Effects of nucleotides and divalent cations on phospholipase activity in Saccharomyces cerevisiae

Divalent cations activate the lysophospholipase and transacylase reactions catalyzed by the same enzymes in the yeast Saccharomyces cerevisiae. The activation was observed at neutral pH, but not at the pH optimum of lysophospholipase/transacylase, near 3.5. Adenine nucleotides, especially AMP and ADP, are strong inhibitors of the same group of enzymes. Half maximal inhibition by AMP was found at a concentration of about 20 M. The inhibition by nucleotides in low concentrations is enhanced by divalent cations.     

Effect of temperature on the pathways of NADH-oxidation in broad-bean mitochondria

1. Respiration rates of broad-bean (Vicia faba) mitochondria were studied as a function of temperature. Arrhenius plots of all membrane-bound enzymes, as obtained with saturating substrate concentrations, revealed a break in the lower temperature range. That break was considered to indicate a phase transition of membrane phospholipids, characteristic for chilling-sensitive plants. A second discontinuity at 30°C occurred only with activities linked to energy conservation. — 2. The activation energies for the oxidation of NAD⁺-linked substrates differ between states 3 and 4. State 3 respiration of NAD⁺-linked substrates is the result a superimposition of two branches of electron transport, which can be separated by different sensibilities to rotenone. A characteristic temperature dependency of the respiratory control, as well as a shift of the low temperature break in the Arrhenius plot toward a higher temperature after state 4 to state 3 transition, are calculated to be caused by the superimposition of the two branches. — 3. The temperature dependency of the oxidation of extra-mitochondrial NADH and of succinate differs remarkably from that of the oxidation of matrix-NADH. It has been concluded that the rotenone-resistant oxidation of matrix-NADH and the oxidation of external NADH are mediated via different pathways with individual regulation sites.Abbreviations BSA bovine serum albumin - CCCP carbonylcyanide-m-chlorophenylhydrazone - TPP thiaminepyrophosphate     

cAMP-dependent protein kinase regulates Polysphondylium pallidum development

In eukaryotic cells, the universal second messenger cAMP regulates various aspects of development and differentiation. The primary target for cAMP is the regulatory subunit of cAMP-dependent protein kinase A (PKA), which, upon cAMP binding, dissociates from the catalytic subunit and thus activates it. In the soil amoeba Dictyostelium discoideum, the function of PKA in growth, development and cell differentiation has been thoroughly investigated and substantial information is available. To obtain a more general view, we investigated the influence of PKA on development of the related species Polysphondylium pallidum. Cells were transformed to overexpress either a dominant negative mutant of the regulatory subunit (Rm) from Dictyostelium that cannot bind cAMP, or the catalytic subunit (PKA-C) from Dictyostelium. Cells overexpressing Rm rarely aggregated and the few multicellular structures developed slowly into very small fruiting bodies without branching of secondary sorogens, the prominent feature of Polysphondylium. Few round spores with reduced viability were formed. When mixed with wild-type cells and allowed to develop, the Rm cells were randomly distributed in aggregation streams, but were later found in the posterior region of the culminating slug or were left behind on the surface of the substratum. The PKA-C overexpressing cells exhibited precocious development and formed more aggregates of smaller size. Moreover, expression of PKA-C under the control of the prestalk-specific ecmB promoter of Dictyostelium leads to protrusions from aggregation streams. We conclude that Dictyostelium PKA subunits introduced into Polysphondylium cells are functional as signal components, indicating that a biochemically similar PKA mechanism works in Polysphondylium.     

The influence of externally supplied sucrose on phloem transport in the maize leaf strip

Sucrose (2,5–1000 mmol l^–1), labeled with [¹⁴C]sucrose, was taken up by the xylem when supplied to one end of a 30-cm-long leaf strip of Zea mays L. cv. Prior. The sugar was loaded into the phloem and transported to the opposite end, which was immersed in diluted Hoagland's nutrient solution. When the Hoagland's solution at the opposite end was replaced by unlabeled sucrose solution of the same molarity as the labeled one, the two solutions met near the middle of the leaf strip, as indicated by radioautographs. In the dark, translocation of ¹⁴C-labeled assimilates was always directed away from the site of sucrose application, its distance depending on sugar concentration and translocation time. When sucrose was applied to both ends of the leaf strip, translocation of ¹⁴C-labeled assimilates was directed toward the lower sugar concentration. In the light, transport of ¹⁴-C-labeled assimilates can be directed (1) toward the morphological base of the leaf strip only (light effect), (2) toward the base and away from the site of sucrose application (light and sucrose effect), or (3) away from the site of sucrose application independent of the (basipetal or acropetal) direction (sucrose effect). The strength of a sink, represented by the darkened half of a leaf strip, can be reduced by applying sucrose (at least 25 mmol l^–1) to the darkened end of the leaf strip. However, equimolar sucrose solutions applied to both ends do not affect the strength of the dark sink. Only above 75 mmol l^–1 sucrose was the sink effect of the darnened part of the leaf strip reduced. Presumably, increasing the sucrose concentration replenishes the leaf tissue more rapidly, and photosynthates from the illuminated part of the leaf strip are imported to a lesser extent by the dark sink.Supported by Deutsche Forschungsgemeinschaft     

Pressurized gas transport in two Japanese alder species in relation to their natural habitats

Oxygen uptake measurements have shown that pressurized gas transport, resulting from the physical effect of thermo-osmosis of gases, improves oxygen supply to the roots of the seedlings in two alder speciesAlnus japonica (Thunb.) Steud. andAlnus hirsuta (Spach) Rupr., which are both native in Japan. When gas transport conditions were established by irradiation of the tree stems the internal aeration was increased to a level nearly equal to the oxygen demand of the root system in leafless seedlings ofA. hirsuta, but was higher inA. japonica so that excess oxygen was excreted into the environment. An increase of superoxide dismutase (SOD) activity, which protects plants from toxic oxygen radicals and post-anoxic injury, has been observed in root tissues ofA. japonica when the seedlings were flooded for 3 days. The increase of SOD activity, in concert with high gas transport rates, may enable this tree species to grow in wet sites characterized by low oxygen partial pressure in the soil and by varying water tables. A less effective gas transport, flood-induced reduction of SOD activity in root tissues, and reduced height growth in waterlogged soil may be responsible for the fact thatA. hirsuta is unable to inhabit wettland sites.     

Asymmetric distribution of mitochondria and of spindle microtubules in opposite directions in differential mitosis of germ line cells in Acricotopus

Additional chromosomes present only in the germ line are a specific feature of the Orthocladiinae, a subfamily of the Chironomidae. During the complex chromosome cycle in the orthocladiid Acricotopus lucidus, about half of the germ-line-limited chromosomes (Ks) are eliminated in the first division of the primary germ cells. Following normal gonial mitoses, the reduction in the number of Ks is compensated for, in the last mitosis prior to meiosis, by a monopolar movement of the unseparated Ks, while the somatic chromosomes (Ss) segregate equally. This differential mitosis produces daughter cells with different chromosome constitutions and diverse developmental fates. A preferential segregation of mitochondria occurs to one pole associated with an asymmetric formation of the mitotic spindle. This has been detected in living gonial cells in both sexes by using MitoTracker probes and fluorochrome-labelled paclitaxel (taxol). In males, the resulting unequal partitioning of mitochondria to the daughter cells is equalised by the transport of mitochondria through a permanent cytoplasmic bridge from the aberrant spermatocyte to the primary spermatocyte. This asymmetry in the distribution and in the segregation of cytoplasmic components in differential gonial mitosis in Acricotopus may be involved in the process of cell-fate determination. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorised users.     

Quantitative structure analysis of genetic diversity among spring bread wheats (<Emphasis Type="Italic">Triticum aestivum</Emphasis> L.) from different geographical regions

Genetic diversity in spring bread wheat (T.␣aestivum L.) was studied in a total of 69 accessions. For this purpose, 52 microsatellite (SSR) markers were used and a total of 406 alleles were detected, of which 182 (44.8%) occurred at a frequency of <5% (rare alleles). The number of alleles per locus ranged from 2 to 14 with an average of 7.81. The largest number of alleles per locus occurred in the B genome (8.65) as␣compared to the A (8.43) and D (5.93) genomes, respectively. The polymorphism index content (PIC) value varied from 0.24 to 0.89 with an average of 0.68. The highest PIC for all accessions was found in the B␣genome (0.71) as compared to the A (0.68) and D␣genomes (0.63). Genetic distance-based method (standard UPGMA clustering) and a model-based method (structure analysis) were used for cluster analysis. The two methods led to analogical results. Analysis of molecular variance (AMOVA) showed that 80.6% of the total variation could be explained by the variance within the geographical groups. In comparison to the diversity detected for all accessions (H _e = 0.68), genetic diversity among European spring bread wheats was H _e = 0.65. A comparatively higher diversity was observed between wheat varieties from Southern European countries (Austria/Switzerland, Portugal/Spain) corresponding to those from other regions.     

Penicillium chrysogenum glucose oxidase -- a study on its antifungal effects

AIMS: Purification and characterization of the high molecular mass Candida albicans-killing protein secreted by Penicillium chrysogenum. METHODS AND RESULTS: The protein was purified by a combination of ultrafiltration, chromatofocusing and gel filtration. Enzymological characteristics [relative molecular mass (M(r)) = 155 000, subunit structure alpha(2) with M(r,alpha) = 76 000, isoelectric point (pI) = 5.4] were determined using SDS-PAGE and 2D-electrophoresis. N-terminal amino acid sequencing and homology search demonstrated that the antifungal protein was the glucose oxidase (GOX) of the fungus. The enzyme was cytotoxic for a series of bacteria, yeasts and filamentous fungi. Vitamin C (1.0 mg ml(-1)) prevented oxidative cell injuries triggered by 0.004 U GOX in Emericella nidulans cultures but bovine liver catalase was ineffective even at a GOX : catalase activity ratio of 0.004 : 200 U. A secondary inhibition of growth in E. nidulans cultures by the oxygen-depleting GOX-catalase system was likely to replace the primary inhibition exerted by H(2)O(2). CONCLUSIONS: Penicillium chrysogenum GOX possesses similar enzymological features to those described earlier for other Penicillium GOXs. Its cytotoxicity was dependent on the inherent antioxidant potential of the test micro-organisms. SIGNIFICANCE AND IMPACT OF THE STUDY: Penicillium chrysogenum GOX may find future applications in glucose biosensor production, the disinfection of medical implants or in the food industry as an antimicrobial and/or preservative agent.     

Seed weight increases with altitude in the Swiss Alps between related species but not among populations of individual species

Seed weight is a crucial plant life history trait, determining establishment success and dispersal ability. Especially in stressful environments, larger seeds may be selected at the expense of seed number, because larger seeds have a better chance of giving rise to an established offspring. We tested the hypotheses that between related species-pairs and among populations of single species a similar trend for increasing seed weight with increasing altitude should be present. Firstly, we measured seed weights from 29 species-pairs, with one species occurring in lowland areas and a congeneric species from high altitudes. Seeds of the alpine species were 28±8% larger than seeds from lowland species (P<0.01). Compared to the related lowland species, 55% of the alpine species had heavier seeds, 3% (one species) had lighter, and 41% had seeds of approximately equal weight. Secondly, we compared seed weights among populations of four species from different habitats and with different life histories. Seeds from between 11 and 34 populations per species were sampled along altitudinal gradients of 800–1,500 m (ca. 800 m in Scabiosa lucida, ca. 1,000 m in Saxifraga oppositifolia, ca. 1,000 m in Epilobium fleischeri, and ca. 1,500 m in Carex flacca). In all the four species, we found no indication for heavier seeds at higher altitudes. Our results indicate a selection pressure for species with heavier seeds at higher altitude, but the trend does not seem to operate across all cases. Phylogenetic constraints may limit the correlation among altitude and seed weight, operating particularly against selection for larger seed size, the closer populations and species are related to each other.