Characterization of gamma-tocopherol methyltransferases from Capsicum annuum L and Arabidopsis thaliana.

Tocopherols are essential micronutrients in human and animal nutrition due to their function as lipophilic antioxidants. They are exclusively synthesized by photosynthetic organisms including higher plants. Despite the attributed beneficial health effects and many industrial applications, research on the tocopherol biosynthetic pathway and its regulation in plants is still limited. In the work presented here we performed a detailed biochemical characterization of a gamma-tocopherol methyltransferase (gamma-TMT) from Arabidopsis thaliana and of a gamma-TMT purified from Capsicum annuum fruits, a tissue with high accumulation of tocopherols. The biochemical characteristics of both enzyme preparations were remarkably similar including substrate specificities. Both enzymes converted delta- and gamma- into beta- and alpha-tocopherol, respectively, but beta-tocopherol was not accepted as a substrate, pointing to a specific methylation at the C(5)-position of the tocopherol aromatic head group. A kinetic analysis performed with the Arabidopsis enzyme was consistent with an iso-ordered bi-bi type reaction mechanism. Our results emphasize the role of gamma-TMT in regulating the spectrum of accumulated tocopherols in plants.     

Impaired expression of the plastidic ferrochelatase by antisense RNA synthesis leads to a necrotic phenotype of transformed tobacco plants

Protoporphyrin IX is the last common intermediate of tetrapyrrole biosynthesis. The chelation of a Mg2+ ion by magnesium chelatase and of a ferrous ion by ferrochelatase directs protoporphyrin IX towards the formation of chlorophyll and heme, respectively. A full length cDNA clone encoding a ferrochelatase was identified from a Nicotiana tabacum cDNA library. The encoded protein consists of 497 amino acid residues with a molecular weight of 55.4 kDa. In vitro import of the protein into chloroplasts and its location in stroma and thylakoids confirm its close relationship to the previously described Arabidopsis thaliana plastid-located ferrochelatase (FeChII). A 1700-bp tobacco FeCh cDNA sequence was expressed in Nicotiana tabacum cv. Samsun NN under the control of the CaMV 35S promoter in antisense orientation allowing investigation into the consequences of selective reduction of the plastidic ferrochelatase activity for protoporphyrin IX channeling in chloroplasts and for interactions between plastidic and mitochondrial heme synthesis. Leaves of several transformants showed a reduced chlorophyll content and, during development, a light intensity-dependent formation of necrotic leaf lesions. In comparison with wild-type plants the total ferrochelatase activity was decreased in transgenic lines leading to an accumulation of photosensitizing protoporphyrin IX. Ferrochelatase activity was reduced only in plastids but not in mitochondria of transgenic plants. By means of the specifically diminished ferrochelatase activity consequences of the selective inhibition of protoheme formation for the intracellular supply of heme can be investigated in the future.     

Bacillus anthracis pXO1 virulence plasmid encodes a type 1 DNA topoisomerase

The virulence plasmid pXO1 is responsible for toxin production in Bacillus anthracis. A DNA fragment from pXO1 was isolated and was shown, by sequence analysis, to contain part of a type 1 DNA topoisomerase gene. Attempts to clone the entire wild-type gene, designated topX, in Escherichia coli, were unsuccessful. In order to obtain the complete gene, it was first insertionally inactivated and then cloned in the mutated form. The deduced amino acid sequence of Topo X1 shows similarities to that of the two E. coli type 1 DNA topoisomerases. The N-terminal two-thirds of the putative B. anthracis protein exhibits strongest sequence similarity to topoisomerase III, whereas the C-terminal portion contains cysteine residues that could form three zinc-binding domains, as they do in topoisomerase I. The suggested active-site tyrosine is conserved in all three proteins. The regulation of expression from the topX promoter is modified by addition of a gyrase inhibiting antibiotic. The Topo X1 protein is likely to be involved in the stability of pXO1.     

Carbon dioxide induces endotrophic germ tube formation in Candida albicans

Candida albicans formed germ tubes when exposed to air containing 5 to 15% carbon dioxide (CO2). The CO2-mediated germ tube formation occurred optimally at 37 degrees C in a pH range of 5.5 to 6.5. No germ tubes were produced at 25 degrees C, even when the optimal concentration of CO2 (10%) was present in the environment. The requirement of CO2 for germ tube formation could be partially substituted by sodium bicarbonate but not by N2. Carbon dioxide was required to be present throughout the entire course of germ tube emergence suggesting that its role is not limited to an initial triggering of morphogenic change. We suggest that carbon dioxide may be a common effector responsible for the germ tube promoting activity of certain chemical inducers for C. albicans.     

Effect of visible light on carotenogenesis in arthroconidiating Trichophyton mentagrophytes

The effect of visible light on carotenoid content in the dermatophyte Trichophyton mentagrophytes ATCC 26323 was investigated. The fungus T. mentagrophytes accumulated several carotenoids when arthroconidiated on Sabouraud glucose agar at 37°C. When this fungus was irradiated with moderate fluence rates of white light, the resultant arthroconidia contained considerably less carotenoids in comparison with dark controls although growth and arthroconidiation of this fungus were not at all affected by visible light. The reduction of carotenoid content in arthroconidia was due primarily to blue light, although red light caused a slight decrease in pigmentation. The suppressive effect of visible light on pigmentation was fluence rate dependent. Carotenoid accumulation in arthroconidia was inversely and exponentially related to the fluence rate of light. Carotenoid formation in arthroconidiating T. mentagrophytes was neither photoinducible nor photostimulative. An analysis of isolated carotenoids revealed that visible light caused a quantitative reduction in pigmentation, and no single carotenoid was selectively decreased.Non-standard abbreviations PI pigmentation index - r coefficient of correlation     

Mouse chromosome 15

Committee Members: R. Duncan and J. Todd.     

An exotic species spreads through Europe: Tygarrup javanicus (Chilopoda: Geophilomorpha: Mecistocephalidae) is reported from the Slovakia and the Czech Republic

Geophilomorph centipedes, due to their soil life style, are frequently transported with plants. This report tries to underline some adaptations that facilitate them being introduced and becoming alien species. Tygarrup javanicus (Attems, 1907), tropical species reported recently from different greenhouses in Central Europe, is used as an example of such a spreading species.     

DIEL RHYTHM OF ALGAL PHOSPHATE UPTAKE RATES IN P‐LIMITED CYCLOSTATS AND SIMULATION OF ITS EFFECT ON GROWTH AND COMPETITION1

Oscillations in the phosphate (P_i) uptake rates for three species of green algae were examined in a P‐limited cyclostat. For Ankistrodesmus convolutus Corda and Chlorella vulgaris Beyerinck, the P_i uptake rates increased during the daytime and decreased at night. In contrast, Chlamydomonas sp. exhibited the opposite uptake pattern. Cell densities also oscillated under a light:dark cycle, dividing at a species‐specific timing rather than continuously. In general, the cell densities exhibited an inverse relationship with the P_i uptake rates. A competition experiment between A. convolutus and C. vulgaris in a P‐limited cyclostat resulted in the dominance of C. vulgaris, regardless of the relative initial cell concentrations. Chlorella vulgaris also dominated in a mixed culture with Chlamydomonas sp., irrespective of the initial seeding ratio and dilution rate. However, Chlamydomonas sp. and A. convolutus coexisted in the competition experiment with gradual decrease of Chlamydomonas sp. when equally inoculated. Mathematical expressions of the oscillations in the P_i uptake rate and species‐specific cell division gate were used to develop a simulation model based on the Droop equation. The simulation results for each of the species conformed reasonably well to the experimental data. The results of the competition experiments also matched the competition simulation predictions quite well, although the experimental competition was generally more delayed than the simulations. In conclusion, the model simulation that incorporated the effect of diel rhythms in nutrient uptake clearly demonstrated that species diversity could be enhanced by different oscillation patterns in resource uptake, even under the condition of limitation by the same resource.     

Uptake and metabolism of biotin by human peripheral blood mononuclear cells

We studied the uptake of biotin into human peripheral bloodmononuclear cells (PBMC) using[³H]biotin and studiedthe catabolism of biotin in PBMC using[¹⁴C]biotin. Over 30 min, [³H]biotin uptakewas greater at 37°C than at 25°C(K_T = 2.6 ± 0.4 nM, maximal velocity = 2.9 ± 0.2 fmol · 10⁶cells¹ · 30 min¹). Ouabain reduced[³H]biotin uptake to65% of control values, suggesting that biotin uptake is Na-K-ATPasedependent. Unlabeled biotin and biotin analogs reduced the uptake of[³H]biotin to22-70% of control values, suggesting the presence of acompetition for a structurally specific biotin transporter. Whenendocytosis by PBMC was stimulated by various acyl glycerols, [³H]biotin uptake was40-73% of control values; these data are consistent with thehypothesis that stimulated endocytosis reduces biotin transporterdensity on the cell surface. During a 168-h incubation, PBMC did notcatabolize[¹⁴C]biotin.