Cloning of a Picea abies monosaccharide transporter gene and expression – analysis in plant tissues and ectomycorrhizas

Ectomycorrhizas are formed between certain soil fungi and fine roots of woody plants. An important feature of this symbiosis is the supply of photoassimilates to the fungus. Hexoses, formed from sucrose in the common apoplast at the root/fungus interface, can be taken up by both plant and fungal monosaccharide transporters. Recently we characterised a monosaccharide transporter from the ectomycorrhizal fungus Amanita muscaria. This transporter was up-regulated in mycorrhizas, thus increasing the hexose uptake capacity of the fungal partner in symbiosis. In order to characterise host (Picea abies) root monosaccharide transporters, degenerate oligonucleotide primers, designed to match conserved regions from known plant hexose transporters, were used to isolate a cDNA fragment of a transporter by PCR. This fragment was used to identify a presumably full length clone (PaMST1) in a P. abies/A. muscaria mycorrhizal cDNA library. The entire cDNA code for an open reading frame of 513 amino acids, revealing best homology to H⁺/monosaccharide transporters from Ara- bidopsis, Saccharum and Ricinus. PaMST1 was highly expressed in the hypocotyl and in roots of P. abies seedlings, but not in needles. Mycorrhiza formation led to a slight reduction of PaMST1 expression. The results are discussed with special reference to carbon allocation in ectomycorrhizas. Received: 9 October 1999 / Accepted: 22 December 1999     

Resibufogenin corrects hypertension in a rat model of human preeclampsia

The study of the pathogenesis of preeclampsia has been hampered by a relative dearth of animal models. We developed a rat model of preeclampsia in which the excretion of a circulating inhibitor of Na/K ATPase, marinobufagenin (MBG), is elevated. These animals develop hypertension, proteinuria, and intrauterine growth restriction. The administration of a congener of MBG, resibufogenin (RBG), reduces blood pressure to normal in these animals, as is the case when given to pregnant animals rendered hypertensive by the administration of MBG. Studies of Na/K ATPase inhibition by MBG and RBG reveal that these agents are equally effective as inhibitors of the enzyme.     

Biological suppression of potato ring rot by fluorescent pseudomonads.

Three strains of fluorescent pseudomonads (IS-1, IS-2, and IS-3) isolated from potato underground stems with roots showed in vitro antibiosis against 30 strains of the ring rot bacterium Clavibacter michiganensis subsp. sepedonicus. On the basis of morphological and biochemical tests and fatty acid analysis, IS-1 and IS-2 were identified as Pseudomonas aureofaciens and IS-3 was identified as P. fluorescens biovar III. IS-1 was the most inhibitory to C. michiganensis subsp. sepedonicus strains in vitro, followed by IS-3 and IS-2. Suppression of ring rot by these antagonists was demonstrated in greenhouse trials with stem-cultured potato (cv. Russet Burbank) seedlings. Although each antagonist significantly reduced C. michiganensis subsp. sepedonicus populations, only IS-1 reduced infection by C. michiganensis subsp. sepedonicus. In a second experiment, treatment with IS-1 (10(9) CFU/ml) significantly reduced ring rot infection by 23.4 to 26.7% after 5 to 8 weeks. The average C. michiganensis subsp. sepedonicus population was also significantly reduced by 50 to 52%. Application of different combinations of antagonist strains was not more effective than single-strain treatment.     

Comparative Metabolism of Fluorinated 3,4-Dihydroxyphenylalanine Isomers in Aggregating Brain Cell Cultures

Fluorinated analogues of 3,4-dihydroxyphenylalanine (DOPA) were tested for intracellular metabolic conversion in aggregating cell cultures prepared from fetal rat brain. 5-Fluoro-D/L-DOPA was methylated almost exclusively to 3-O-methyl-5-fluoro-D/L-DOPA. Metabolism of 6-fluoro-D/L-DOPA resulted in 6-fluorodopamine, 6-fluoro-3,4-dihydroxyphenylacetic acid, and 3-O-methyl-6-fluoro-D/L-DOPA, but with a qualitatively and quantitatively different metabolite pattern compared with that of L-DOPA and D/L-DOPA, respectively. Homovanillic acid and fluorohomovanillic acid have not been found intracellularly in the cultures. On the basis of these data, the model development of the cerebral metabolism of tracers used in positron emission tomography can be improved.     

Gut Microbiota in Children Hospitalized with Oedematous and Non-Oedematous Severe Acute Malnutrition in Uganda

Background

Severe acute malnutrition (SAM) among children remains a major health problem in many developing countries. SAM manifests in both an oedematous and a non-oedematous form, with oedematous malnutrition in its most severe form also known as kwashiorkor. The pathogenesis of both types of malnutrition in children remains largely unknown, but gut microbiota (GM) dysbiosis has recently been linked to oedematous malnutrition. In the present study we aimed to assess whether GM composition differed between Ugandan children suffering from either oedematous or non-oedematous malnutrition.

Methodology/Principal Findings

As part of an observational study among children hospitalized with SAM aged 6–24 months in Uganda, fecal samples were collected at admission. Total genomic DNA was extracted from fecal samples, and PCR amplification was performed followed by Denaturing Gradient Gel Electrophoresis (DGGE) and tag-encoded 16S rRNA gene-targeted high throughput amplicon sequencing. Alpha and beta diversity measures were determined along with ANOVA mean relative abundance and G-test of independence followed by comparisons between groups. Of the 87 SAM children included, 62% suffered from oedematous malnutrition, 66% were boys and the mean age was 16.1 months. GM composition was found to differ between the two groups of children as determined by DGGE (p = 0.0317) and by high-throughput sequencing, with non-oedematous children having lower GM alpha diversity (p = 0.036). However, beta diversity analysis did not reveal larger differences between the GM of children with oedematous and non-oedematous SAM (ANOSIM analysis, weighted UniFrac, R = -0.0085, p = 0.584; unweighted UniFrac, R = 0.0719, p = 0.011).

Conclusions/Significance

Our results indicate that non-oedematous SAM children have lower GM diversity compared to oedematous SAM children, however no clear compositional differences were identified.     

ON SEXUAL AGGLUTINATION AND MATING-TYPE SUBSTANCES IN ISOGAMOUS DIOECIOUS CHLAMYDOMONADS. III. THE SENSITIVITY OF SEX CELL CONTACT TO VARIOUS ENZYMES

The components responsible for sex cell contact in dioecious Chlamydomonas species have been characterized by their sensitivity to enzymes. The mating-type substances, which are glycoprotein complexes according to the analysis of the corresponding isoagglutinins, were incubated with proteases, glycosidases, and with sulfatase. An enzyme may or may not affect both gamete types of a species or it may discriminate between the sexes, eliminating one mating-type activity only. In no case was an incapacitation of a given gamete type based upon the detachment of the proper contact component as an isoagglutinin or as a functionally univalent structure with a specific receptor-blocking power against the complementary type. Inactivating enzymes seem to interfere with the surface configuration essential for gamete contact or to eliminate actual contact sites. The compilation of data for six taxa reveals a complex sensitivity pattern of the gametes’ agglutinability. Two types of interactions appear to occur. In C. reinhardti and C. chlamydogama the contact mechanism seems to depend on an interaction between two proteinaceous structures. In C. eugametos and C. moewusii definite proof was obtained for a contact based upon a carbohydrate-protein interaction. In the (+) sexes of C. eugametos and C. moewusii syngen II, α-glycosidically bound terminal mannose residues are essential for the contact. Neither β-galactosidase, neuraminidase, or sulfatase incapacitate the mating-type substances in situ.     

On the evolution of anisogamy from isogamous monoecy and on the origin of sex

A new hypothesis is established for the evolution of anisogamy in isogamous monoecious haploid unicells. Contrary to present concepts, it is assumed that in such isogamous ancestors the genetically identical gametes are bipolarly different as (+) and (?). The gametic differentiation of a monoecious taxon may be perceived as a phenocopy of that of a related dioecious form. The two gametic phenotypes result from alternative pathways of sexual differentiation. The alternative must be controlled by a sensible switch mechanism that responds to minor differences between vegetative cells when gametogenesis is triggered. Cell-size dependent factors are assumed to be able to influence the switch mechanism causing bigger cells to be of one sex, smaller cells of the other. Fertilization then occurs selectively between big and small gametes because of their sexual difference. Size-different gametes within one species may arise from different types of gametogenesis depending on how many gametes one vegetative cell produces. These assumptions, i.e. the bipolarity of isogametes, in monoecious taxa, size-dependent sex determination, and different types of gametogenesis within one taxon, are justifiable since they are realized independently from each other in various species. It is postulated that mutational fixation of the size-dependent sex determination in such monoecious species creates cell lineages producing sex-different macro- and microgametes and establishes constitutional anisogamous dioecy. The proposed hypothesis clearly separates the evolution of anisogamy as a problem of sexual reproduction from the evolution of sex per se, sex being defined as a bipolarity effecting fertilization.     

Proteomics characterization of mouse kidney peroxisomes by tandem mass spectrometry and protein correlation profiling

The peroxisome represents a ubiquitous single membrane-bound key organelle that executes various metabolic pathways such as fatty acid degradation by alpha- and beta-oxidation, ether-phospholipid biosynthesis, metabolism of reactive oxygen species, and detoxification of glyoxylate in mammals. To fulfil this vast array of metabolic functions, peroxisomes accommodate approximately 50 different enzymes at least as identified until now. Interest in peroxisomes has been fueled by the discovery of a group of genetic diseases in humans, which are caused by either a defect in peroxisome biogenesis or the deficient activity of a distinct peroxisomal enzyme or transporter. Although this research has greatly improved our understanding of peroxisomes and their role in mammalian metabolism, deeper insight into biochemistry and functions of peroxisomes is required to expand our knowledge of this low abundance but vital organelle. In this work, we used classical subcellular fractionation in combination with MS-based proteomics methodologies to characterize the proteome of mouse kidney peroxisomes. We could identify virtually all known components involved in peroxisomal metabolism and biogenesis. Moreover through protein localization studies by using a quantitative MS screen combined with statistical analyses, we identified 15 new peroxisomal candidates. Of these, we further investigated five candidates by immunocytochemistry, which confirmed their localization in peroxisomes. As a result of this joint effort, we believe to have compiled the so far most comprehensive protein catalogue of mammalian peroxisomes.     

The human pyridoxal kinase, a plausible target for ginkgotoxin from Ginkgo biloba

Ginkgotoxin (4'-O-methylpyridoxine) occurring in the seeds and leaves of Ginkgo biloba, is an antivitamin structurally related to vitamin B(6). Ingestion of ginkgotoxin triggers epileptic convulsions and other neuronal symptoms. Here we report on studies on the impact of B(6) antivitamins including ginkgotoxin on recombinant homogeneous human pyridoxal kinase (EC 2.7.1.35). It is shown that ginkgotoxin serves as an alternate substrate for this enzyme with a lower K(m) value than pyridoxal, pyridoxamine or pyridoxine. Thus, the presence of ginkgotoxin leads to temporarily reduced pyridoxal phosphate formation in vitro and possibly also in vivo. Our observations are discussed in light of Ginkgo medications used as nootropics.     

Choosing tree genotypes for phytoremediation of landfill leachate using phyto-recurrent selection

Information about the response of poplar (Populus spp.) genotypes to landfill leachate irrigation is needed, along with efficient methods for choosing genotypes based on leachate composition. Poplar clones were irrigated during three cycles of phyto-recurrent selection to test whether genotypes responded differently to leachate and water, and to test whether the methodology had merit as a tool for plant selection during remediation. Fifteen below- and above-ground traits were evaluated. Twenty-five clones were tested in cycle 1, while the best l2 genotypes were evaluated in cycles 2 and 3. Eight clones were selected and subsequently tested in an in situ landfill study (cycle 4). Results from cycles 1, 2, and 3 are presented here. Overall, clones responded differently to irrigation treatments, with certain genotypes exhibiting better below- and above-ground growth with water than leachate. However, growth was greater with leachate irrigation for some clones. In addition, differences between treatments within clones decreased with days after planting (DAP). There were no treatment differences for number of leaves, height, and root length at the end of cycle 2 (45 DAP) or cycle 3 (30 DAP). These results detail the extensive variation in clonal responses to leachate irrigation, along with the need and efficacy of using phyto-recurrent selection to choose superior genotypes.