Nicotinic postsynaptic membranes from Torpedo: sidedness, permeability to macromolecules, and topography of major polypeptides

Experiments were conducted to examine the topographic arrangement of the polypeptides of the acetylcholine receptor (AcChR) and the nonreceptor Mr 43,000 protein in postsynaptic membranes isolated from Torpedo electric organ. When examined by electron microscopy, greater than 85% of vesicles were not permeable to ferritin or lactoperoxidase (LPO). Exposure to saponin was identified as a suitable procedure to permeabilize the vesicles to macromolecules with minimal alteration of vesicle size or ultrastructure. The sidedness of vesicles was examined morphologically and biochemically. Comparison of the distribution of intramembrane particles on freeze-fractured vesicles and the distribution found in situ indicated that greater than 85% of the vesicles were extracellular-side out. Vesicles labeled with alpha- bungarotoxin (alpha-Bgtx) were reacted with antibodies against alpha- BgTx or against purified AcChR of Torpedo. Bound antibodies were detected by the use of ferritin-conjugated goat anti-rabbit antibody and were located on the outside of greater than 99% of labeled vesicles. Similar results were obtained for normal vesicles or vesicles exposed to saponin. Quantification of the amount of [3H]-alpha-BgTx bound to vesicles before and after they were made permeable with saponin indicated that less than 5% of alpha-BgTx binding sites were cryptic in normal vesicles. It was concluded that greater than 95% of postsynaptic membranes were oriented extracellular-side out. LPO- catalyzed radioiodinations were performed on normal and saponin-treated vesicles and on vesicles from which the Mr (relative molecular mass) 43,000 protein had been removed by alkaline extraction. In normal vesicles, polypeptides of the AcChR were iodinated while the Mr 43,000 protein was not. In vesicles made permeable with saponin, the pattern of labeling of AcChR polypeptides was unchanged, but the Mr 43,000 protein was heavily iodinated. The relative iodination of AcChR polypeptides was unchanged in membranes equilibrated with agonist or with alpha-BgTx or after alkaline-extraction. It was concluded that the Mr 43,000 protein is present on the intracellular surface of the postsynaptic membrane and that AcChR polypeptides are exposed on the extracellular surface.     

Compaction and particle segregation in myelin membrane arrays

Compacted membrane arrays are formed in the nerve myelin sheath by lowering the water activity (through evaporation or immersion in hypertonic solutions of nonelectrolytes or monovalent salts) or by binding specific cations (Ca(++), La(+++), and tetracaine at concentrations above 5-10 mM). X-ray diffraction observations on intact, hydrated nerves treated to induce compaction provide a control to assess the significance of structural changes seen by freeze-fracture electron microscopy. Compaction inevitably leads to lateral segregation of particles away from the closely packed membrane arrays into contiguous normal, or slightly expanded, period arrays. In the particle-enriched layers, the E fracture face is more particle-dense than the P face, whereas no particles are found on either face in the compacted layers. Morphologically, compaction induced by the all-or-nothing, relatively irreversible action of specific cations cannot be distinguished from compaction to the same extent induced by the graded, reversible effects of nonelectrolytes. Compaction by sodium chloride resembles that by specific- cation binding in that the repeat period is independent of reagent concentration; but, like dehydration by nonelectrolytes, the extent of compaction is reversibly related to reagent concentration. Sodium chloride-compacted myelin can be distinguished morphologically by a lack of the elongated border particles at the boundary between smooth and particle-enriched membrane observed for other compacting treatments. Fracture faces in compacted arrays are not always smooth, but the unusual appearances can be duplicated in purified myelin lipid multilayers subjected to similar treatments, which indicates that the particle-free membrane fracture faces are uninterrupted lipid hydrocarbon layers. Correlation of x-ray diffraction and electron microscopy observations provides a direct basis for identifying the intramembrane particles with transmembrane protein. The transmembrane protein appears to play a significant role in maintaining the normal membrane separation; swelling of the particle-enriched arrays in myelin compacted by tetracaine at low ionic strength provides information about the charge distribution on the transmembrane protein. Swelling of the compacted arrays following irreversible particle segregation shows that the interaction properties of the particle-free membranes are similar to those of pure lipid multilayers. Compaction and the consequent particle segregation in lyelin results from conditions stabilizing close apposition of the lipid bilayers. Particle segregation in areas of close contact between other cell membranes may also be driven by interbilayer attractive forces.     

Monocaryotization of Cultures of Lenzites trabea (Pers.) Fr. and other Wood-destroying Basidiomycetes

Cultures of Lenzites trabea and some other basidiomycetes grownon nutrient agar containing high concentrations of sodium arsenateconsistently reverted from the dicaryotic to the monocaryoticcondition. This effect could also be induced by a wide varietyof other toxicants, including copper sulphate, zinc sulphate,sodium dichromate, sodium pentachlorophenate, creosote, crystalviolet, boric acid, and sodium taurocholate, but sodium arsenatewas the most reliable monocaryotizing agent, being effectivewith eight of the fourteen basidio-mycete species tested withit. The neohaplonts formed by sodium arsenate and other toxicantsmate normally with each other and with compatible monosporeisolates. The value of chemical monocaryotization as a techniquein fungal genetics and experimental taxonomy is discussed. Byusing the mating factor alleles as genetic markers, it was demonstratedthat when L. trabea is monocaryo-tized by sodium arsenate orcopper sulphate, only one of the two nuclei in the dicaryoncan be recovered in the neohaplonts, the direction of this completenuclear selection depending on the toxicant and the dicaryonconcerned. It was shown to occur in treated wood blocks duringroutine soil-block decay tests of inorganic wood preservativesand it is probably a frequent, though hitherto unrecognized,occurrence in the laboratory testing of wood preservatives andother fungicides against basidiomycetes.     

Genetic relationships among populations of Aedes aegypti from Uruguay and northeastern Argentina inferred from ISSR‐PCR data

Aedes aegypti (L.) (Diptera: Culicidae), the main vector of yellow fever and dengue viruses, was eradicated from Argentina between 1955 and 1963, but reinvaded the country in 1986. In Uruguay, the species was reintroduced in 1997. In this study we used highly polymorphic inter‐simple sequence repeats (ISSR) markers to analyse the genetic structure of Ae. aegypti populations from Uruguay and northeastern Argentina to identify possible colonization patterns of the vector. Overall genetic differentiation among populations was high (F_ST = 0.106) and showed no correlation with geographic distance, which is consistent with the short time since the reintroduction of the species in the area. Differentiation between pairs of Argentine populations (F_ST 0.072 to 0.221) was on average higher than between Uruguayan populations (F_ST?0.044 to 0.116). Bayesian estimation of population structure defined four genetic clusters and most populations were admixtures of two of them: Mercedes and Treinta y Tres (Uruguay) were mixtures of clusters 1 and 3; Salto (Uruguay) and Paraná (Argentina) of clusters 1 and 4; Fray Bentos (Uruguay) of clusters 2 and 3, and Gualeguaychú (Argentina) of clusters 2 and 3. Posadas and Buenos Aires in Argentina were fairly genetically homogeneous. Our results suggest that Ae. aegypti recolonized Uruguay from bordering cities in Argentina via bridges over the Uruguay River and also from Brazil.     

The considerable adult size variability in wood feeders is optimal

1. The life history of wood feeders was modelled in order to explain the multiseasonality of development and the great variability of adult size in this group. 2. The model was parameterised with experimental bioenergetic and reproductive data for the xylem feeder Aredolpona rubra (Coleoptera: Cerambycidae). 3. The length of the developmental period, which together with food quality directly determines adult size and indirectly determines the number of eggs laid, was optimised. 4. The results show that multi‐seasonal larval development maximises fitness under conditions of low food quality, relatively low predation pressure, and the presence of hostile periods during the year. 5. The variability of the number of seasons needed to complete development within a wood‐feeder population is a consequence of development time optimisation and the unavoidable extension of the egg‐laying period. These insects have an evolutionary dilemma: to eclose late in a given season at smaller size, bringing about low egg production and low offspring value, or to grow bigger to the next season, jeopardising their lives. 6. The results of the model predict wood‐feeder developmental patterns that depend on the tree tissue inhabited.     

MYGALOMORPH SPIDERS (ARANEAE: DIPLURIDAE) FROM THE LOWER CRETACEOUS CRATO LAGERSTÄTTE, ARARIPE BASIN, NORTH-EAST BRAZIL

Abstract:  The first mygalomorph spiders from the Lower Cretaceous Crato Lagerstätte of Cearà Province, north-east Brazil, are described, from adult males and females, in two new genera and species: Cretadiplura ceara Selden, gen. et sp. nov. and Dinodiplura ambulacra Selden, gen. et sp. nov. They belong to the extant family Dipluridae, hitherto known as fossils only from Tertiary strata; thus this occurrence extends the family record by some 90 myr.     

Effects of MEK inhibitor U0126 on meiotic progression in mouse oocytes： microtuble organization, asymmetric division and metaphase Ⅱ arrest

In this study we used U0126, a potent and specific inhibitor of MEK, to study the roles of MEK/ERK/p90~(rsk) signaling pathway in the meiotic cell cycle of mouse oocytes. The phosphorylation of MAP kinase and p90~(rsk) in the oocytes treated with 1.5 μM U0126 was the same as that in oocytes cultured in drug-free medium. With 1.5 μM U0126 treatment, the spindles appeared normal as they formed in oocytes, but failed to maintain its structure. Instead, the spindle lost one pole or elongated extraordinarily. After further culture, some oocytes extruded gigantic polar bodies (>30 μm) that later divided into two small ones. Some oocytes underwent symmetric division and produced two equal-size daughter cells in which normal spindles formed. In oocytes with different division patterns, MAP kinase was normally phosphorylated. When the concentration of U0126 was increased to 15 mM, the phosphorylation of both MAPK and p90~(rsk) were inhibited, while symmetric division was decreased. When incubating in medium c