Olaf Döring

Oxidoreductases in plant plasma membranes.

Electron transporting oxidoreductases at biological membranes mediate several physiological processes. While such activities are well known and widely accepted as physiologically significant for other biological membranes, oxidoreductase activities found at the plasma membrane of plants are still being neglected. The ubiquity of the oxidoreductases in the plasma membrane suggests that the activity observed is of major importance in fact up to now no plant without redox activity at the plasmalemma is known. Involvement in proton pumping, membrane energization, ion channel regulation, iron reduction, nutrient uptake, signal transduction, and growth regulation has been proposed. However, positive proof for one of the numerous theories about the physiological function of the system is still missing. Evidence for an involvement in signalling and regulation of growth and transport activities at the plasma membrane is strong, but the high activity of the system displayed in some experiments also suggests function in defense against pathogens.

Molecular components and biochemistry of electron transport in plant plasma membranes (review).

It is worthwhile emphasizing the importance of electron transport across lipid membranes. Mitochondrial and electron transport in chloroplasts were elucidated in great detail many years ago. Plasma membrane-bound electron transfer may be involved in several processes such as membrane energization, signalling, regulation of transport and/or growth, and generation or scavenging of free radicals. We here give an overview of plasma membrane-bound electron transfer, of possible compounds of the electron transporting systems isolated from plasma membranes, and of their biochemical characteristics. Both the progress made in purification of redox enzymes and compounds and data from biochemical characterization of the activities found, support the discussion concerning models of the molecular structure of the electron transport systems of plant plasma membranes.

The effect of ascorbate free radical on the energy state of the plasma membrane of onion (Allium cepa L.) root cells: alteration of K+ efflux by ascorbate?

In this work we demonstrate the effect of different ascorbate redox states on the plasma membrane potential and on proton secretion of onion root cells. Ascorbate and dehydroascorbate induced plasmalemma hyperpolarization and stimulated acidification of the incubation medium. Both effects were transient and a repolarization of the membrane to the resting potential was reached within 10–15 min. The free radical of ascorbate had a permanent stimulatory effect on both parameters: a concentration dependent hyperpolarization of about 25 mV, partially inhibited by vanadate, could be observed immediately after addition. Medium acidification was increased by a factor of 20 and sustained for hours in the presence of the free radical. Additionally, ascorbate may increase the potassium permeability of the membrane, thus hyperpolarizing at low external potassium concentrations. Possible interactions of the free radical with the energization of the plasma membrane via an NAD(P)H-ascorbate free radical reductase (EC 1.6.5.4) will be discussed.

Are Plants Stacked Neutrophiles? Comparison of Pathogen-Induced Oxidative Burst in Plants and Mammals

Are plants stacked neutrophils? Curious question, one might think. However, after reading some up-to-date reviews on the topic of plant pathogen defense the idea becomes inescapable. In literature, on journal covers [e.g., Plant & Cell Physiol 38 (10), 1997], in congress presentations and, we fear, also in the minds of scientists, the NADPH oxidase complex known from neutrophil phagolysosomes has been transplanted, complete with all its regulatory components, to the plasma membrane (PM) of plants. In other words, the plant defense system is proposed to be a clone of the mammalian’s oxidative burst.

To Be or Not to Be — A Question of Plasma Membrane Redox?

At the beginning of the 1980s scientists working on plasma membrane (PM)-bound oxidoreductase (redox) activities had to go through thorough discussion in order to persuade their colleagues that electron transport at the PM of plants was no mere fiction but reality. Nowadays, the need to prove the sheer existence of the system(s) no longer exists; however, because the PM redox system of intact plants has, up till now, refused detection without electron acceptors that were added to the experimental solution, the most difficult of the old arguments of doubt still lingers on: What might such a system be good for? Today there are some answers to this question (Bienfait and Luttge 1988; Crane et al. 1991a,b; Navas 1991; Rubinstein 1992; see also the complete issue of Protoplasma 184 (1–4), 1995); yet it appears that there are too many answers and no plain way to decide which one is right. In experiments with PM-redox activity an impermeant electron acceptor, e.g. ferricyanide, is almost always added to the experimental solution, and it is therefore not clear what the system would do, if anything, without such a treatment.

The role of active oxygen in iron tolerance of rice (Oryza sauva L.)

SummaryIron tolerance of rice (Oryza sativa L.) was investigated using an oxygen depleted hydroculture system. Treatment with high concentrations of Fe2+ induced yellowing and bronzing symptoms as well as iron coatings at the root surface. Root and shoot growth were inhibited by increasing iron concentration in the medium. All symptoms were more pronounced in an iron sensitive cultivar (IR 64) compared to an iron tolerant one (IR 9764-45-2). Superoxide dismutase and peroxidase activity of root extracts of IR 97 were about twice that of IR 64 in untreated control plants. No significant increase of peroxidase activity was detected with increasing iron concentration in the medium. Catalase activity of IR 64 was slightly higher than that of IR 97, independent of iron concentration.

Inhibitors of the plasma membrane redox system of Zea mays L. roots. The vitamin K antagonists dicumarol and warfarin

The action of the 4-hydroxycoumarins dicumarol and warfarin, antagonists of probable vitamin K type components of the plasma membrane electron-transport system, on plasma membrane redox activity of intact maize roots was compared. Both effectors inhibited electron transfer to extracellular hexacyanoferrate III. While the effect of the strongly lipophilic dicumarol on the electron-transport system was irreversible by rinsing, the inhibition caused by the hydrophilic warfarin could be reverted completely by exchange of the incubation medium. We take these results as possible evidence for the integration of dicumarol into the plasma membrane. The action of warfarin may be confined to enzymic sites freely accessible from the aqueous apoplasmic solution.

INHIBITION OF MAIZE (ZEA MAYS L.) ROOT PLASMA MEMBRANE-BOUND REDOX ACTIVITIES BY COUMARINS

Modulation of plasma membrane-bound NADH:hexacyanoferrate III oxidoreductase activities by dicumarol and warfarin was investigated with plasma membrane vesicles of Zea mays L. (cv. Sil Anjou 18) roots, prepared by aqueous two phase partitioning. Vesicles were about 65% right-side out orientated as demonstrated by enzyme latency of vanadate sensitive ATPase activity. Dicumarol or warfarin, respectively, inhibited NADH:hexacyanoferrate III oxidoreductase activity in a concentration-dependent manner and inhibition could be reversed partially by addition of quinones

The effect of calcium and calmodulin inhibitors on NADH oxidation by isolated plasma membrane vesicles preloaded with NADH

Abstract Right-side-out plasma membrane vesicles of soybean (Glycine max (L.) Merr.) hypocotyl were loaded with NADH by electroporation. After washing it was possible to measure a decrease of NADH within the vesicles. Addition of the membrane impermeant electron acceptor hexacyanoferrate III to the experimental solution caused an increase of NADH oxidation, while the acceptor was reduced. NADH oxidation and reduction of the external acceptor were influenced by calcium and by calmoudin inhibitors. Ca2+ up to 100 μM stimulates the NADH:hexacyanoferrate III oxidoreductase activity, while millimolar concentrations inhibit. The stimulation is Ca2+ specific and sensitive to calmodulin inhibitors. The membrane impermeant inhibitor, calmidazolium, is by far the strongest one, while the permeant compounds fluphenazine, trifluoperazine and W7 are less active. Compound W5 turned out to be practically inactive.

Interaction between electron transport at the plasma membrane and nitrate uptake by maize (Zea mays L.) roots

SummaryIn the present study nitrate uptake by maize (Zea mays L.) roots was investigated in the presence or absence of ferricyanide (hexacyanoferrate III) or dicumarol. Nitrate uptake caused an alkalization of the medium. Nitrate uptake of intact maize seedlings was inhibited by ferricyanide while the effect of dicumarol was not very pronounced. Nitrite was not detected in the incubation medium, neither with dicumarol-treated nor with control plants after application of 100 μM nitrate to the incubation solution. In a second set of experiments interactions between nitrate and ferricyanide were investigated in vivo and in vitro. Nitrate (1 or 3 mM) did neither influence ferricyanide reductase activity of intact maize roots nor NADH-ferricyanide oxidoreductase activity of isolated plasma membranes. Nitrate reductase activity of plasma-membrane-enriched fractions was slightly stimulated by 25 μM dicumarol but was not altered by 100 μM dicumarol, while NADH-ferricyanide oxidoreductase activity was inhibited in the presence of dicumarol. These data suggest that plasma-membrane-bound standard-ferricyanide reductase and nitrate reductase activities of maize roots may be different. A possible regulation of nitrate uptake by plasmalemma redox activity, as proposed by other groups, is discussed.