetics fit well to a rapid equilibrium, random sequential Bi Bi mechanism, with intrasubunit modulation between the GSH binding site (G-site) and the electrophile binding site (H-site). The rate-limiting step of the reaction is viscosity-dependent, and thermodynamic data suggest that product release is rate-limiting. Five residues of GST I (Ser(11), His(40), Lys(41), Gln(53) and Ser(67)), which are located in the G-site, were individually replaced with alanine and their structural and functional roles in the 1-chloro-2,4-dinitrobenzene (CDNB) conjugation reaction were investigated. On the basis of steady-state kinetics, difference spectroscopy and limited proteolysis studies it is concluded that these residues: (1) contribute to the affinity of the G-site for GSH, as they are involved in side-chain interaction with GSH; (2) influence GSH thiol ionization, and thus its reactivity; (3) participate in k(cat) regulation by affecting the rate-limiting step of the reaction; and (4) in the cases of His(40), Lys(41) and Gln(53) play an important role in the structural integrity of, and probably in the flexibility of, the highly mobile short 3(10)-helical segment of alpha-helix 2 (residues 35-46), as shown by limited proteolysis experiments. These structural perturbations are probably transmitted to the H-site through changes in Phe(35) conformation. This accounts for the modulation of K(CDNB)(m) by His(40), Lys(41) and Gln(53), and also for the intrasubunit communication between the G- and H-sites. Computer simulations using CONCOORD were applied to maize GST I monomer and dimer structures, each with bound lactoylglutathione, and the results were analysed by the essential dynamics technique. Differences in dynamics were found between the monomer and the dimer simulations showing the importance of using the whole structure in dynamic analysis. The results obtained confirm that the short 3(10)-helical segment of alpha-helix 2 (residues 35-46) undergoes the most significant structural rearrangements. These rearrangements are discussed in terms of enzyme catalytic mechanism.

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Dataset Information

Expression Patterns of Genes Involved in Ascorbate-Glutathione Cycle in Aphid-Infested Maize (Zea mays L.) Seedlings.

Publications

Expression Patterns of Genes Involved in Ascorbate-Glutathione Cycle in Aphid-Infested Maize (Zea mays L.) Seedlings.

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