Regulation of the amylase secretion by the yeast Filobasidium capsuligenum and a 2-deoxy-d-glucose resistant mutant

Summary The regulation of extracellular amylase production by the basidiomycetous yeast Filobasidium capsuligenum CCY 64-5-1 was characterized using growing and resting cells. A basal level of amylolytic activity was produced with various carbon sources including glucose. Amylase secretion was repressed by glucose and, more severely, by 2-deoxy-d-glucose, whereas compounds with -1,4-linked glucose, such as methyl glucoside, maltose, -cyclodextrin and soluble starch, served as inducers. Repression was not relieved by exogenously added cAMP. The effects of several metabolic inhibitors on amylase secretion were studied. Following UV-mutagenesis a mutant strain (FC-5) capable of growing in a 2-deoxy-d-glucose supplemented corn starch medium was selected for further characterization. This strain produced more amylase, had acquired an increased resistance against repression by glucose, and retained a growth rate comparable to the wild type. FC-5 was also characterized by a reduced glucokinase activity and an increased hexokinase activity.     

Biphasic fluence-response curves for phytochrome-mediated kalanchoë seed germination : sensitization by gibberellic Acid

The fluence-response curves for the effect of two red pulses separated by 24 hours on the germination of Kalanchoe blossfeldiana Poelln. cv Vesuv seeds, incubated on gibberellic acid (GA₃) are biphasic for suboptimal concentrations. The response in the low fluence range corresponds with a classical red/far-red reversible phytochrome mediated reaction. GA₃ induces an additional response in the very low fluence range, which is also phytochrome mediated. The sensitivity to phytochrome-far-red absorbing form (Pfr), however, is increased about 20,000-fold, so that even far-red fluences become saturating. Both in the very low and low fluence response range, the maximal responses induced by saturating fluences are modulated by the GA₃ concentration. GA₃ having no direct influence on the phytochrome phototransformations, alters the Pfr requirement and determines the responding seed population fraction in the very low and low fluence range. The effet of GA₃ appears to be on the transduction chain of the phytochrome signal.     

Relative biological effectiveness (RBE) of neutrons produced by 50 MeV deuterons and by 34, 45, 65 and 75 MeV protons

RBE of p(34) + Be, p(45) + Be, p(65), + Be, p(75) + Be and d(50) + Be neutron beams produced at the cyclotron "Cyclone" of Louvain-la-Neuve were measured. The biological criterion was the regeneration of the crypts of the intestinal mucosa (50 regenerated crypts per circumference) after abdominal irradiation in mice. Taking the p(65) + Be neutrons as reference, RBE values were found equal to 1.12, 1.07, 1.00 (Ref.), 0.96 and 1.02 respectively. These results are consistent with those published for cell lethality in vitro. However, the RBE variation is smaller than this previously obtained in the laboratory for growth inhibition in Vicia faba.     

The nuclear migration signal of Xenopus laevis nucleoplasmin.

Nucleoplasmin is the most abundant protein in the nucleus of Xenopus laevis oocytes. Its ability to target to the nucleus when microinjected into the cytoplasm has been the subject of many studies central to our understanding of how proteins segregate into nuclei. Using a cDNA clone we constructed beta-galactosidase-nucleoplasmin hybrids in modified bacterial expression vectors. The fusion proteins were expressed in Escherichia coli, purified and injected into the cytoplasm of X. laevis oocytes. The distribution of the fusion proteins between the cytoplasmic and nuclear compartments were analysed after incubation of various lengths of time. The results show that the signal sequence for nuclear transport is located close to the carboxy terminus of the protein. The signal sequence has been mapped to a small stretch of amino acids, containing a stretch of four lysines analogous to the SV40 large-T antigen signal.     

Heparan sulfate proteoglycans of human lung fibroblasts. Structural heterogeneity of the core proteins of the hydrophobic cell-associated forms

Heparan sulfate proteoglycans (HSPG) were solubilized from human lung fibroblast monolayers with detergent. Presumptive membrane-associated forms displaying hydrophobic properties were purified by gel filtration on Sepharose CL-4B, by ion-exchange chromatography on Mono Q and by incorporation in lipid vesicles. The HSPG preparations were 125I-iodinated and treated with heparitinase before sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Five radiolabeled proteins with apparent molecular weights of 125,000, 90,000, 64,000, 48,000, and 35,000 were visualized by autoradiography. A sixth protein, identified in nonreduced 125I-HSPG preparations, appeared as a non-HS chain-bearing Mr 35,000 peptide which was disulfide-linked to an HS chain-bearing peptide of similar size. This multiplicity of core proteins did not seem to result from proteolysis during the heparitinase treatment itself, since some of the core proteins migrated independently during gel filtration before heparitinase digestion. Moreover, heparitinase digestion of 125I-HSPG purified by affinity chromatography on an immobilized monoclonal antibody yielded only the Mr 64,000 protein. Alternative depolymerizations of the HS chains by heparinase or HNO2 also yielded multiple protein bands. These results imply that heterogeneity of the core protein moiety may be a genuine property of the hydrophobic HSPG of human lung fibroblasts. The occurrence of multiple integral membrane HSPG forms may be relevant for the multiple functions that have been ascribed to cell-surface HSPG.     

Dynamics of long-leaved sundew Drosera intermedia populations at two extremes of a hydrological gradient

Densities of Drosera intermedia were low in two studied habitats (10–25 ramets m⁻¹), a path through a wet heath (short inundation in spring, low soil moisture in summer) and a pool edge (longlasting inundation, high soil moisture in summer). The low densities could be explained by the observed low recruitment and high adult mortality.
The low recruitment resulted from: (1) a high first year mortality of the large number of seedlings that emerged each year in the path population, caused by summer drought and cover with algae after heavy rainfall; (2) the absence in two years out of three of seedling emergence at the pool edge, due to the longlasting inundation. In neither population any seedlings survived to flower; (3) low vegetative reproduction rate.
Adult mortality during the growing season was caused by drought, which did not occur at the pool edge. Rapid senescence in autumn, caused by summer drought on the path and by a rapid submersion after heavy rainfall at the pool edge, was associated with a high winter mortality.     

Differentiation between Xanthomonas campestris pv. graminis (ISPP List 1980), pv. phleipratensis (ISPP List 1980) emend., pv. poae Egli and Schmidt 1982 and pv. arrhenatheri Egli and Schmidt 1982, by Numerical Analysis of Phenotypic Features and Protein Gel Electrophoregrams

A numerical analysis of 257 phenotypic features of 45 bacterial isolates from grasses, revealed three phenons corresponding to (i) X. campestris pv. graminis (ISPP List 1980), (ii) X. campestris pv. phleipratensis (ISPP List 1980) and (iii) X. campestris pv. poae Egli and Schmidt 1982 and X. campestris pv. arrhenatheri Egli and Schmidt 1982. In each phenon, the strains clustered together regardless of the geographical origin of the isolates orthe year of isolation. Polyacrylamide gel electrophoresis of soluble proteins and host range studies, revealed four groups corresponding to the pathovars mentioned above. The four pathovars constitute definite biological entities that can be differentiated by phenotypic, gel electrophoretic and host range features.     

Interdependence of coenzyme-induced conformational work and binding potential in yeast alcohol and porcine heart lactate dehydrogenases: a hydrogen-deuterium exchange study

Binding of NAD coenzymes to yeast alcohol dehydrogenase (YADH) and porcine heart lactate dehydrogenase (PHLDH) was studied by hydrogen-deuterium exchange with the infrared technique. Conformational changes in the enzymes specific to the coenzymes and their fragments were observed, and the pH dependence of the exchange reaction shows that it conforms to the EX-2 scheme. In both YADH and PHLDH the magnitude of the conformational change of measured by exchange retardation is considerably larger for NAD+ than for NADH. Studies with coenzyme fragments like ADP-ribose, ADP, and AMP also highlight the lack of rigorous correlation between structural features such as charge and size and their influence on exchange behavior. Ternary complexes such as YADH-NAD+-pyrazole, PHLDH-NAD+-oxalate, and PHLDH-NADH-oxamate, which mimic the transition state, have a significantly more pronounced effect on exchange rates than the corresponding binary complexes. The outstanding feature of this study is the demonstration that in the binary enzyme-coenzyme complexes the more loosely bound NAD+ is more effective in retarding exchange than the more firmly bound NADH. These differences are attributed to the unequal structural constraints exerted by the two coenzymes upon the enzymes, which translate to unequal expenditure of transconformational work in the formation of the two complexes. The opposing variation in the free energy of binding and the transconformational work expended can be viewed as an unequal partitioning of the net free energy gain resulting from the protein-ligand interaction into a binding term and that required for conformational change.     

Incorporation of the carbocyclic analogue of (E)-5-(2-bromovinyl)-2'-deoxyuridine 5'-triphosphate into a synthetic DNA

The carbocyclic analogue of (E)-5-(2-bromovinyl)-2'-deoxyuridine, C-BVDU, is a very potent and selective anti-herpes-virus compound. In order to synthesize and study the properties of a DNA that contains C-BVDU, the 5'-triphosphate, C-BVDUTP was prepared and evaluated as a potential substrate of the E. coli Klenow DNA polymerase enzyme. Although C-BVDUTP proved to be a very poor substrate also of this enzyme, it could be incorporated up to 3.6% into the synthetic DNA, poly(dA-dT, C-BVDU). This level of substitution decreased significantly the template activity for DNA and RNA polymerases, as compared to that of poly(dA-dT).