Regulation of malate dehydrogenase activity by glutamate, citrate, alpha-ketoglutarate, and multienzyme interaction

Binding experiments indicate that mitochondrial aspartate aminotransferase can associate with the alpha-ketoglutarate dehydrogenase complex and that mitochondrial malate dehydrogenase can associate with this binary complex to form a ternary complex. Formation of this ternary complex enables low levels of the alpha-ketoglutarate dehydrogenase complex, in the presence of the aminotransferase, to reverse inhibition of malate oxidation by glutamate. Thus, glutamate can react with the aminotransferase in this complex without glutamate inhibiting production of oxalacetate by the malate dehydrogenase in the complex. The conversion of glutamate to alpha-ketoglutarate could also be facilitated because in the trienzyme complex, oxalacetate might be directly transferred from malate dehydrogenase to the aminotransferase. In addition, association of malate dehydrogenase with these other two enzymes enhances malate dehydrogenase activity due to a marked decrease in the Km of malate. The potential ability of the aminotransferase to transfer directly alpha-ketoglutarate to the alpha-ketoglutarate dehydrogenase complex in this multienzyme system plus the ability of succinyl-CoA, a product of this transfer, to inhibit citrate synthase could play a role in preventing alpha-ketoglutarate and citrate from accumulating in high levels. This would maintain the catalytic activity of the multienzyme system because alpha-ketoglutarate and citrate allosterically inhibit malate dehydrogenase and dissociate this enzyme from the multienzyme system. In addition, citrate also competitively inhibits fumarase. Consequently, when the levels of alpha-ketoglutarate and citrate are high and the multienzyme system is not required to convert glutamate to alpha-ketoglutarate, it is inactive. However, control by citrate would be expected to be absent in rapidly dividing tumors which characteristically have low mitochondrial levels of citrate.     

Restoration of malonyl-CoA sensitivity of soluble rat liver mitochondria carnitine palmitoyltransferase by reconstitution with a partially purified malonyl-CoA binding protein.

Solubilization of rat liver mitochondria in 5% Triton X-100 followed by chromatography on a hydroxylapatite column resulted in the identification of malonyl-CoA binding protein(s) distinct from a major carnitine palmitoyltransferase activity peak. Further purification of the malonyl-CoA binding protein(s) on an acyl-CoA affinity column followed by sodium dodecyl sulfate gel electrophoresis indicated proteins with Mr mass of 90 and 45-33 kDa. A purified liver malonyl-CoA binding fraction, which was devoid of carnitine palmitoyltransferase, and a soluble malonyl-CoA-insensitive carnitine palmitoyltransferase were reconstituted by dialysis in a liposome system. The enzyme activity in the reconstituted system was decreased by 50% in the presence of 100 microM malonyl-CoA. Rat liver mitochondria carnitine palmitoyltransferase may be composed of an easily dissociable catalytic unit and a malonyl-CoA sensitivity conferring regulatory component.     

Determination of a unique solution to parallel proton transfer reactions using the genetic algorithm

Kinetic analysis of the dynamics as measured in multiequilibria systems is readily attained by curve-fitting methodologies, a treatment that can accurately retrace the shape of the measured signal. Still, these reconstructions are not related to the detailed mechanism of the process. In this study we subjected multiple proton transfer reactions to rigorous kinetic analysis, which consists of solving a set of coupled-nonlinear differential rate equations. The manual analysis of such systems can be biased by the operator; thus the analysis calls for impartial corroboration. What is more, there is no assurance that such a complex system has a unique solution. In this study, we used the Genetic Algorithm to investigate whether the solution of the system will converge into a single global minimum in the multidimensional parameter space. The experimental system consisted of proton transfer between four proton-binding sites with seven independent adjustable parameters. The results of the search indicate that the solution is unique and all adjustable parameters converge into a single minimum in the multidimensional parameter space, thus corroborating the accuracy of the manual analysis.     

Determinants of cyanobacterial species composition in the splash zone of two Croatian islands

Cyanobacterial communities of the splash zone of two Croatian islands, Veruda and Ugljan, were surveyed. At each island, we studied eight localities with different aspects. From all 336 samples, a total of 42 cyanobacterial species, one green alga, and one red alga were found. In both islands Hyella spp. and Gloeocapsopsis crepidinum dominated and Entophysalis deusta, Solentia spp., Kyrtuthrix dalmatica and Mastigocoleus testarum were found frequently. Multivariate statistical analysis of species composition shows that the vertical gradient is the strongest determinant of species composition and that there are statistically significant but not very pronounced differences in species composition between the two islands. Species composition among individual sites within islands differed slightly, and between localities decreased rather slowly with the distance of individual samples (i.e. the similarity distance decay was very slow), showing that even close samples can have quite different species composition. The species accumulation curves do not reach their asymptote, indicating that many samples are needed to reasonably cover the species richness of a site. This study highlights the importance of in-depth species counts and high density sampling along vertical gradients for satisfactory cyanobacterial community assessments in marine coastal splash zones.     

Disentangling the Diversity of Arboreal Ant Communities in Tropical Forest Trees

Tropical canopies are known for their high abundance and diversity of ants. However, the factors which enable coexistence of so many species in trees, and in particular, the role of foragers in determining local diversity, are not well understood. We censused nesting and foraging arboreal ant communities in two 0.32 ha plots of primary and secondary lowland rainforest in New Guinea and explored their species diversity and composition. Null models were used to test if the records of species foraging (but not nesting) in a tree were dependent on the spatial distribution of nests in surrounding trees. In total, 102 ant species from 389 trees occurred in the primary plot compared with only 50 species from 295 trees in the secondary forest plot. However, there was only a small difference in mean ant richness per tree between primary and secondary forest (3.8 and 3.3 sp. respectively) and considerably lower richness per tree was found only when nests were considered (1.5 sp. in both forests). About half of foraging individuals collected in a tree belonged to species which were not nesting in that tree. Null models showed that the ants foraging but not nesting in a tree are more likely to nest in nearby trees than would be expected at random. The effects of both forest stage and tree size traits were similar regardless of whether only foragers, only nests, or both datasets combined were considered. However, relative abundance distributions of species differed between foraging and nesting communities. The primary forest plot was dominated by native ant species, whereas invasive species were common in secondary forest. This study demonstrates the high contribution of foragers to arboreal ant diversity, indicating an important role of connectivity between trees, and also highlights the importance of primary vegetation for the conservation of native ant communities.     

Measuring size and composition of species pools: a comparison of dark diversity estimates

Ecological theory and biodiversity conservation have traditionally relied on the number of species recorded at a site, but it is agreed that site richness represents only a portion of the species that can inhabit particular ecological conditions, that is, the habitat‐specific species pool. Knowledge of the species pool at different sites enables meaningful comparisons of biodiversity and provides insights into processes of biodiversity formation. Empirical studies, however, are limited due to conceptual and methodological difficulties in determining both the size and composition of the absent part of species pools, the so‐called dark diversity. We used >50,000 vegetation plots from 18 types of habitats throughout the Czech Republic, most of which served as a training dataset and 1083 as a subset of test sites. These data were used to compare predicted results from three quantitative methods with those of previously published expert estimates based on species habitat preferences: (1) species co‐occurrence based on Beals' smoothing approach; (2) species ecological requirements, with envelopes around community mean Ellenberg values; and (3) species distribution models, using species environmental niches modeled by Biomod software. Dark diversity estimates were compared at both plot and habitat levels, and each method was applied in different configurations. While there were some differences in the results obtained by different methods, particularly at the plot level, there was a clear convergence, especially at the habitat level. The better convergence at the habitat level reflects less variation in local environmental conditions, whereas variation at the plot level is an effect of each particular method. The co‐occurrence agreed closest the expert estimate, followed by the method based on species ecological requirements. We conclude that several analytical methods can estimate species pools of given habitats. However, the strengths and weaknesses of different methods need attention, especially when dark diversity is estimated at the plot level.     

Modelling the Population Dynamics of Root Hemiparasitic Plants Along a Productivity Gradient

Root hemiparasitic plants interact with their host plants through parasitism and competition. The interactions can be divided into aboveground and belowground interactions. Because both groups of plants are autotrophic, they compete for light aboveground. Belowground interactions are more complex. The host plants compete for resources in the soil and the hemiparasitic plants prey on the host plants through haustoria, using the hosts as the main source of water and nutrients. In this paper, we modeled the relationship between these two plant types, extending the well-known Rosenzweig-MacArthur predator-prey model to cover both light competition and intra-specific parasitism among hemiparasites. We included a realistic relationship of carrying capacity to environmental productivity and followed model behavior on a productivity gradient. The model shows that, at very low productivities, there are only a few poor hosts and hemiparasites have no chance to persist. As productivity increases, there is a range of productivity where both plant types coexist. A further increase in productivity gets the system out of the coexistence range, and only host plants survive. This final prediction successfully explains patterns observed in empirical data, contrary to the results of an earlier, oversimplified model of the explored interaction. Comparison of various models demonstrates that the model is able to reproduce the decline of hemiparasites with increasing productivity only when competition for light is included.     

Seed dispersal and realized gene flow of two forest orchids in a fragmented landscape

• Species with vast production of dust‐like windborne seeds, such as orchids, should not be limited by seed dispersal. This paradigm, however, does not fit recent studies showing that many sites suitable for orchids are unoccupied and most seeds land close to their maternal plant. To explore this issue, we studied seed dispersal and gene flow of two forest orchid species, Epipactis atrorubens and Cephalanthera rubra, growing in a fragmented landscape of forested limestone hills in southwest Bohemia, Czech Republic.
• We used a combination of seed trapping and plant genotyping methods (microsatellite DNA markers) to quantify short‐ and long‐distance dispersal, respectively. In addition, seed production of both species was estimated.
• We found that most seeds landed very close to maternal plants (95% of captured seeds were within 7.2 m) in both species, and dispersal distance was influenced by forest type in E. atrorubens. In addition, C. rubra showed clonal reproduction (20% of plants were of clonal origin) and very low fruiting success (only 1.6% of plants were fruiting) in comparison with E. atrorubens (25.7%). Gene flow was frequent up to 2 km in C. rubra and up to 125 km in E. atrorubens, and we detected a relatively high dispersal rate among regions in both species.
• Although both species occupy similar habitats and have similar seed dispersal abilities, C. rubra is notably rarer in the study area. Considerably low fruiting success in this species likely limits its gene flow to longer distances and designates it more sensitive to habitat loss and fragmentation.

     

Kinetics of luminal proton binding to the SR Ca-ATPase

An open membrane preparation containing SR Ca-ATPase was prepared from sarcoplasmic-reticulum vesicles to study the ion binding kinetics in the P-E₂ conformation. Because Ca²⁺ and H⁺ binding are electrogenic reactions, fluorescent styryl dyes could be used to determine changes in the binding site occupation in equilibrium titration experiments and time-resolved relaxation processes triggered by a pH jump. By photo release from caged proton the pH of the electrolyte could be decreased in a step of 0.1 pH units by a single ultraviolet-laser flash. Analysis of the pH-jump induced relaxation process in the P-E₂ conformation showed that three Ca-ATPase-specific processes could be identified, fast H⁺ binding (τ < 100 μs) and pH-insensitive conformational relaxations after the release of the Ca²⁺ ion (τ ∼160 ms), and a slow process (τ ∼3.4 s) whose origin could not be unambiguously revealed. The Ca²⁺-binding affinity in the P-E₂ conformation was reduced with increasing pH, a behavior that can be explained by a reversible transition of the empty P-E₂ state to an inactivated state of the ion pump. All findings are interpreted in the framework of the Post-Albers pump cycle introduced previously, supplemented by an additional transition to an inhibited state of the ion pump.     

Plant density affects measures of biodiversity effects

Aim s: We tested for the effect of final sowing plant density (i.e. density of established seedlings) on the values of biodiversity effects [transgressive overyielding, net effect, complementarity effect (CE) and selection effect (SE), trait-dependent complementarity and dominance effect] in a glasshouse pot experiment.Methods: We conducted a single-season (4 months) glasshouse experiment. Species monocultures and mixtures containing up to four common meadow species from different functional groups were sown and subsequently thinned to five density levels (8–128 individuals per pot, i.e. 200–3200 individuals m ?2). Community functioning was characterized by yield (both living and dead biomass) of all constituent species.Important Findings: Our results show that plant density (final sowing density in our case, but this finding can be generalized) affects the yields of both monocultures and mixtures. As these and their relationships are the basis for calculation of biodiversity effects, these effects also varied along the density gradient. Net biodiversity effect, CE and SE all increased with density. The net biodiversity effect and the CE switched from negative to quite positive in the four-species mixture. Using Fox's tripartite partitioning, trait-dependent complementarity was minor in comparison to the dominance effect. One of our experimental species did not follow the density-productivity relationship, called constant final yield (CFY), which was reflected in the biodiversity measures. The shape of the density-productivity relationship for experimental species affects also the values of biodiversity indices, particularly when species do not follow the CFY relationship. According to our data and recent simulation experiments, the values of commonly used biodiversity effects can be rather misleading if a species has, e.g. a unimodal dependence of yield for the density gradient and the density level used in the experiment is higher than the peak density.