Light-activated proton-motive force generation in lipid vesicles containing cytochrome b-c1 complex and bacterial reaction centres

(1) Purified bovine heart mitochondrial cytochrome b-c1 complex (ubiquinone-cytochrome c oxidoreductase) and photosynthetic reaction centres isolated from Rhodopseudomonas sphaeroides strain R-26 have been incorporated into lipid vesicles. In the presence of cytochrome c and ubiquinone-2, light activation caused a cyclic electron transfer involving both components. (2) Since cytochrome c is added outside the vesicles, it is both reduced by the cytochrome b-c1 complex and oxidised by the reaction centre on the outside of the vesicles. Ubiquinone-2, however, is reduced by the reaction centres at a site in contact with the inside of the vesicles, but the reduced form, ubiquinol-2, is oxidised by the cytochrome b-c1 complex at a site in contact with the outer aqueous phase. (3) In the presence of valinomycin plus K+, initiation of cyclic electron flow causes protons to move from inside the vesicles to the outer medium and the H +/2e- ratio was calculated to be close to 4.     

The suitability of some plant hosts for the development of the peach-potato aphid, Myzus persicae (Sulzer)

Myzus persicae develop well on Brassica species, particularly those that grow rapidly, such as mustard and turnip, but much less well on sugar beet, spinach and lettuce. They usually develop better on young, rapidly growing leaves than on mature leaves. Two clones of M. persicae behaved similarly on the host plants tested.

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Zusammenfassung Die Entwicklung von Myzus persicae-Alienicolen auf den Sekundärwirten Kohlrüben, Kopfkohl, Chinakohl, Brokkoli, Blumenkohl, Senf, Raps, Grünkohl sowie Zuckerrübe, Spinat und Salat wurde durch wiederholte Erfassung der durchschnittlichen und totalen Larvenproducktion einzelner Jungfern sowie ihrer Mortalität in Gewächshausversuchen verfolgt und mit Auszählung des Befalls von Freilandpflanzen verglichen. Myzus persicae entwickelte sich gut an Brassica-Arten, besonders an raschwüchsigen wie Senf und Kohlrüben, aber viel weniger gut an Zuckerrübe, Spinat and Salat. Sie entwickelt sich gewöhnlich an jungen schnell wachsenden Blättern besser als an ausgereiften. Zwei Klone von Myzus persicae verhielten sich an den geprüften Wirtspflanzen ähnlich.

     

Gravitropically-stimulated seedlings show autotropism in weightlessness

In a spaceflight experiment, autotropism by oat ( Avena sativa L.) coleoptiles following gravitropic responses was prominent in weightlessness: counter-reactions led to the straightening of the curved coleoptiles. This was not the case during clinorotation on earth. The autotropic reactions appeared to be related to the stimulus received during the stimulus period, i.e. the greater the response the greater the autotropic counter-reaction. Previous models of the gravitropic system which predicted that coleoptiles would not straighten in weightlessness are disproved. A modification to one of the models is proposed which includes the autotropic response observed in spaceflight. The nature of the counter-reactions in the absence of gravitropic stimulation is discussed.     

Quantitative electron-paramagnetic-resonance measurements of the electron-transfer components of the photosystem-I reaction centre. The reaction-centre chlorophyll (P700), the primary electron acceptor X and bound iron-sulphur centre A.

An e.p.r. spectrum of the reduced form of the electron-transport component (X), thought to be the primary electron acceptor of Photosystem I, was obtained. By using line-shape simulations of this component and the free-radical e.p.r. signal I of the oxidized reaction-centre chlorophyll (P700), it was possible to determine the ratio of the number of electron spins to which these signals correspond in Photosystem-I particles under a variety of conditions. On illumination at cryogenic temperatures of Photosystem-I preparations, in which both bound iron-sulphur centres A and B were reduced, the measured ratio of free radical to component X varied between 1.04 and 2.23, with an average value of 1.54 +/- 0.18 where a Gaussian line-shape is assumed for the component-X signal in the simulation. The error in this measurement is estimated to be up to 50%. In a similar way component X and centre A of the bound iron-sulphur protein were quantified, the ratio between these two components varying between 1.26 and 0.61 with an average value of 0.75 +/- 0.06. These results indicate that the quantitative relationship, in terms of net electron spins, between centre A, component X and P700 is of the order to be expected if component X is indeed the primary electron acceptor in Photosystem I and a component of the photosynthetic electron-transport chain.     

The reactions of hydrogen peroxide with bovine cytochrome c oxidase

Oxidised cytochrome c oxidase is known to react with two molecules of hydrogen peroxide to form consecutively 607 nm 'Peroxy' and 580-nm 'Ferryl' species. These are widely used as model compounds for the equivalent P and F intermediates of the catalytic cycle. However, kinetic analysis of the reaction with H(2)O(2) in the pH range 6.0-9.0 reveals a more complex situation. In particular, as the pH is lowered, a 580-nm compound can be formed by reaction with a single H(2)O(2). This species, termed F(&z.rad;), is spectrally similar, but not identical, to F. The reactions are equivalent to those previously reported for the bo type quinol oxidase from Escherichia coli (T. Brittain, R.H. Little, C. Greenwood, N.J. Watmough, FEBS Lett. 399 (1996) 21-25) where it was proposed that F(&z.rad;) is produced directly from P. However, in the bovine oxidase F(&z.rad;) does not appear in samples of the 607-nm form, P(M), produced by CO/O(2) treatment, even at low pH, although this form is shown to be identical to the H(2)O(2)-derived P state, P(H), on the basis of spectral characteristics and kinetics of reaction with H(2)O(2). Furthermore, lowering the pH of a sample of P(M) or P(H) generated at high pH results in F(&z.rad;) formation only on a minutes time scale. It is concluded that P and F(&z.rad;) are not in a rapid, pH-dependent equilibrium, but instead are formed by distinct pathways and cannot interconvert in a simple manner, and that the crucial difference between them lies in their patterns of protonation.     

Reaction of bovine cytochrome c oxidase with hydrogen peroxide produces a tryptophan cation radical and a porphyrin cation radical

Oxidized bovine cytochrome c oxidase reacts with hydrogen peroxide to generate two electron paramagnetic resonance (EPR) free radical signals (Fabian, M., and Palmer, G. (1995) Biochemistry 34, 13802-13810). These radicals are associated with the binuclear center and give rise to two overlapped EPR signals, one signal being narrower in line width (DeltaHptp = 12 G) than the other (DeltaHptp = 45 G). We have used electron nuclear double resonance (ENDOR) spectrometry to identify the two different chemical species giving rise to these two EPR signals. Comparison of the ENDOR spectrum associated with the narrow signal with that of compound I of horseradish peroxidase (formed by reaction of that enzyme with hydrogen peroxide) demonstrates that the two species are virtually identical. The chemical species giving rise to the narrow signal is therefore identified as an exchange-coupled porphyrin cation radical similar to that formed in horseradish peroxidase compound I. Comparison of the ENDOR spectrum of compound ES (formed by the reaction of hydrogen peroxide with cytochrome c peroxidase) with that of the broad signal indicates that the chemical species giving rise to the broad EPR signal in cytochrome c oxidase is probably an exchange coupled tryptophan cation radical. This is substantiated using H(2)O/D(2)O solvent exchange experiments where the ENDOR difference spectrum of the broad EPR signal of cytochrome c oxidase shows a feature consistent with hyperfine coupling to the exchangeable N(1) proton of a tryptophan cation radical.     

Dystroglycan overexpression in vivo alters acetylcholine receptor aggregation at the neuromuscular junction

Dystroglycan is a member of the transmembrane dystrophin glycoprotein complex in muscle that binds to the synapse-organizing molecule agrin. Dystroglycan binding and AChR aggregation are mediated by two separate domains of agrin. To test whether dystroglycan plays a role in receptor aggregation at the neuromuscular junction, we overexpressed it by injecting rabbit dystroglycan RNA into one- or two-celled Xenopus embryos. We measured AChR aggregation in myotomes by labeling them with rhodamine-alpha-bungarotoxin followed by confocal microscopy and image analysis. Dystroglycan overexpression decreased AChR aggregation at the neuromuscular junction. This result is consistent with dystroglycan competition for agrin without signaling AChR aggregation. It also supports the hypothesis that dystroglycan is not the myotube-associated specificity component, (MASC) a putative coreceptor needed for agrin to activate muscle-specific kinase (MuSK) and signal AChR aggregation. Dystroglycan was distributed along the surface of muscle membranes, but was concentrated at the ends of myotomes, where AChRs normally aggregate at synapses. Overexpressed dystroglycan altered AChR aggregation in a rostral-caudal gradient, consistent with the sequential development of neuromuscular synapses along the embryo. Increasing concentrations of dystroglycan RNA did not further decrease AChR aggregation, but decreased embryo survival. Development often stopped during gastrulation, suggesting an essential, nonsynaptic role of dystroglycan during this early period of development.     

Provisioning calls of the cooperatively breeding bell miner Manorina melanophrys encode sufficient information for individual discrimination

Acoustically-mediated individual discrimination has been the focus of much investigation in ornithology. For cooperatively breeding species, strong selection pressure favouring individual recognition during acts of altruism is predicted under many of the most interesting hypotheses proposed to account for helping behaviour. We investigated differences in 157 calls given by 12 different individuals as they provisioned nestlings, including both breeding and helper bell miners Manorina melanophrys (Meliphagidae), a cooperatively breeding honeyeater endemic to south-eastern Australia. Individual differences were apparent in all 15 call parameters analysed, many with a high level of repeatability. Moreover, the information capacity of mew calls allows as many as 515 different signatures to exist in the system. As many parameters showed strong sex differences, separate discriminant functions were used to predict individual identity within each sex. Five parameters were used in a function that correctly identified over 90% of female calls in both a training (n = 35) and test dataset (n = 11) not used to generate the function. Among males, a separate function used six parameters to correctly assign individual identity in 89.3% (n=84; training) and 77.8% (n=27; test) of cases. Three original parameters, including spectral and spatial characteristics, were highly correlated with functions predicting identity in both sexes. The accuracy of functions was not influenced by a signaller's sex, breeding status, or by sampling period which was spread over as much as two breeding seasons within individuals. Significant potential therefore exists for bell miners to use these simple provisioning calls in individual discrimination. If this is the case, call information also has the potential to be used by receivers as predicted by signalling hypotheses proposed to account for cooperative helping behaviour, such as the pay-to-stay and social prestige hypotheses.