Photosynthetic characteristics and tolerance to photo-oxidation of transgenic rice expressing C4 photosynthesis enzymes

The photosynthetic characteristics of four transgenic rice lines over-expressing rice NADP-malic enzyme (ME), and maize phosphoenolpyruvate carboxylase (PC), pyruvate,orthophosphate dikinase (PK), and PC+PK (CK) were investigated using outdoor-grown plants. Relative to untransformed wild-type (WT) rice, PC transgenic rice exhibited high PC activity (25-fold increase) and enhanced activity of carbonic anhydrase (more than two-fold increase), while the activity of ribulose-bisphosphate carboxylase/oxygenase (Rubisco) and its kinetic property were not significantly altered. The PC transgenic plants also showed a higher light intensity for saturation of photosynthesis, higher photosynthetic CO₂ uptake rate and carboxylation efficiency, and slightly reduced CO₂ compensation point. In addition, chlorophyll a fluorescence analysis indicates that PC transgenic plants are more tolerant to photo-oxidative stress, due to a higher capacity to quench excess light energy via photochemical and non-photochemical means. Furthermore, PC and CK transgenic rice produced 22–24% more grains than WT plants. Taken together, these results suggest that expression of maize C₄ photosynthesis enzymes in rice, a C₃ plant, can improve its photosynthetic capacity with enhanced tolerance to photo-oxidation. This revised version was published online in June 2006 with corrections to the Cover Date.     

Biosynthesis of anthraquinones in cell cultures of Cinchona 'Robusta' proceeds via the methylerythritol 4-phosphate pathway.

Robustaquinone B was found as a major anthraquinone in cell cultures of Cinchona 'Robusta' after treatment with a fungal elicitor. Anthraquinones in Cinchona are considered to be of the Rubia type, i.e. rings A and B are derived from chorismate and alpha-ketoglutarate, whereas ring C is formed from isopentenyl diphosphate (IPP). To determine the origin of IPP, either formed via the mevalonic acid pathway or the 2-C-methyl-D-erythritol 4-phosphate pathway, the incorporation of [1-13C]glucose into robustaquinone B was studied. The 13C labeling of robustaquinone B was analyzed by one- and two-dimensional NMR spectroscopy and the labeling pattern was compared with the hypothetical labeling patterns obtained via the different biosynthetic pathways. The results clearly show that the IPP, constituting the ring C of robustaquinone B, is biosynthesized via the 2-C-methyl-D-erythritol 4-phosphate pathway. Moreover, the data also confirm that rings A and B of robustaquinone B are formed from chorismate and alpha-ketoglutarate via o-succinylbenzoate.     

NMR spectroscopy and chemical studies of an arabinan-rich system from the endosperm of the seed of Gleditsia triacanthos

Exhaustive extraction of the endosperm from the seed of Gleditsia triacanthos using water at room temperature and 50 degrees C left a residue, which was further extracted at 95 degrees C. Precipitation of this extract with 2-propanol yielded major amounts of galactomannan components, while the supernatant was mainly composed of arabinose-rich constituents. Two fractions were obtained by anion-exchange chromatography. The fraction that eluted with water is an arabinan with (1-->5) alpha-L linkages and branching mainly on C-2, accompanied with equal amounts of a low-galactose galactomannan oligosaccharide, and a small proportion of a beta-(1-->4)-galactan. The fraction eluted with an increased ionic strength consists mainly of a similar arabinan, and lower proportions of a high-galactose galactomannan, galactan, and protein. The arabinan moiety in both fractions was characterized by chemical analysis and 1D and 2D NMR spectroscopic techniques.     

Suppression of tumor promoter phorbolmyristate acetate-induced chromosome breakage by antioxidants and inhibitors of arachidonic acid metabolism

To gain insight into the mechanism of formation of chromosomal aberrations by the tumor promoter phorbolmyristate acetate (PMA) in human lymphocytes, we investigated the effect of antioxidants and inhibitors of arachidonic acid metabolism. Among the antioxidants bovine erythrocyte CuZn superoxide dismutase, glutathione peroxidase, mannitol (a scavenger of hydroxyl radicals), butylated hydroxytoluene and butylated hydroxyanisole were anticlastogenic while catalase and dimethylfuran (a scavenger of singlet oxygen) were inactive. These results show that the induction of aberrations by PMA occurs via indirect action, i.e. the intermediacy of superoxide and hydroxyl radicals. The following inhibitors of arachidonic acid metabolism were strongly anticlastogenic: the cyclo-oxygenase inhibitors indomethacin and flufenamic acid and the lipoxygenase inhibitor BN1015. Imidazole, nordihydroguaiaretic acid BN 1048 and 5,8,11,14-eicosatetraynoic acid were moderately active. The inhibitor of phospholipase A₂, fluocinolone acetonide, was also anticlastogenic.

We conclude that the oxidative metabolism of arachidonic acid is involved in the induction of chromosomal aberrations by PMA in human lymphocytes. However, because of the limited selectivity of these drugs, it is not yet possible to identify unambiguously the step(s) in the arachidonic acid cascade responsible for PMA clastogenicity.     

The rapid polyphosphoinositide metabolism may be a triggering event for thrombin-mediated stimulation of human platelets

The metabolism of polyphosphoinositides was examined in human platelets activated by thrombin. The addition of thrombin to [3H]glycerol-labeled platelets induced an initial loss and a subsequent increase of the radioactivity in phosphatidylinositol-4,5-bisphosphate (TPI) without any significant change in phosphatidylinositol-4-phosphate (DPI). A marked enhancement of [32P]Pi incorporation into TPI occurred in parallel with an increase in this lipid content, which was accompanied with a conccurent decrease in phosphatidylinositol (PI). The rate of this subsequent increase in TPI was smaller than that observed in [3H]arachidonic acid-labeled platelets, suggesting that formed TPI in activated platelets may contain much greater amount of arachidonate than preexisting TPI in resting platelets. These data indicate that thrombin causes a rapid change in TPI metabolism (initial degradation of preexisting TPI and subsequent production of arachidonate-rich TPI), which might be a primary candidate to modulate thrombin-induced function in human platelets.     

Hemoglobin Dallas (α97(G4)Asn→Lys):functional characterization of a high oxygen affinity natural mutant

Hemoglobin Dallas, an α-chain variant with a substitution of lysine for asparagine at position 97(G4), was found to have increased oxygen affinity ($\text{p}{}_{\mathrm{<mtext>1</mtext><mtext>2</mtext>}}\text{= 1}\text{mmHg at pH}\text{7.3}\text{and}\text{20°C), diminished cooperativity (0n,}\text{the Hill coefficient}\text{= 1.7}$) and reduced Bohr effect (about 50%). Addition of allosteric effectors (such as 2,3-diphosphoglycerate, inositol hexakisphosphate and bezafibrate) led to a decrease in oxygen affinity and increase in cooperative energy. Kinetic studies at pH 7.0 and 20°C revealed that (i), the overall rate of oxygen dissociation is 1.4-fold slower than that for HbA and (ii), the carbon monoxide dissociation rate is unaffected. The abnormal properties of this hemoglobin variant can be atttributed to a more ‘relaxed’ T-state.     

A compound CP-31398 suppresses excitotoxicity-induced neurodegeneration

Neurodegeneration causes dysfunction and degeneration of neurons and is triggered by various factors including genetic defects, free radicals, injury, and glutamate excitotoxicity. Among those, glutamate excitotoxicity is implicated in chronic disorders including AD and ALS, and in acute insults in the CNS including traumatic brain injury. Neurological disorders show hallmark morphological abnormalities such as axon degeneration and cell body death. The molecular mechanisms underlying excitotoxicity-induced neurodegeneration are complex and deciphering a molecular mechanism from one angle is beneficial to understand the process, however, still difficult to develop strategies to suppress excitotoxicity-induced degeneration due to existence of other mechanisms. Thus, directly identifying compounds that can modulate excitotoxicity-induced neurodegeneration and subsequently clarifiying the molecular mechanism is a valid approach to develop effective strategies to suppress neurodegeneration. We searched for compounds that can suppress excitotoxicity-induced neurodegeneration and found that CP-31398, a known compound that can rescue the structure and function of the tumor suppressor protein p53 mutant form and stabilize the active conformation of the p53 wild-type form, suppresses excitotoxicity-induced axon degeneration and cell body death. Moreover, CP-31398 suppresses mitochondrial dysfunction which has a strong correlation with excitotoxicity. Thus, our findings identify a compound that can serve as a novel modulator of neurodegeneration induced by glutamate excitotoxicity.