Melon resistance to the aphid Aphis gossypii: behavioural analysis and chemical correlations with nitrogenous compounds

In the melon, the Vat (monogenic, dominant) resistance gene governs both an antixenotic reaction to the melon aphid Aphis gossypii Glover (Homoptera, Aphididae) and a resistance to non-persistent virus transmission, restricted to this vector species. We investigated the behavioural features and tissue localisation of the antixenosis resistance by the electrical penetration graph technique (EPG, DC system). We also compared the chemical composition in amino compounds and proteins of the phloem sap collected from two isogenic lines of melon (Cucumis melo L.), carrying the Vat gene or not. All behavioural and chemical data indicated that this resistance is constitutive. EPG analysis clearly showed that access to phloem, although delayed by alterations in pathway activities, was not impaired in terms of frequency of access or initiation of feeding. The most striking feature was, however, a very reduced duration of ingestion from phloem of resistant plants, making this compartment one of the tissues where the effects of the Vat gene are unambiguously expressed. This was confirmed by clear differential activity of phloem extracts in artificial no-choice bioassays. Chemical analyses have shown that phloem saps from the two isogenic lines were extremely similar in profiles of ninhydrin positive compounds, and contained a low total amount of free amino acids (less than 10 mM). Out of more than 40 distinguishable peaks in the chromatograms (protein and non-protein amino acids, as well as small peptides), only five differentiated the two genotypes. Two of them were increased in the resistant genotype: glutamic acid and a major unknown peak, probably a non-protein amino acid (different from pyrazolyl-alanine, a Cucumis-specific amino acid). The three others were depressed in resistant plants, and included the sulphur amino acid cystine and a peptide peak partly composed of the cysteine-containing peptide glutathione (reduced form). Sap collection also showed that phloem exudation rates, as well as total protein and glutathione levels, were depressed in phloem sap from resistant plants. Such data are all indicative of a modified phloem-sealing physiology, linked to sulfhydryl oxidation processes, in plants carrying the Vat gene. The originality of the mechanism of Vat resistance to aphids is discussed.     

Selective Sensitization of Zinc Finger Protein Oxidation by Reactive Oxygen Species through Arsenic Binding

Cysteine oxidation induced by reactive oxygen species (ROS) on redox-sensitive targets such as zinc finger proteins plays a critical role in redox signaling and subsequent biological outcomes. We found that arsenic exposure led to oxidation of certain zinc finger proteins based on arsenic interaction with zinc finger motifs. Analysis of zinc finger proteins isolated from arsenic-exposed cells and zinc finger peptides by mass spectrometry demonstrated preferential oxidation of C3H1 and C4 zinc finger configurations. C2H2 zinc finger proteins that do not bind arsenic were not oxidized by arsenic-generated ROS in the cellular environment. The findings suggest that selectivity in arsenic binding to zinc fingers with three or more cysteines defines the target proteins for oxidation by ROS. This represents a novel mechanism of selective protein oxidation and demonstrates how an environmental factor may sensitize certain target proteins for oxidation, thus altering the oxidation profile and redox regulation.     

Measurement and meaning of cellular thiol:disufhide redox status

The functional group of cysteine is a thiol group (SH) that, due to its chemical reactivity, is able to undergo a wide array of modifications each with the potential to confer a different property or function to the molecule harboring this residue. Most of these modifications involve the reversible oxidation of the thiol to sulfenic acid (SOH), and disulfide, including intra- and intermolecular disulfides between polypeptides and glutathione (glutathionylation). The reversibility of these oxidations allows thiol groups to serve as versatile chemical and structural transducing elements in several low molecular mass metabolites and proteins. A plethora of cellular functions such as DNA and protein synthesis, protein secretion, cytoskeleton architecture, differentiation, apoptosis, and anti-oxidant defense, are recognized to be modulated, at certain stage, by thiol–disulfide exchange mechanisms of redox active thiol groups. All organisms are equipped with enzymatic systems composed by NADPH-dependent reductases, redoxins, and peroxidases that provide kinetic control of global thiol-redox homeostasis as well as target selectivity. These redox systems are distributed in different subcellular compartments and are not in equilibrium with each other. In consequence, measuring cellular thiol–disulfide status represents a challenge for studies aimed to obtain dynamic and spatio-temporal resolution. This review provides a summary of the methods and tools available to quantify the thiol redox status of cells.     

Conserved cysteine residues in the mammalian lamin A tail are essential for cellular responses to ROS generation

Pre-lamin A and progerin have been implicated in normal aging, and the pathogenesis of age-related degenerative diseases is termed 'laminopathies'. Here, we show that mature lamin A has an essential role in cellular fitness and that oxidative damage to lamin A is involved in cellular senescence. Primary human dermal fibroblasts (HDFs) aged replicatively or by pro-oxidants acquire a range of dysmorphic nuclear shapes. We observed that conserved cysteine residues in the lamin A tail domain become hyperoxidized in senescent fibroblasts, which inhibits the formation of lamin A inter- and intramolecular disulfide bonds. Both in the absence of lamin A and in the presence of a lamin A cysteine-to-alanine mutant, which eliminates these cysteine residues (522, 588, and 591), mild oxidative stress induced nuclear disorganization and led to premature senescence as a result of decreased tolerance to ROS stimulators. Human dermal fibroblasts lacking lamin A or expressing the lamin A cysteine-to-alanine mutant displayed a gene expression profile of ROS-responsive genes characteristic of chronic ROS stimulation. Our findings suggest that the conserved C-terminal cysteine residues are essential for lamin A function and that loss or oxidative damage to these cysteine residues promotes cellular senescence.     

Chlorophyll a is the crucial redox sensor and transmembrane signal transmitter in the cytochrome b6f complex. Components and mechanisms of state transitions from the hydrophobic mismatch viewpoint

The cytochrome (cyt) b₆f complex is involved in the transmembrane redox signaling that triggers state transitions in cyanobacteria and chloroplasts. However, the components and molecular mechanisms are still unclear. In an attempt to solve this long-standing problem, we first focused on the unknown role of a single chlorophyll a (Chla) in cyt b₆f with a new approach based on Chla structural properties. Various b₆f X-ray crystal structures were analyzed to identify their differences, which correlate with differences in Chla molecular volume. We found that the distance of the Rieske [2Fe-2S] cluster to Chla correlates with the distance between a pair of residues at the Q_o-site and the distance between a pair of residues at the opposite membrane side. These correlations were accompanied by the rotation of a key peripheral residue and by changes in the hydrophobic thickness of cyt b₆f. Parallel analysis of cyt bc₁ crystal structures allowed us to conclude that Chla acts as the crucial redox sensor and transmembrane signal transmitter in b₆f for changes in the plastoquinone pool redox state. The hydrophobic mismatch induced by the changed hydrophobic thickness of cyt b₆f is the driving force for the structural reorganizations of the photosynthetic apparatus during induction and the progression of state transitions in cyanobacteria and chloroplasts. A mechanism for LHCII kinase activation in chloroplasts is also proposed. Our understanding of the dynamic structural changes in bc-complexes during turnover at the Q_o-site and state transitions is augmented by the time-sequence ordering of 56 bc crystal structures.     

The yeast autophagy protease Atg4 is regulated by thioredoxin

Autophagy is a membrane-trafficking process whereby double-membrane vesicles called autophagosomes engulf and deliver intracellular material to the vacuole for degradation. Atg4 is a cysteine protease with an essential function in autophagosome formation. Mounting evidence suggests that reactive oxygen species may play a role in the control of autophagy and could regulate Atg4 activity but the precise mechanisms remain unclear. In this study, we showed that reactive oxygen species activate autophagy in the model yeast Saccharomyces cerevisiae and unraveled the molecular mechanism by which redox balance controls Atg4 activity. A combination of biochemical assays, redox titrations, and site-directed mutagenesis revealed that Atg4 is regulated by oxidoreduction of a single disulfide bond between Cys338 and Cys394. This disulfide has a low redox potential and is very efficiently reduced by thioredoxin, suggesting that this oxidoreductase plays an important role in Atg4 regulation. Accordingly, we found that autophagy activation by rapamycin was more pronounced in a thioredoxin mutant compared with wild-type cells. Moreover, in vivo studies indicated that Cys338 and Cys394 are required for the proper regulation of autophagosome biogenesis, since mutation of these cysteines resulted in increased recruitment of Atg8 to the phagophore assembly site. Thus, we propose that the fine-tuning of Atg4 activity depending on the intracellular redox state may regulate autophagosome formation.     

Cu(II)-Binding N-Terminal Sequences of Human Proteins

Proteins anchor copper(II) ions mainly by imidazole from histidine residues located in different positions in the primary protein structures. However, the motifs with histidine in the first three N-terminal positions (His¹, His², and His³) show unique Cu(II)-binding properties, such as availability from the surface of the protein, high flexibility, and high Cu(II) exchangeability with other ligands. It makes such sequences beneficial for the fast exchange of Cu(II) between ligands. Furthermore, sequences with His¹ and His², thus, non-saturating the Cu(II) coordination sphere, are redox-active and may play a role in Cu(II) reduction to Cu(I). All human protein sequences deposited in UniProt Knowledgebase were browsed for those containing His¹, His², or His³. Proteolytically modified sequences (with the removal of a propeptide or Met residue) were taken for the analysis. Finally, the sequences were sorted out according to the subcellular localization of the proteins to match the respective sequences with the probability of interaction with divalent copper.     

Optimized Bicompartment Two Solution Cells for Effective and Stable Operation of Li–O2 Batteries

Lithium–oxygen batteries are in fact the only rechargeable batteries that can rival internal combustion engines, in terms of high energy density. However, they are still under development due to low‐efficiency and short lifetime issues. There are problems of side reactions on the cathode side, high reactivity of the Li anode with solution species, and consumption of redox mediators via reactions with metallic lithium. Therefore, efforts are made to protect/block the lithium metal anode in these cells, in order to mitigate side reactions. However, new approach is required in order to solve the problems mentioned above, especially the irreversible reactions of the redox mediators which are mandatory to these systems with the Li anode. Here, optimized bicompartment two solution cells are proposed, in which detrimental crossover between the cathode and anode is completely avoided. The Li metal anode is cycled in electrolyte solution containing fluorinated ethylene carbonate, in which its cycling efficiency is excellent. The cathode compartment contains ethereal solution with redox mediator that enables oxidation of Li₂O₂ at low potentials. The electrodes are separated by a solid electrolyte membrane, allowing free transport of Li ions. This approach increases cycle life of lithium oxygen cells and their energy efficiency.