The multi-protein family of Arabidopsis sulphotransferases and their relatives in other plant species

All members of the sulphotransferase (SOT, EC 2.8.2.-) protein family use 3'-phosphoadenosine 5'-phosphosulphate (PAPS) as the sulphuryl donor and transfer the sulphonate group to an appropriate hydroxyl group of several classes of substrates. These enzymes have highly conserved domains and can be found in eubacteria and eukaryotes. In mammals, sulphate conjugation catalysed by SOTs constitutes an important reaction in the transformation of xenobiotics, and in the modulation of the biological activity of steroid hormones and neurotransmitters. In plants, sulphate-conjugation reactions seem to play an important role in plant growth, development, and adaptation to stress. To date only a few plant SOTs have been characterized in detail. The flavonol 3- and 4'-SOTs from Flaveria species (Asteraceae), which catalyse the sulphonation of flavonol aglycones and flavonol 3-sulphates, respectively, were the first plant SOTs for which cDNA clones were isolated. The plasma membrane associated gallic acid SOT of Mimosa pudica L. pulvini cells may be intrinsic to signalling events that modify the seismonastic response. In Brassica napus L. a SOT catalyses the O-sulphonation of brassinosteroids and thereby abolishes specifically the biological activity of 24-epibrassinolide. The fully sequenced genome of Arabidopsis thaliana Heynh. contains in total 18 genes that are likely to encode SOT proteins based on sequence similarities of the translated products with an average identity of 51.1%. So far only one SOT from A. thaliana (At5g07000) was functionally characterized: the protein was shown to catalyse the sulphonation of 12-hydroxyjasmonate and thereby inactivate excess jasmonic acid in plants. The substrates and, therefore, the physiological roles of SOTs are very diverse. By using the numerous informative databases and methods available for the model plant A. thaliana, the elucidation of the functional role of the SOT protein family will be accelerated.     

Na+ -D-glucose cotransporter in the kidney of Leucoraja erinacea: molecular identification and intrarenal distribution

Studies on membrane vesicles from the kidney of Leucoraja erinacea suggested the sole presence of a sodium-D-glucose cotransporter type 1 involved in renal D-glucose reabsorption. For molecular characterization of this transport system, an mRNA library was screened with primers directed against conserved regions of human sglt1. A cDNA was cloned whose nucleotide and derived amino acid sequence revealed high homology to sodium glucose cotransporter 1 (SGLT1). Xenopus laevis oocytes injected with the respective cRNA showed sodium-dependent high-affinity uptake of D-glucose. Many positions considered functionally essential for sodium glucose cotransporter 1 (SGLT1) are also found in the skate protein. High conservation preferentially in transmembrane helices and small linking loops suggests early appearance and continued preservation of these regions. Larger loops, especially loop 13, which is associated with phlorizin binding, were more variable, as is the interaction with the specific inhibitor in various species. To study the intrarenal distribution of the transporter, a skate SGLT1-specific antibody was generated. In cryosections of skate kidney, various nephron segments could be differentiated by lectin staining. Immunoreaction with the antibody was observed in the proximal tubule segments PIa and PIIa, the early distal tubule, and the collecting tubule. Thus Leucoraja, in contrast to the mammalian kidney, employs only SGLT1 to reabsorb d-glucose in the early, as well as in the late segments of the proximal tubule and probably also in the late distal tubule (LDT). Thereby, it differs also partly from the kidney of the close relative Squalus acanthias, which uses SGLT2 in more distal proximal tubule segments but shows also expression in the later nephron parts.     

Metabolism and Plant Hormone Action During Clubroot Disease

Infection of Brassicaceae with the obligate biotrophic pathogen Plasmodiophora brassicae results in the development of root galls (clubroots). During the transformation of a healthy root to a root gall a plethora of changes in primary and secondary metabolism occur. The upper part of an infected plant is retarded in growth due to redirection of assimilates from the shoot to the root. In addition, changes in the levels of plant growth regulators, especially auxins and cytokinins, contribute to the hypertrophy of infected roots. Also, defense reactions are manipulated after inoculation of suitable host plants with P. brassicae. This review summarizes our current knowledge on the changes in these parameters. A model is presented for how primary metabolism and secondary metabolism, including plant hormones, interact to induce clubroot formation.     

In vitro anticancer studies of α- and β-d-glucopyranose betulin anomers

Four derivatives of betulin containing a d-glucopyranosyl moiety at C3 position were synthesized and characterized by ¹H and ¹³C NMR spectroscopy as well as mass spectrometry. The crystal structure of 28-O-acetylbetulin-3-yl-β-d-(2′,3′,4′,6′-tetra-O-acetyl)glucopyranoside was determined. The compounds were tested against fifteen tumor cell lines of different histogenic origins. The α- and β-anomers of 28-O-acetylbetulin-3-yl-d-glucopyranoside, exerted a dose dependent antiproliferative action towards the tumor cell lines. Treatment of HCT-116 cells for 24 h induced apoptosis, which was confirmed by the appearance of a typical ladder pattern in the DNA fragmentation assay and cell cycle analysis. The α- and β-anomers of 28-O-acetylbetulin-3-yl-d-glucopyranoside seem to induce apoptosis by activation of different upstream caspases on colon cancer HCT-116 cell line.     

Extracellular Neuroactive Amino Acids in the Rat Striatum During Ischaemia: Comparison Between Penumbral Conditions and Ischaemia with Sustained Anoxic Depolarisation

Abstract: Changes in the extracellular levels of excitatory and inhibitory amino acid transmitters were studied in the rat striatum during penumbral ischaemia using intracerebral microdialysis. Effects of penumbral forebrain ischaemia were compared with those of ischaemia with sustained anoxic depolarisation and K⁺ (100 m M ). Comparisons were also made between different groups of animals at 2 and 24 h after dialysis probe implantation. The K⁺ stimulus did not provoke any release of excitatory amino acids in the 24-h group, probably reflecting a decrease of functional synapses adjacent to the probe. During 30 min of penumbral ischaemia, excitatory amino acids did not reach critical concentrations in the extracellular fluid, and increases in levels of inhibitory/modulatory amino acids were similar. On the other hand, severe transient ischaemia resulted in massive synchronous release of many neuroactive excitatory and inhibitory compounds, in both the 2- and 24-h groups. These and other data suggest that changes during severe ischaemia may arise from both neurotransmitter and metabolic pools. It is concluded that is- chaemic damage in the penumbra may not be related to extracellular neuroactive amino acid changes generated within this region.     

A microtest system for the serial assay of phytotoxic compounds using photoautotrophic cell suspension cultures of Chenopodium rubrum

Chenopodium rubrum photoautotrophic cell suspensions were grown in plastic tissue culture dishes under photoautotrophic conditions. Growth was monitored by measuring cell number, packed cell volume, chlorophyll content and oxygen production. Such microtiter dishes are suitable systems for the serial assay of growth inhibition and various physiological effects (i.e. chlorophyll fluorescence, cell viability, oxygen production) of photoautotrophic cells as caused by herbicides and fungal phytotoxins. The applicability of the test system is discussed.Abbreviations pcv packed cell volume - fr.w. fresh weight - rpm revol. per minute - DMSO dimethyl sulfoxide - PMS phenazine methosulfate - NBT nitro-blue tetrazolium chloride     

Seminal prostaglandins in men with subnormal sperm motility and therapeutic treatment

The contents of prostaglandins in seminal plasma from a total of 73 men were evaluated. The subjects were grouped as follows: normospermic men, patients with impaired motility, patients with small untreated varicocelle and patients with impaired motility and Kallikrein therapy. Sperm density, morphology and motility were examined. High performance reversed phase liquid chromatography (HPLC) in combination with specific radioimmunoassays were used for the determination of PGE₂, PGI₂ and PGF_2α. There was a significant difference (p < 0, 025; F-test) between the PGI₂ concentrations in patients with impaired motility (5,6 ± 1,4 pg/mg protein) and normal men (8,8 ± 3,7 pg/mg protein). PGE₂ and PGF_2α were significantly different in patients with varicocele (p < 0,025, F-test). Wide ranges of prostaglandins occured in the Kallikrein-group with no significant differences. We conclude that: a) PGI₁ is an additional prostaglandin compound in seminal plasma. b)its measurement may not be useful as diagnostic parameter in subfertile men and c) Kallikrein has no influence on the prostaglandin content in seminal plasma and other seminal parameters as motility, motility index and sperm counts.