Markers for trans-Golgi Membranes and the Intermediate Compartment Localize to Induced Membranes with Distinct Replication Functions in Flavivirus-Infected Cells

Replication of the flavivirus Kunjin virus is associated with virus-induced membrane structures within the cytoplasm of infected cells; these membranes appear as packets of vesicles associated with the sites of viral RNA synthesis and as convoluted membranes (CM) and paracrystalline arrays (PC) containing the components of the virus-specified protease (E. G. Westaway, J. M. Mackenzie, M. T. Kenney, M. K. Jones, and A. A. Khromykh, J. Virol. 71:6650-6661, 1997). To determine the cellular origins of these membrane structures, we compared the immunolabelling patterns of several cell markers in relation to these sites by immunofluorescence and immunoelectron microscopy. A marker for the trans-Golgi membranes and the trans-Golgi network, 1,4-galactosyltransferase (GalT), was redistributed to large foci in the cytoplasm of Kunjin virus-infected cells, partially coincident with immunofluorescent foci associated with the putative sites of viral RNA synthesis. As determined by immunoelectron microscopy, the induced vesicle packets contained GalT, whereas the CM and PC contained a specific protein marker for the intermediate compartment (ERGIC53). A further indicator of the role of cellular organelles in their biogenesis was the observation that the Golgi apparatus-disrupting agent brefeldin A prevented further development of immunofluorescent foci of induced membranes if added before the end of the latent period but that once formed, these membrane foci were resistant to brefeldin A dispersion. Reticulum membranes emanating from the induced CM and PC were also labelled with the rough endoplasmic reticulum marker anti-protein disulfide isomerase and were obviously redistributed during infection. This is the first report identifying trans-Golgi membranes and the intermediate compartment as the apparent sources of the flavivirus-induced membranes involved in events of replication.     

Im-trityl protection of histidine.

A rational attempt to prepare FmocHis(piTrt)OH regiospecifically gave in fact the well-known tau-trityl isomer, and experiments with model systems indicate that the prospects for access to pi-trityl histidine derivatives, which would be of great value for the racemization-free synthesis of histidine-containing peptides, are poor.     

What Do We Know About Metal Recycling Rates?

The recycling of metals is widely viewed as a fruitful sustainability strategy, but little information is available on the degree to which recycling is actually taking place. This article provides an overview on the current knowledge of recycling rates for 60 metals. We propose various recycling metrics, discuss relevant aspects of recycling processes, and present current estimates on global end‐of‐life recycling rates (EOL‐RR; i.e., the percentage of a metal in discards that is actually recycled), recycled content (RC), and old scrap ratios (OSRs; i.e., the share of old scrap in the total scrap flow). Because of increases in metal use over time and long metal in‐use lifetimes, many RC values are low and will remain so for the foreseeable future. Because of relatively low efficiencies in the collection and processing of most discarded products, inherent limitations in recycling processes, and the fact that primary material is often relatively abundant and low‐cost (which thereby keeps down the price of scrap), many EOL‐RRs are very low: Only for 18 metals (silver, aluminum, gold, cobalt, chromium, copper, iron, manganese, niobium, nickel, lead, palladium, platinum, rhenium, rhodium, tin, titanium, and zinc) is the EOL‐RR above 50% at present. Only for niobium, lead, and ruthenium is the RC above 50%, although 16 metals are in the 25% to 50% range. Thirteen metals have an OSR greater than 50%. These estimates may be used in considerations of whether recycling efficiencies can be improved; which metric could best encourage improved effectiveness in recycling; and an improved understanding of the dependence of recycling on economics, technology, and other factors.     

Changes in the levels of wheat- and barley-germ agglutinin during embryogenesis in vivo,in vitro and during germination

Radioimmunoassay has been used to measure levels of wheat-germ agglutinin and barley-germ agglutinin during embryogenesis and germination. The two lectins exhibited similar patterns of accumulation during grain maturation in vivo and both decreased to low levels after imbibition of harvest-ripe grains for 3 d. Precocious germination of immature wheat and barley embryos excised and cultured in vitro could be prevented either by inclusion of abscisic acid or mannitol in the culture medium. Changes in the level of wheat-germ agglutinin induced by in vitro culture depended on the maturation stage of the embryo. No direct correlation was found between application of exogenous abscisic acid and accumulation of the lectin.     

Collagen biosynthesis by human skin fibroblasts III. The effects of ascorbic acid on procollagen production and prolyl hydroxylase activity

Human skin fibroblasts were cultured under conditions optimized for collagen synthesis, and the effects of ascorbic acid on procollagen production, proline hydroxylation and the activity of prolyl hydroxylase were examined in cultures. The results indicated that addition of ascorbic acid to confluent monolayer cultures of adult human skin fibroblasts markedly increased tha amount of [³H]hydroxyproline syntehsized. Ascorbic acid, however, did not increase the synthesis of ³H-labeled collagenous polypeptides assayed independently of hydroxylation of proline residues, nor did it affect the amount of prolyl hydroxylase detectable by an in vitro enzyme assay. Also long-term cultures of the cells or initiation of fibroblast cultures in the presence of ascorbic acid did not lead to an apparent selection of a cell population which might be abnormally responsive to ascorbic acid. Thus, ascorbic acid appears to have one primary action on the synthesis of procollagen by cultured human skin fibroblasts: it is necessary for synthesis of hydroxyproline, and consequently for proper triple helix formation and selection of procollagen.     

Construction of improved bacteriophage phi 105 vectors for cloning by transfection in Bacillus subtilis

A series of improved phage vectors have been constructed, based on Bacillus subtilis bacteriophage phi 105, which can be used to clone genes in B. subtilis by direct transfection of protoplasts. The new vectors, designated phi 105J23, phi 105J24, phi 105J27 and phi 105J28, show frequencies of plaque formation that are equal to those of wild-type phi 105. This represents at least a 10-fold improvement over phi 105J9, the vector used in previous cloning experiments. Two of the new vectors phi 105J27 and phi 105J28 incorporate a mutation, cts-52, that renders the prophage temperature inducible. This has made it possible to devise a rapid small-scale procedure for screening progeny phage for the presence of inserted DNA. The usefulness of the new vectors is illustrated in the accompanying paper by cloning more than 20 B. subtilis sporulation genes.