TY - JOUR A1 - Sakuraba, Yasuhito A1 - Bülbül, Selin A1 - Piao, Weilan A1 - Choi, Giltsu A1 - Paek, Nam-Chon T1 - Arabidopsis EARLY FLOWERING3 increases salt tolerance by suppressing salt stress response pathways JF - The plant journal KW - Arabidopsis thaliana KW - salt stress response KW - EARLY FLOWERING3 (ELF3) KW - reactive oxygen species KW - PHYTOCHROME INTERACTING FACTOR4 (PIF4) KW - JUNGBRUNNEN1 (JUB1/ANAC042) KW - ORESARA1 (ORE1/ANAC092) KW - SAG29 Y1 - 2017 U6 - https://doi.org/10.1111/tpj.13747 SN - 0960-7412 SN - 1365-313X VL - 92 SP - 1106 EP - 1120 PB - Wiley CY - Hoboken ER - TY - JOUR A1 - Petrov, Veselin A1 - Hille, Jacques A1 - Müller-Röber, Bernd A1 - Gechev, Tsanko S. T1 - ROS-mediated abiotic stress-induced programmed cell death in plants JF - Frontiers in plant science N2 - During the course of their ontogenesis plants are continuously exposed to a large variety of abiotic stress factors which can damage tissues and jeopardize the survival of the organism unless properly countered. While animals can simply escape and thus evade stressors, plants as sessile organisms have developed complex strategies to withstand them. When the intensity of a detrimental factor is high, one of the defense programs employed by plants is the induction of programmed cell death (PCD). This is an active, genetically controlled process which is initiated to isolate and remove damaged tissues thereby ensuring the survival of the organism. The mechanism of PCD induction usually includes an increase in the levels of reactive oxygen species (ROS) which are utilized as mediators of the stress signal. Abiotic stress-induced PCD is not only a process of fundamental biological importance, but also of considerable interest to agricultural practice as it has the potential to significantly influence crop yield. Therefore, numerous scientific enterprises have focused on elucidating the mechanisms leading to and controlling PCD in response to adverse conditions in plants. This knowledge may help develop novel strategies to obtain more resilient crop varieties with improved tolerance and enhanced productivity. The aim of the present review is to summarize the recent advances in research on ROS-induced PCD related to abiotic stress and the role of the organelles in the process. KW - abiotic stress KW - programmed cell death KW - reactive oxygen species KW - signal transduction KW - stress adaptation Y1 - 2015 U6 - https://doi.org/10.3389/fpls.2015.00069 SN - 1664-462X VL - 6 PB - Frontiers Research Foundation CY - Lausanne ER - TY - JOUR A1 - Krylov, Andrey. V. A1 - Adamzig, H. A1 - Walter, A. D. A1 - Loechel, B. A1 - Kurth, E. A1 - Pulz, O. A1 - Szeponik, Jan A1 - Wegerich, Franziska A1 - Lisdat, Fred T1 - Parallel generation and detection of superoxide and hydrogen peroxide in a fluidic chip JF - Sensors and actuators : B, Chemical N2 - A fluidic chip system was developed, which combines a stable generation of superoxide radicals and hydrogen peroxide with their sensorial detection. The generation of both reactive oxygen species was achieved by immobilization of xanthine oxidase on controlled pore glass in a reaction chamber. Antioxidants can be introduced into the fluidic chip system by means of mixing chamber. The detection of both species is based on the amperometric principle using a biosensor chip with two working electrodes. As sensing protein for both electrodes cytochrome c was used. The novel system was designed for the quantification of the antioxidant efficiency of different potential scavengers of the respective reactive species in an aqueous medium. Several model antioxidants such as ascorbic acid or catalase have been tested under flow conditions. KW - biosensor KW - cytochrome c KW - flow system KW - reactive oxygen species KW - antioxidant Y1 - 2006 U6 - https://doi.org/10.1016/j.snb.2005.11.062 SN - 0925-4005 VL - 119 IS - 1 SP - 118 EP - 126 PB - Elsevier CY - Lausanne ER - TY - THES A1 - Foti, Alessandro T1 - Characterization of the human aldehyde oxidase T1 - Charakterisierung der menschlichen Aldehydoxidase BT - Studies on the FAD active site and ROS generation N2 - In this work the human AOX1 was characterized and detailed aspects regarding the expression, the enzyme kinetics and the production of reactive oxygen species (ROS) were investigated. The hAOX1 is a cytosolic enzyme belonging to the molybdenum hydroxylase family. Its catalytically active form is a homodimer with a molecular weight of 300 kDa. Each monomer (150 kDa) consists of three domains: a N-terminal domain (20 kDa) containing two [2Fe-2S] clusters, a 40 kDa intermediate domain containing a flavin adenine dinucleotide (FAD), and a C-terminal domain (85 kDa) containing the substrate binding pocket and the molybdenum cofactor (Moco). The hAOX1 has an emerging role in the metabolism and pharmacokinetics of many drugs, especially aldehydes and N- heterocyclic compounds. In this study, the hAOX1 was hetereogously expressed in E. coli TP1000 cells, using a new codon optimized gene sequence which improved the expressed protein yield of around 10-fold compared to the previous expression systems for this enzyme. To increase the catalytic activity of hAOX1, an in vitro chemical sulfuration was performed to favor the insertion of the equatorial sulfido ligand at the Moco with consequent increased enzymatic activity of around 10-fold. Steady-state kinetics and inhibition studies were performed using several substrates, electron acceptors and inhibitors. The recombinant hAOX1 showed higher catalytic activity when molecular oxygen was used as electron acceptor. The highest turn over values were obtained with phenanthridine as substrate. Inhibition studies using thioridazine (phenothiazine family), in combination with structural studies performed in the group of Prof. M.J. Romão, Nova Universidade de Lisboa, showed a new inhibition site located in proximity of the dimerization site of hAOX1. The inhibition mode of thioridazine resulted in a noncompetitive inhibition type. Further inhibition studies with loxapine, a thioridazine-related molecule, showed the same type of inhibition. Additional inhibition studies using DCPIP and raloxifene were carried out. Extensive studies on the FAD active site of the hAOX1 were performed. Twenty new hAOX1 variants were produced and characterized. The hAOX1 variants generated in this work were divided in three groups: I) hAOX1 single nucleotide polymorphisms (SNP) variants; II) XOR- FAD loop hAOX1 variants; III) additional single point hAOX1 variants. The hAOX1 SNP variants G46E, G50D, G346R, R433P, A439E, K1231N showed clear alterations in their catalytic activity, indicating a crucial role of these residues into the FAD active site and in relation to the overall reactivity of hAOX1. Furthermore, residues of the bovine XOR FAD flexible loop (Q423ASRREDDIAK433) were introduced in the hAOX1. FAD loop hAOX1 variants were produced and characterized for their stability and catalytic activity. Especially the variants hAOX1 N436D/A437D/L438I, N436D/A437D/L438I/I440K and Q434R/N436D/A437D/L438I/I440K showed decreased catalytic activity and stability. hAOX1 wild type and variants were tested for reactivity toward NADH but no reaction was observed. Additionally, the hAOX1 wild type and variants were tested for the generation of reactive oxygen species (ROS). Interestingly, one of the SNP variants, hAOX1 L438V, showed a high ratio of superoxide prodction. This result showed a critical role for the residue Leu438 in the mechanism of oxygen radicals formation by hAOX1. Subsequently, further hAOX1 variants having the mutated Leu438 residue were produced. The variants hAOX1 L438A, L438F and L438K showed superoxide overproduction of around 85%, 65% and 35% of the total reducing equivalent obtained from the substrate oxidation. The results of this work show for the first time a characterization of the FAD active site of the hAOX1, revealing the importance of specific residues involved in the generation of ROS and effecting the overall enzymatic activity of hAOX1. The hAOX1 SNP variants presented here indicate that those allelic variations in humans might cause alterations ROS balancing and clearance of drugs in humans. N2 - Aldehydoxidasen (AOX) sind Molybdo-enzyme, die durch breite Substratspezifität gekennzeichnet sind, aromatische/aliphatische Aldehyde in die entsprechenden Carbonsäuren oxidieren und verschiedene heteroaromatische Ringe hydroxylieren. Die Enzyme verwenden Sauerstoff als terminalen Elektronenakzeptor und produzieren reduzierte Sauerstoffspezies während des Umsatzes. Die physiologische Funktion von Säugetier-AOX-Isoenzymen ist noch unklar, aber menschliches AOX (hAOX1) ist ein Enzym von Phase-I-Wirkstoff-Metabolismus. Weiterhin, wurden zahlreiche Einzelnukleotidpolymorphismen (SNP) und weitere hAOX1-Mutanten im hAOX1-Gen identifiziert. SNPs sind eine Hauptquelle für die interindividuelle Variabilität in der menschlichen Population, und SNP-basierte Aminosäureaustausche in hAOX1 modulieren die katalytische Funktion des Enzyms entweder positiv oder negativ. In diesem Bericht haben wir zehn neue SNPs ausgewählt, die zu Aminosäureaustauschen in der Nähe der FAD-Cofaktor von hAOX1 führen und die gereinigten Enzyme nach heterologen Expression in Escherichia coli charakterisieren. Darüber hinaus haben wir zehn weitere FAD-Varianten produziert. Die hAOX1-Varianten wurden sorgfältig durch quantitative Unterschiede in ihrer Fähigkeit zur Herstellung von Superoxidradikal charakterisiert. ROS repräsentieren markante Schlüsselmoleküle in physiologischen und pathologischen Zuständen in der Zelle. Unsere Daten zeigen signifikante Veränderungen der Superoxid-Anionenproduktion unter den Varianten. Insbesondere führte der Rest L438 in der Nähe des Isoalloxanzinringes des FAD-Cofaktors zu einer erhöhten Superoxid-Radikalproduktion von 75-85%. In Anbetracht der hohen Toxizität des Superoxid-Anions in der Zelle ist die hAOX1-L438V SNP-Variante ein eventueller Kandidat für kritische oder pathologische Rollen dieser natürlichen Variante innerhalb der menschlichen Population. KW - aldehyde KW - oxidase KW - ROS KW - reactive oxygen species Y1 - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:517-opus4-410107 ER - TY - JOUR A1 - Dongmo, Saustin A1 - Leyk, Janina A1 - Dosche, Carsten A1 - Richter-Landsberg, Christiane A1 - Wollenberger, Ursula A1 - Wittstock, Gunther T1 - Electrogeneration of O-2(center dot-) and H2O2 Using Polymer-modified Microelectrodes in the Environment of Living Cells JF - Electroanalysis : an international journal devoted to fundamental and practical aspects of electroanalysis N2 - Microelectrodes modified with electropolymerized plumbagin (PLG) were used for the generation of superoxide radical (O-2(center dot-)) and hydrogen peroxide (H2O2) during oxygen reduction reaction (ORR) in an aqueous medium, specifically in serum-free cell culture media. This is enabled by the specific design of a polymer film on the microelectrode. The generation and diffusion of O-2(center dot-) during electrocatalytic ORR at a positionable PLG polymer-modified microelectrode was followed by fluorescence microscopy with the selective dye 4-chloro-7-nitrobenzo-2-oxa-1,3-diazole (NBD-Cl) and by amperometric detection using a cytochrome c-modified electrode at + 0.13 V. H2O2 production, either by direct oxygen reduction or as product of O-2(center dot-) disproportionation, was monitored by the reaction with Amplex UltraRed. The PLG polymer-modified microelectrodes were used to expose mammalian B6-RPE07 retinal cells to defined local fluxes of reactive oxygen species (ROS), and cellular responses and morphological alterations were observed. The use of a controllable source of ROS opens many possibilities to study how living cells respond to the presence of a certain flux of specific ROS. KW - reactive oxygen species KW - microelectrode KW - scanning electrochemical microscopy KW - biosensor KW - polymer-modified electrode KW - oxygen reduction reaction Y1 - 2016 U6 - https://doi.org/10.1002/elan.201600267 SN - 1040-0397 SN - 1521-4109 VL - 28 SP - 2400 EP - 2407 PB - Wiley-VCH CY - Weinheim ER - TY - JOUR A1 - Allu, Annapurna Devi A1 - Soja, Aleksandra Maria A1 - Wu, Anhui A1 - Szymanski, Jedrzej A1 - Balazadeh, Salma T1 - Salt stress and senescence: identification of cross-talk regulatory components JF - Journal of experimental botany N2 - Leaf senescence is an active process with a pivotal impact on plant productivity. It results from extensive signalling cross-talk coordinating environmental factors with intrinsic age-related mechanisms. Although many studies have shown that leaf senescence is affected by a range of external parameters, knowledge about the regulatory systems that govern the interplay between developmental programmes and environmental stress is still vague. Salinity is one of the most important environmental stresses that promote leaf senescence and thus affect crop yield. Improving salt tolerance by avoiding or delaying senescence under stress will therefore play an important role in maintaining high agricultural productivity. Experimental evidence suggests that hydrogen peroxide (H2O2) functions as a common signalling molecule in both developmental and salt-induced leaf senescence. In this study, microarray-based gene expression profiling on Arabidopsis thaliana plants subjected to long-term salinity stress to induce leaf senescence was performed, together with co-expression network analysis for H2O2-responsive genes that are mutually up-regulated by salt induced-and developmental leaf senescence. Promoter analysis of tightly co-expressed genes led to the identification of seven cis-regulatory motifs, three of which were known previously, namely CACGTGT and AAGTCAA, which are associated with reactive oxygen species (ROS)-responsive genes, and CCGCGT, described as a stress-responsive regulatory motif, while the others, namely ACGCGGT, AGCMGNC, GMCACGT, and TCSTYGACG were not characterized previously. These motifs are proposed to be novel elements involved in the H2O2-mediated control of gene expression during salinity stress-triggered and developmental senescence, acting through upstream transcription factors that bind to these sites. KW - Arabidopsis KW - hydrogen peroxide KW - longevity KW - reactive oxygen species KW - salt stress KW - senescence KW - signal cross-talk KW - transcription factor Y1 - 2014 U6 - https://doi.org/10.1093/jxb/eru173 SN - 0022-0957 SN - 1460-2431 VL - 65 IS - 14 SP - 3993 EP - 4008 PB - Oxford Univ. Press CY - Oxford ER -