Cytokinin-induced switch in development in excised cotyledons of radiata pine cultured in vitro

Cotyledons of Pinus radiata D. Don were cultured under shoot-forming (plus cytokinin) and elongating (minus cytokinin) conditions. Using. autoradiographic and precursor incorporation techniques, the sites and rate of macromolecular synthesis were examined during the first five days in culture. Active incorporation of ³H-thymidine, ³ H-uridine and ³H-leucine occurred. In shoot-forming cotyledons the incorporation became preferentially located in the epidermal and sub-epidermal cell layers in contact with the medium. In elongating cotyledons, in contrast, incorporation was randomly distributed, and the amount of incorporation declined with time. Biochemically, differences in DNA, RNA and total protein synthetic patterns were observed. In elongating cotyledons the rates of RNA and protein synthesis were higher during the first 48 h than in shoot-forming tissues, after which the synthetic rates were similar. Two peaks of newly formed DNA were observed in both tissues. These findings indicate that the cytokinin-induced changes in developmental pathways began within 24 h in culture.     

Interactions of the neurotoxic amine 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine with monoamine oxidases.

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), a thermal breakdown product of a meperidine-like narcotic used by drug abusers as a heroin substitute, produces Parkinsonian symptoms in humans and primates. The nigrostriatal toxicity is not due to MPTP itself but to one or more oxidation products resulting from the action of monoamine oxidase (MAO) on this tertiary allylamine. Both MAO A and B catalyse the oxidation of MPTP to the 1-methyl-4-phenyl-2,3-dihydropyridinium species (MPDP+), which undergoes further oxidation to the fully aromatic 1-methyl-4-phenylpyridinium species (MPP+). These bio-oxidations are blocked by selective inhibitors of MAO A and B. Additionally, MPTP, MPDP+ and MPP+ are competitive inhibitors of MAO A and B. The A form of the enzyme is particularly sensitive to this type of reversible inhibition. Both MAO A and B also are irreversibly inactivated by MPTP and MPDP+, but not by MPP+. This inactivation obeys the characteristics of a mechanism-based or 'suicide' process. The inactivation, which is accompanied by the incorporation of radioactivity from methyl-labelled MPTP, is likely to result from covalent modification of the enzyme.     

PEG-mediated expression of GUS and CAT genes in protoplasts from embryogenic suspension cultures of Picea glauca

ß-Glucuronidase (GUS) and chloramphenicol acetyl transferase (CAT) were used as reporter proteins in protoplasts from embryogenic suspension cultures of Picea glauca (Moench) Voss (white spruce). Plasmid DNA enclosing chimeric GUS and CAT constructs, using the cauliflower mosaic virus 35S promoter, was introduced into Picea glauca protoplasts using polyethylene glycol (PEG). Transient expression was detected 12 to 40 h after PEG-mediated DNA delivery. Dose-response curves using covalently closed circular plasmid DNA, in the absence of carrier DNA, have been obtained for each of these reporter genes. Linearized plasmid DNA gave lower levels of expression than covalently closed circular plasmid DNA when assayed 40 h after PEG-mediated DNA transfer. The use of carrier DNA (herring sperm DNA), in combination with covalently closed circular plasmid DNA, increased the level of expression of GUS by about 50%. CAT expression was enhanced if PEG-mediated delivery was performed on ice rather than at room temperature. The highest level of expression for CAT, and the lowest signal-to-noise ratio, was found 24 h after PEG-mediated DNA transfer. Both GUS and CAT provided results that were quantifiable and can therefore be used as reporter genes in Picea glauca.Abbreviations CAT chloramphenicol acetyl transferase - GUS ß-glucuronidase - CaMV cauliflower mosaic virus - NOS nopaline synthase - CCC covalently closed circular DNA - L linear DNA - PEG polyethylene glycol - HS herring sperm DNA - P protoplasts - PCM protoplast culture medium - MES morpholinoethane-sulfonic acid - Cm chloramphenicol - Ac acetylated - MUG 4-methyl umbelliferyl ß-D-glucuronide - TLC thin layer chromatography     

Non-morphogenic metabolism during shoot induction in radiata pine cotyledon explants

Summary The effects of excision, light and cytokinin (N⁶-benzyladenine) on¹⁴C-acetate metabolism in cotyledons ofPinus radiata (D. Don) were determined.¹⁴CO₂ was released and the distribution of radioactivity into lipids, sugars, organic acids and amino acids was determined. While light and cytokinin generally caused some increase in metabolism, the effect of excision, i.e., wounding, was most pronounced. Specific metabolites examined (citrate, malate, succinate, alanine, aspartate, glutamate and glutamine) were at least 50% greater in¹⁴C-labeling in excised cotyledons as compared to intact seedlings. This enhancement of wound metabolism would mask possible morphogenically-related changes occurring at that time. This research was supported by the Natural Sciences and Engineering Research Council of Canada Grant A-6467 to T.A. Thorpe.     

Application of Deuterated A-Tocopherols to the Biokinetics and Bioavailability of Vitamin E

-Tocopherol, a superior chain-breaking, peroxyl radical-trapping antioxidant and the most active component of vitamin E, is elevated in liver tumor cells, contributing to their greater resistance towards lipid peroxidation compared to cells from normal tissues. Also, in regenerating rat liver the level of vitamin E has been found to fluctuate in phase with the rate of cell division. In order to study the biokinetcis and mechanisms of the distribution of vitamin E in organs and within tissues of animals, deuterated forms of -tocopherol have been synthesized and their uptake into blood and tissues has been measured by gas chromatography-mass spectrometry. Measurement of the competitive uptake from a mixture of the RRR-and SRR--tocopherol stereoisomers labelled with different amounts of deuterium shows that the liver exerts a strong preference for secretion of the natural (RRR) stereoisomer into the plasma. It is suggested that a tocopherol-binding protein plays a key role in this process.     

A re-examination of host relations in the Aphelinidae (Hymenoptera: Ghalcidoidea)

Host relations among the Aphelinidae (Hymenoptera: Chalcidoidea) are highly intriguing from an evolutionary view. Females are usually primary endoparasitoids of whiteflies or scale insects, whereas the development of the male is different and has been used for classification. In heteronomous hyperparasitoids (adelphoparasitoids or autoparasitoids) the male develops as a hyperparasitoid of conspecific females or other endoparasitoid species. We review the consequences of this mode of development which is unique because decisions concerning host selection are inextricably linked with progeny sex ratio. Autoparasitoid field sex ratios can fluctuate dramatically concurrent with changes in the relative availability of male and female hosts. A recent adaptive explanation for these sex ratios involves understanding the reproductive constraints acting on heteronomous parasitoids. Host relations in these parasitoids can show a degree of plasticity. We argue that in many instances autoparasitism may be facultative in nature and should not be used for classification. Heterotrophic parasitism, wherein the male develops as a primary parasitoid of lepidopterous eggs, has been poorly understood in the past due to uncertainties in reports of the biology and taxonomy of heterotrophic parasitoids. The host relations of this group are clarified.     

Recolonisation and recruitment of fishes to intertidal rockpools at Wellington, New Zealand

Synopsis A study of recolonisation of rockpools by intertidal fishes on the Wellington south coast, New Zealand, found the assemblage to be resilient and seasonally stable. A total of 26 species from nine families were recorded, dominated by the Tripterygiidae (triplefins) and Gobiesocidae (clingfishes). A pattern of alternating species dominance occurred, with the triplefinsBellapiscis medius andForsterygion lapillum being numerically dominant in summer, but becoming less common in winter and replaced as dominants by the clingfishesTrachelochismus pinnulatus andGastroscyphus hectoris. Juvenile recruits of eleven species occurred in the samples from spring to early summer, however only the aforementioned four species recruit to the intertidal zone in large numbers. The speed of rockpool recolonisation by fishes after extractive sampling is seasonally dependent, being quicker in the summer than winter. In general, recolonisation takes at least one month, but probably fewer than three. While stochastic factors influence assemblage composition in the short term, overall regulation of the fish assemblage of rockpools appears to be primarily deterministic, resulting in an essentially predictable taxonomic structure.     

Iron and copper nutrition-dependent changes in protein expression in a tomato wild type and the nicotianamine-free mutant chloronerva.

The nicotianamine-deficient mutant chloronerva resembles phenotypically an Fe-deficient plant despite the high accumulation of Fe in the leaves, whereas if suffers from Cu deficiency in the shoot. Two-dimensional electrophoretic separation of proteins from root tips and leaves of wild-type Lycopersicon esculentum Mill. cv Bonner Beste and the mutant grown with and without Fe showed a number of consistent differences. In root tips of the Fe-deficient wild type and the Fe-sufficient as well as the Fe-deficient mutant, the expression of glyceraldehyde-3-phosphate dehydrogenase, formate dehydrogenase, and ascorbate peroxidase was increased. In leaves of the Fe-sufficient and -deficient mutant, Cu-containing chloroplastic and cytosolic superoxide dismutase (Cu-Zn) and plastocyanin (Cu) were nearly absent. This low plastocyanin content could be restored by supplying Cu via the xylem, but the superoxide dismutase levels could not be increased by this treatment. The differences in the protein patterns between wild type and mutant indicate that the apparent Fe deficiency of mutant plants led to an increase in enzymes involved in anaerobic metabolism as well as enzymes involved in stress defense. The biosynthesis of plastocyanin was diminished in mutant leaves, but it was differentially induced by increased Cu content.