Studies on the mechanism of photosystem II photoinhibition I. A two-step degradation of D1-protein

The role of D1-protein in photoinhibition was examined. Photoinhibition of spinach thylakoids at 20°C caused considerable degradation of D1-protein and a parallel loss of variable fluorescence, Q_B-independent electron flow and Q_B-dependent electron flow. The breakdown of D1-protein as well as the loss of variable fluorescence and Q_B-independent electron flow were largely prevented when thylakoids were photoinhibited at 0°C. The Q_B-dependent electron flow markedly decreased under the same conditions. This inactivation may represent the primary event in photoinhibition and could be the result of some modification at the Q_B-site of D1-protein. Evidence for this comes from fluorescence relaxation kinetics following photoinhibition at 0°C which indicate a partial inactivation of Q_A ^--reoxidation. These results support the idea of D1-protein breakdown during photoinhibition as a two step process consisting of an initial inactivation at the Q_B-site of the protein followed by its degradation. The latter is accompanied by the loss of PS II-reaction centre function.Abbreviations Asc ascorbate - p-BQ 1, 4-benzoquinone - DAD diaminodurene - DPC diphenylcarbazide - DQH₂ duroquinole - Fecy ferricyanide - MV methylviologen - Q_A primary quinone acceptor of PS II - Q_B secondary quinone acceptor of PS II - SiMo silicomolybdate     

Reductions in genetic variation inDrosophila andE. coli caused by selection at linked sites

Selection at linked sites has important consequences for the properties of neutral variation and for tests of the predictions of the neutral theory of molecular evolution. We review the theory of the effect of adaptive gene substitutions on neutral variability at linked sites (hitchhiking or selective sweeps) and discuss theoretical results on the effect of selection against deleterious alleles on variation at linked sites (background selection). InDrosophila melanogaster there is a clear relation between the frequency of recombination in a given region of the chromosome and the amount of natural variability in that region. Attempts to predict this relation have given rise to models of selective sweeps and background selection. We describe possible methods of discriminating between these models, and also discuss the probable strong influence of selective sweeps on variation in largely nonrecombining genomes, with particular reference toEscherichia coll. Finally we present some unresolved questions and possible directions for future research.     

Developmental and Regional Expression of NMDA Receptor Subtypes Containing the NR2D Subunit in Rat Brain

Abstract: The regional and developmental expression of NMDA receptors containing the NR2D subunit was analyzed on the level of the subunit mRNA and protein in rat brain. RNase protection experiments indicated that among two proposed splice variants of the NR2D subunit, only the NR2D-2 subunit is expressed. The regional distribution of the NR2D subunit protein was visualized with a newly developed NR2D-2 subunit-specific antiserum on brain sections using the histoblot technique. In adult brain, NR2D immunoreactivity was mainly restricted to diencephalic, mesencephalic, and brainstem structures. During postnatal development, the NR2D subunit was detected transiently in certain regions, such as the ventro-basal complex of the thalamus, hippocampus, inferior colliculus, and brainstem reticular formation, suggesting that NR2D subunit-containing receptors play a role in these brain areas only during development. The level of NR2D subunit mRNA and protein decreased during late postnatal development. However, significant levels of NR2D subunit mRNA and protein were present in adulthood, in particular, in the globus pallidus, thalamus, subthalamic nuclei, and superior colliculus. These results indicate a functional relevance for NMDA receptors containing the NR2D subunit in the developing and adult brain, although its expression in the adult brain is less prominent and restricted to a few brain areas.     

Degenerative Joint Disease and enthesopathies in a Bronze Age population from Sindou cave (Senaillac-Lauzes,Lot, France)

The anthropological characteristics of the people who lived during the cultural period of the Late Bronze Age in South West France still remain practically unknown because very few sites have provided skeletal remains which permit of an exhaustive study. The cave of Sindou is, in that sense, one of the scarce exceptions. Although the sample of Sindou cannot be considered as representative of the whole regional population (N=50), we studied the presence and severity of DJD and enthesopathies of microtraumatic origin with the aim of finding some data which contribute to the knowledge of several biological aspects of this human group. From the results of the comparisons of the Sindou remains with two different medieval samples, a great similarity is deduced for these skeletal markers, but the higher frequency and severity of Achilles tendon enthesopathy in Sindou is a probable index of a higher level of physical stress at this specific localisation.     

HSP25 in isolated perfused rat hearts: Localization and response to hyperthermia

Recent investigations concentrate on the correlation between the myocardial expression of the inducible 70-kDa heat shock protein (HSP70i) by different stress conditions and its possible protective effects. Only few studies have focused on the involvement of small heat shock proteins in this process. We analyzed the location of the small heat shock protein HSP25 in isolated cardiomyocytes as well as its location and induction in isolated perfused hearts of rats. By immunofluorescence microscopy HSP25 was found to colocalize with actin in the I-band of myofibrils in cardiomyocytes of isolated perfused hearts as well as in isolated neonatal and adult cardiomyocytes. Hyperthermic perfusion of isolated hearts for 45 min resulted in modulation of different parameters of heart function and in induction of HSP25 and HSP70i. Temperatures higher than 43°C (44–46°C) were lethal with respect to the contractile function of the hearts. Compared to control hearts perfused at 37°C, significant increases during hyperthermic perfusion at 42°C and 43°C were obtained for heart rate, contraction velocity and relaxation velocity. In response to hyperthermia at 43°C and after subsequent normothermic perfusion for 135 min at 37°C, left ventricular pressure, contraction velocity and relaxation velocity remained significantly elevated. However, heart rate returned to control values immediately after the period of heat treatment. HSP25 is constitutively expressed even in normothermic perfused hearts as shown by Western blotting. Hyperthermia increased the content of HSP25 only in the left ventricular tissue. In contrast, HSP70i was strongly induced in all analyzed parts of the myocardium (left ventricle, right ventricle, septum). Our findings suggest a differential regulation of HSP25 and HSP70i expression in response to hyperthermia in isolated perfused hearts. The constitutively expressed HSP25 seems to be located adjacent to the myofibrils which implies a specific role of this protein even under unstressed conditions for the contractile function of the myocardium.     

Regulation and functional significance of phospholipase D in myocardium

There is now clear evidence that receptor-dependent phospholipase D is present in myocardium. This novel signal transduction pathway provides an alternative source of 1,2-diacylglycerol, which activates isoforms of protein kinase C. The members of the protein kinase C family respond differently to various combinations of Ca²⁺, phosphatidylserine, molecular species of 1,2-diacylglycerol and other membrane phospholipid metabolites including free fatty acids. Protein kinase C isozymes are responsible for phosphorylation of specific cardiac substrate proteins that may be involved in regulation of cardiac contractility, hypertrophic growth, gene expression, ischemic preconditioning and electrophysiological changes. The initial product of phospholipase D, phosphatidic acid, may also have a second messenger role. As in other tissues, the question how the activity of phospholipase D is controlled by agonists in myocardium is controversial. Agonists, such as endothelin-1, atrial natriuretic factor and angiotensin 11 that are shown to activate phospholipase D, also potently stimulate phospholipase C- in myocardium. PMA stimulation of protein kinase C inactivates phospholipase C and strongly activates phospholipase D and this is probably a major mechanism by which agonists that promote phosphatidyl-4,5-bisphosphate hydrolysis secondary activate phosphatidylcholine-hydrolysis. On the other hand, one group has postulated that formation of phosphatidic acid secondary activates phosphatidyl-4,5-bisphosphate hydrolysis in cardiomyocytes. Whether GTP-binding proteins directly control phospholipase D is not clearly established in myocardium. Phospholipase D activation may also be mediated by an increase in cytosolic free Ca²⁺ or by tyrosine-phosphorylation.     

Early degradation of photosynthetic membranes in carob and sunflower cotyledons

The assimilatory activity of cotyledons can play an essential role in the survival of seedlings with a slow and delayed development of primary leaves. Changes in the photosynthetic activity of the cotyledon, from the onset of greening through senescence, were studied in two such plants, carob and sunflower, in order to determine its efficiency and duration, also in connection with the achievement of assimilatory autonomy by the plantlet. Chlorophyll analyses showed that the cotyledon's chloroplasts reached maximal greening in plantlets with a pair of expanded leaves. In contrast, the cotyledon's photosynthetic activity, measured as the rate of oxygen release, started to decrease early, before expansion of primary leaves. The decrease was due to the inactivation of a number of photosystem II (PSII) units, as revealed by immunodetection of breackdown products of the reaction centre's D1 and D2 thylakoid proteins. No signals of PSII alteration were noticed in the primary leaf chloroplasts that differentiated under the same environmental conditions. The damage to the cotyledon PSII, occurring in a non-photoinhibitory situation, might be due to a slower rate of turnover of D1 polypeptide than in the leaf thylakoids. The differential turnover of this protein in cotyledons and in leaves might represent an organ-specific regulation of the photosynthetic activity. The peculiarity of the cotyledon thylakoids make these organs useful objects for studying the metabolic cycle of both D1 and D2 proteins in vivo, under non-photoinhibiting conditions.     

Conservation of the alternative oxidase and enhancement of cyanide-resistant respiration in suspension-cultured pear fruit cells by inhibitors of macromolecular synthesis

The contribution of the alternative pathway to the respiration of suspension-cultured pear ( Pyrus communis cv. Passa Crasanne) cells was enhanced, often severalfold, within 2 to 4 days following the addition of cycloheximide, actinomycin D, or 2-(4-methyl-2,6-dinitroanalino)- N -methyl propionamide (D-MDMP). Concomitant inhibition of cellular protein synthesis by cycloheximide and actinomycin D was transient and incomplete. However, inhibition by D-MDMP was virtually complete (>97%) and persisted over several days. [³⁵S]-labelling and polyacrylamide gel separation indicated that cycloheximide precluded the appearance of discernable new proteins in mitochondria. Probes with monoclonal antibodies revealed a conservation of alternative oxidase protein levels in the mitochondria of inhibitor-treated cells. The data, appraised within the complexities of cell-culture dynamics, lead to the conclusion that the observed increases in capacity for cyanide-resistant respiration are the consequence, likely indirect, of inhibited protein synthesis with resultant retention and activation of constitutive alternative oxidase.     

Wave separation versus bandpass filtering: a comparative non-linear analysis of brain α-EEG signals with and without psychotropic drug treatment

Attempts to demonstrate low-dimensional attractor behaviour in the analysis of electroencephalographic (EEG) signals meet with difficulties that in part stem from a departure from single-system dynamics. In order to address this problem, the -waves can be extracted by digital filtering or by wave separation; these two techniques are compared in order to specify the conditions in which finite impulse response (FIR) bandpass filters can be used. The comparison was made using 18 EEG records of 3 min duration under resting conditions (6 subjects, 3 records per subject: prior to apomorphine administration, then 90 min and 150 min post-treatment). No presence of low-dimensional dynamic episodes in -signals was observed without digital processing. Sixty 5 s sections showing attractor behaviour were found after filtering and twenty five 5 s sections after wave separation. The mean correlation dimension was calculated for each experimental condition and for 4 subjects, in order to observe the temporal profile of the drug. When attractors were found after wave separation, bandpass filtering then also showed attractor behaviour, with the same temporal profile. However, the reverse is not true: attractors were found after bandpass filtering that were not present after wave separation; in this case the results deserve confirmation, although the temporal profiles for all cases in which attractors were found after filtering remained comparable.