Cell-specific fatty acylation of proteins in cultured cells of neuronal and glial origin

Distinct sets of cellular proteins were labeled with [³H]myristic and [³H]palmitic acids in primary (rat neurons and astroglia) and continuous (murine N1E-115 neuroblastoma and rat C6 glioma) cell cultures derived from the nervous system. Both soluble and membrane proteins were modified by myristate in a hydroxylamine-stable (amide) linkage, while palmitoylated proteins were esterlinked and almost exclusively membrane bound. Chain elongation of both labeled fatty acids prior to acylation was observed, but no protein amide-liked [³H]myristate originating from [³H]palmitate was detected. Fatty acylation profiles differed considerably among most of the cell lines, except for rat astroglial and glioma cells in which myristoylated proteins appeared to be almost identical based on SDS gel electrophoresis. An unidentified 47 kDa myristoylated protein was labeled to a significantly greater extent in astroglial than in glioma cells; the expression of this protein could be related to transformation or development in cells of glial origin.     

Lignin Peroxidase Activity Is Not Important in Biological Bleaching and Delignification of Unbleached Kraft Pulp by Trametes versicolor

The discovery in 1983 of fungal lignin peroxidases able to catalyze the oxidation of nonphenolic aromatic lignin model compounds and release some CO₂ from lignin has been seen as a major advance in understanding how fungi degrade lignin. Recently, the fungus Trametes versicolor was shown to be capable of substantial decolorization and delignification of unbleached industrial kraft pulps over 2 to 5 days. The role, if any, of lignin peroxidase in this biobleaching was therefore examined. Several different assays indicated that T. versicolor can produce and secrete peroxidase proteins, but only under certain culture conditions. However, work employing a new lignin peroxidase inhibitor (metavanadate ions) and a new lignin peroxidase assay using the dye azure B indicated that secreted lignin peroxidases do not play a role in the T. versicolor pulp-bleaching system. Oxidative activity capable of degrading 2-keto-4-methiolbutyric acid (KMB) appeared unique to ligninolytic fungi and always accompanied pulp biobleaching.     

A protein-free medium for the growth of hybridomas and other cells of the immune system

Summary A protein-free medium, termed ABC, has been developed which essentially eliminates the need for serum proteins. ABC supports the long-term growth of murine hybridomas as well as other transformed cells of the immune system. The requirement of hybridoma growth for transferrin has been met by substituting the soluble organo-iron compound, sodium nitroprusside. Substantial improvement in the growth of hybridomas was afforded by the inclusion of 18 trace elements complexed to disodium ethylene diaminetetraacetate (EDTA). The medium was further improved by the inclusion of components not found in Ham's F12 medium or by raising the concentrations of existing low molecular weight components. Murine hybridomas can be cultured routinely in this protein-free medium in an anchorage-independent manner with doubling times generally under 24 h. Visualized on electrophoretic gels, levels of monoclonal antibody taken from those cultures often exceeded 80% of the total protein. The medium was also able to support the growth of HuT 78 and H9 cells as well as certain other transformed cells of the immune system. In addition, normal human peripheral blood lymphocytes, activated with phytohemagglutinin and cultured with 50 U/ml recombinant interleukin 2, could be grown for 2 wk with a 50-fold expansion over input cell number.     

Sperm motility initiation factor is a minor component of the Pacific herring egg chorion

Polyclonal antibodies were generated to the 105 kDa herring sperm motility initiation factor (SMIF) and used to explore the role of SMIF in sperm-egg interaction. Using sodium dodecyl sulfate polyacrylamide gel electrophoresis and immunoblotting with SMIF antibodies, it was demonstrated that SMIF is present as a minor (4–7% of total chorion protein) component of the chorion. The major polypeptides in the chorion migrated at 117 kDa and in a grouping between 48–54 kDa, with other minor bands above and below. The only detectable glycosylated component was the 105 kDa band, which was resolved at two isoelectric points (8.22 and 8.31) after isoelectric focusing gel electrophoresis. Using antibodies to SMIF, fertilization was blocked, sperm motility was inhibited in vitro in the presence of solubilized SMIF and SMIF binding sites on sperm were localized. Lastly, SMIF was localized to the region of the herring egg that encircles the micropyle.     

Behavioral changes in pottos (Perodicticus potto): Effects of naturalizing an exhibit

Observations of the potto (Perodicticus potto),a nocturnal prosimian primate,indicated a limited behavioral repertoire and prompted renovation of their exhibit at the Frankin Park Zoo (Boston, MA). We used the natural history of this species to direct the exhibit modifications, which used only nonsynthetic items. We added live plants, soil, bamboo, shelf fungi, grapevines, a hollow tree, and a log containing an insect dispenser to the exhibit at little expense. They provided new textures, odors, pathways, stimulation and cover for this secretive species. The changes produced a heightened aesthetic appeal to the nocturnal exhibit and improved its educational value by representing the animals’ habitat more accurately. The exhibit modifications also resulted in a significant increase in activity, an expansion of the pottos’ behavioral repertoire,the emergence of sexual behaviors, and an increased visibility to the public.     

Using XAD resins to study the effects of reduced organic micropollutant concentrations in River Rhine and Meuse water on phytoplankton growth

Laboratory incubation experiments were used to study the effect of reduced concentrations of organic micropollutants in water from the rivers Rhine and Meuse on the specific growth rate of the river phytoplankton community. Before incubation, part of the water sampled was treated with XAD-4 and XAD-8 resins to absorb dissolved organic compounds. Four dilutions were made by mixing untreated water with XAD-treated water in the ratios 100:0 (control), 70:30, 40:60 and 0:100. The phytoplankton specific growth rate increased significantly with the increased fraction treated with XAD in all but one incubation experiment. In these experiments, the specific growth rate was on average 9% higher in the fraction in which 100% was treated with XAD than in the controls. In the Rhine and Meuse river water, phytoplankton growth seemed to be inhibited by organic compounds. This inhibition was ascribed to the presence of dissolved organic micropollutants. Removing organic micropollutants using XAD resins to study the toxic effects of these compounds on field phytoplankton communities can be concluded to be a promising tool for risk assessment of micropollutants but needs to be supported by additional methodological research.     

Functional response of Neoseiulus barkeri Hughes on two-spotted spider mite (Acari: Tetranychidae)

Generalist predatory mites are the common phytoseiid fauna in many agroecosystems, but little attention has been paid to their potential as biological control agents. In this study, we determined the functional responses of adult females of the generalist predator Neoseiulus barkeri Hughes on eggs, larvae, and adults of the two-spotted spider mite, Tetranychus urticae Koch, in the laboratory. Predation experiments were conducted on pepper leaf discs over a 24 h period at 25±1°C, 70–80% RH and 16L:8D photoperiod. Prey densities ranged 5 to 80 eggs, or 5 to 40 larvae, or 1 to 8 female adults of T. urticae per disc. The predation rate of N. barkeri adult females on T. urticae eggs was the same as on its larvae, but the predation rate on adult females was much lower. The role of generalist predatory mites in integrated and biological control of greenhouse pests was discussed.     

Kinetics of ligand binding to a cluster of membrane-associated receptors

The process of ligand binding to a cluster of membrane-associated receptors is examined theoretically. The theoretical model proposed involves the diffusion of ligands from the solution to the disc-like cluster of receptors on the surface of the spherical cell. When the ligand hits the internal part of the disc-like cluster, it begins to move laterally until it leaves the disc through its outer surface or is bound by one of the receptors inside the disc. If the ligand leaves the cluster, it returns to the solution and hits the disc again after a certain period, etc. According to our model the transition from a diffusion-limited to a reaction-limited process of binding is determined by the dimensionless parameter Dt _c/a ², where D is the lateral diffusion coefficient,t _c is the characteristic time of reaction, anda is the radius of the disc-like cluster. The forward rate constantk _f turns out to be a function of . Comparing the results of our calculations ofk _f with some experimental data we found that agreement is achieved at high , i.e. the process of ligand binding by clustered receptors is predominantly reaction-limited.     

USE OF RIBOSOMAL DNA INTERNAL TRANSCRIBED SPACERS FOR PHYLOGENETIC STUDIES IN DIATOMS1

Sequence variation of ribosomal DNA internal transcribed spacers (ITS) among populations, species, and genera of the diatom genus Stephanodiscus was investigated. ITS 1 and ITS 2, including the 5.8S gene, were sequenced from geographically distant and nearby populations of S. niagarae Ehrenberg. In addition, repeats from S. hantzschii Grunow and Cyclotella meneghiniana Kützing were sequenced to determine the taxonomic range over which the ITS region could be used for diatom systematics. The morphologically distinct S. yellowstonensis Theriot & Stoermer, thought to have evolved from S. niagarae in Yellowstone Lake between 12,000 and 8000 years ago, also was sequenced to assess its relationship to nearby S. niagarae populations. The organization and relative sizes of ITS 1 and ITS 2 in Stephanodiscus species were similar to those reported for other eukaryotes. In general, ITS 2 was slightly larger and more variable than ITS 1. Cladistic analysis of ITS sequences did not resolve relationships of nearby S. niagarae and S. yellowstonensis populations. However, central North American S. niagarae populations were in a clade supported by two nucleotide changes. For Cyclotella, much of the ITS region was not alignable with that for Stephanodiscus species; therefore, generic-level comparison within the Thalassiosiraceae may not be possible. The variation (95–96% similarity) between S. hantzschii and other Stephanodiscus species suggests that interspecific relationships could be assessed with ITS sequences. Although S. yellowstonensis is morphologically distinct from S. niagarae, no autapomorphic nucleotide sites were identified. Two S. niagarae populations (Heart and Lewis Lakes), however, did possess autapomorphic ITS sites.