La dormance embryonnaire chez Taxus baccata: Influence de la composition du milieu liquide sur I'induction de la germination Par

Embryo dormancy of Taxus baccata var. fastigiata is eliminated when cultured continuously in nutritive liquid medium. An equivalent percentage of germination is obtained when the embryos are transferred to agar medium after 8 days of liquid culture. There is no morphological development of the embryo during the period in the liquid medium. But we have ascertained that water-soluble germination inhibitors present in the embryo are leached out into the medium, permitting germination. Germination is totally absent when the embryos are cultured continuously in distilled water, alone or with minerals; incidental in sucrose solution; and maximal when the medium contains sucrose and Ca²⁺ or K⁺ ions. The extent of germination on agar medium depends upon the composition of the liquid medium in which the embryos are cultured for the initial 8 days. But this preliminary culture in the liquid medium does not always remove the endogenous inhibitors, irrespective of its composition. This can be achieved only in the presence of sucrose; and this process can be made more effective by the addition of Ca²⁺ ions.     

Facilitated Penetration of Amanitin—Albumin Conjugates into Hepatocytes after Coupling with Fluorescein

α-AMANITIN, a cyclic peptide of the toadstool Amanita phalloides^1,2, causes necrosis of liver and kidney cells, the first morphological lesions occurring in the nuclei^3,4. It acts by binding to RNA polymerase in eukaryotic .cells and inhibiting the enzyme^5–9. The hepatotoxicity of amanitin increases several times when it is conjugated to albumin, probably because of a slower rate of elimination of the toxin through the glomeruli^4,10. It is unlikely that the amanitin-albumin conjugate enters the hepatocyte by a mechanism involving its albumin moiety; it was therefore suggested¹¹ that penetration of the liver cells is consequent on binding of the amanitin group to the carrier involved in transport of this peptide. This led us to consider more generally the facilitation of penetration into cells by large molecules by means of binding to another molecule for which a carrier exists on the cell membrane^12,13.     

Efficient secretion of two fungal cellobiohydrolases by Saccharomyces cerevisiae

Two different cellobiohydrolases, CBHI and CBHII, of the filamentous fungus Trichoderma reesei both hydrolyse highly crystalline cellulose. Cellulolytic strains of the yeast Saccharomyces cerevisiae were constructed by transferring cDNAs coding for these enzymes into yeast on an expression plasmid. These cellulolytic yeasts were able to secrete efficiently the large, heterologous proteins to the culture medium. The recombinant cellulases were observed to be heterogeneous in Mr due, at least partly, to variable N-glycosylation. Recombinant CBHII was able to bind to crystalline cellulose, although slightly less efficiently than the native enzyme. Both of the two recombinant cellulases were able to degrade amorphous cellulose. In a fermenter cultivation, around 100 micrograms/ml of CBHII was secreted into the yeast growth medium.     

Expression of dicistronic transcriptional units in transgenic tobacco.

We investigated whether the two cistrons of a dicistronic mRNA can be translated in plants to yield both gene products. The coding sequences of various reporter genes were combined in dicistronic units, and their expression was analyzed in stably transformed tobacco plants at the RNA and protein levels. The presence of an upstream cistron resulted in all cases in a drastically reduced expression of the downstream cistron. The translational efficiency of the gene located downstream in the dicistronic units was 500- to 1,500-fold lower than that in a monocistronic control; a 500-fold lower value was obtained with a dicistronic unit in which both cistrons were separated by 30 nucleotides, whereas a 1,500-fold lower value was obtained with a dicistronic unit in which the stop codon of the upstream cistron and the start codon of the downstream cistron overlapped. As a strategy to select indirectly for transformants with enhanced levels of expression of a gene which is by itself nonselectable, the gene of interest can be cloned upstream from a selectable marker in a dicistronic configuration. This strategy can be used provided that the amount of dicistronic mRNA is high. If, on the other hand, the expression of the dicistronic unit is too low, selection of the downstream cistron will primarily give clones with rearranged dicistronic units.     

Gerbera hybrida (Asteraceae) imposes regulation at several anatomical levels during inflorescence development on the gene for dihydroflavonol-4-reductase

In the ornamental cut flower plant Gerbera hybrida the spatial distribution of regulatory molecules characteristic of differentiation of the composite inflorescence is visualized as the various patterns of anthocyanin pigmentation of different varieties. In order to identify genes that the plant can regulate according to these anatomical patterns, we have analysed gene expression affecting two enzymatic steps, chalcone synthase (CHS) and dihydroflavonol-4-reductase (DFR), in five gerbera varieties with spatially restricted anthocyanin pigmentation patterns. The dfr expression profiles vary at the levels of floral organ, flower type and region within corolla during inflorescence development according to the anthocyanin pigmentation of the cultivars. In contrast, chs expression, although regulated in a tissue-specific manner during inflorescence development, varies only occasionally. The variation in the dfr expression profiles between the varieties reveals spatially specific gene regulation that senses the differentiation events characteristic of the composite inflorescence.     

Molecular modelling study of antigen binding to oxazolone-specific antibodies: the Ox1 idiotypic IgG and its mature variant with increased affinity to 2-phenyloxazolone

Structural models of the variable domains of the murine anti-2-phenyloxazolone IgG (Ox1 idiotype) and its somatic variant, which has higher affinity to the hapten 2-phenyloxazolone, were constructed by computer-aided model building using known structures of highly homologous immunoglobulins as templates. Molecular dynamics simulations were used to dock the hapten between the VL and VH domains. The hapten is predicted to bind to slightly different sites in the two models. Hypotheses concerning the role of a number of preferred mutations in anti-oxazolone variants are presented. These can be tested by mutagenesis and crystallography. In particular, the higher binding affinities of the different antibody variants are shown to correlate with better complementarity of electrostatics. The molecular dynamic simulations also suggest that two mobile tryptophans at the mouth of the pocket may play an important role in the binding of hapten.     

Genetic polymorphism and evolution in parthenogenetic animals

Summary The enzyme gene variability within parthenogenetic clones of Acyrtosiphon pisum has been followed by gel electrophoresis. No variation was observed within any clone. One enzyme locus was found to vary between clones. No evidence was found to support gene recombination due to the alleged endomeiosis. This hypothesis is proven to be also theoretically untenable. The low average heterozygosity in aphids is explained as a result of directional selection operating upon the parthenogenetic aphid clones, as a consequence of which the heterozygosity is lowered.Dedicated to Professor Friedrich Mechelke on the occasion of his sixtieth birthday     

Conversion of Phosphatidyl Choline to Phosphatidic Acid in Freeze Injured Rye and Wheat Cultivars

The effect of exposure to freezing temperature (?15°C) on leaf phospholipid composition of hardened rye (Secale cereale L.) and hardened wheat cultivars (‘Miranovskaja 808’, ‘Bezostaja 1’, ‘Short Mexican’ and ‘Penjamo 62’), which differ in their resistance to frost, was investigated. Hardening took place under natural conditions. All the seedlings attained an equal level of linolenic acid in their leaves during hardening. Exposure to freezing temperatures resulted in a loss of phosphatidyl choline and accumulation of phosphatidic acid in the leaves. The ratio of phosphatidic acid to phosphatidyl choline, but not the level of poly-unsaturated fatty acids in the leaves, was related to their ability to survive at low temperatures. As freezing injury is caused by the formation of ice crystals in both extra- and intracellular space, it is probable that the plasma membranes of the investigated cultivars differed with respect to their water permeability. It is concluded that the plants, depending on the degree of their resistance to cold, produce an unknown substance of lipidic nature upon exposure to cold, with the aid of which they adjust the transitional state of their membranes to the prevailing temperature and, at the same time, facilitate the efflux of water from the cells.